What's hot today
Sunday, November 30th, 2014
Rodrigues-Campos, M. and Thompson, B. J. (2014). The ubiquitin ligase FbxL7 regulates the Dachsous-Fat-Dachs system in Drosophila. Development 141: 4098-4103. PubMed ID: 25256343
The atypical cadherins Dachsous (Ds) and Fat (Ft) are required to control the size and shape of tissues and organs in animals. In Drosophila, a key effector of Ds and Ft is the atypical myosin Dachs, which becomes planar polarised along the proximal-distal axis in developing epithelia to regulate tissue size via the Hippo pathway and tissue shape via modulating tension at junctions. How Ds and Ft control Dachs polarisation remains unclear. This study identified a ubiquitin ligase, FbxL7, as a novel component of the Ds-Ft-Dachs system that is required to control the level and localisation of Dachs. Loss of FbxL7 results in accumulation of Dachs, similar to loss of Ft. Overexpression of FbxL7 causes downregulation of Dachs, similar to overexpression of the Ft intracellular domain. In addition to regulating Dachs, FbxL7 also influences Ds in a similar manner. GFP-tagged FbxL7 localises to the plasma membrane in a Ft-dependent manner and is planar polarised. It is proposed that Ft recruits FbxL7 to the proximal side of the cell to help restrict Ds and Dachs to the distal side of the cell.
Abeysundara, N., Leung, A. C., Primrose, D. A. and Hughes, S. C. (2014). Regulation of cell proliferation and adhesion by means of a novel region of Drosophila Merlin interacting with Sip1. Dev Dyn 243: 1554-1570. PubMed ID: 25403428
The tumor suppressor protein Merlin is thought to regulate cell proliferation and cell adhesion through interaction with protein partners. Loss of merlin is associated with Neurofibromatosis Type 2 (NF2) tumors. NHERF1 or EBP50 is a scaffolding protein that functions in apical organization of polarized cells. Merlin and NHERF1 have been shown to interact in vitro in vertebrates. This study investigated how the Drosophila NHERF1 orthologue, Sip1, and Merlin function to regulate cell proliferation and adhesion. Two conserved arginine residues (R325 and R335) were identified in Merlin that addition to the FERM domain, are required for interaction with Sip1. Mutation of the arginine residues result in reduced Sip1 binding to Merlin and loss of Merlin growth suppressor function. Over-expression of Merlin(R325A) and/or Merlin(R335L) in Drosophila wings result in increased proliferation in the adult wing (increase in size), which is rescued by co-over-expression of constitutively active Merlin protein. Reduced Sip1 binding to Merlin also produces defects in adhesion in follicle epithelial cells. It is concluded that Sip1 facilitates the activation of Merlin as a tumor suppressor protein. Thus, this work provides insight into how Merlin functions as a tumor suppressor and in adhesion and this provides insight into the mechanism of NF2 pathogenesis.
Dejima, K., Kang, S., Mitani, S., Cosman, P. C. and Chisholm, A. D. (2014). Syndecan defines precise spindle orientation by modulating Wnt signaling in C. elegans. Development 141: 4354-4365. PubMed ID: 25344071
Wnt signals orient mitotic spindles in development, but it remains unclear how Wnt signaling is spatially controlled to achieve precise spindle orientation. This study shows that C. elegans syndecan (SDN-1; see Drosophila Syndecan) is required for precise orientation of a mitotic spindle in response to a Wnt cue. SDN-1 is the predominant heparan sulfate (HS) proteoglycan in the early C. elegans embryo, and that loss of HS biosynthesis or of the SDN-1 core protein results in misorientation of the spindle of the ABar blastomere. The ABar and EMS spindles both reorient in response to Wnt signals, but only ABar spindle reorientation is dependent on a new cell contact and on HS and SDN-1. SDN-1 transiently accumulates on the ABar surface as it contacts C, and is required for local concentration of Dishevelled (MIG-5; see Drosophila Dishevelled) in the ABar cortex adjacent to C. These findings establish a new role for syndecan in Wnt-dependent spindle orientation.
Castel, S. E., Ren, J., Bhattacharjee, S., Chang, A. Y., Sanchez, M., Valbuena, A., Antequera, F. and Martienssen, R. A. (2014). Dicer promotes transcription termination at sites of replication stress to maintain genome stability. Cell 159: 572-583. PubMed ID: 25417108
Nuclear RNAi is an important regulator of transcription and epigenetic modification, but the underlying mechanisms remain elusive. Using a genome-wide approach in the fission yeast S. pombe, this study found that Dcr1 (see Drosophila Dicer-1), but not other components of the canonical RNAi pathway, promotes the release of Pol II (see Drosophila Pol2) from the 3' end of highly transcribed genes, and, surprisingly, from antisense transcription of rRNA and tRNA genes, which are normally transcribed by Pol I and Pol III. These Dcr1-terminated loci correspond to sites of replication stress and DNA damage, likely resulting from transcription-replication collisions. At the rDNA loci, release of Pol II facilitates DNA replication and prevents homologous recombination, which would otherwise lead to loss of rDNA repeats especially during meiosis. These results reveal a novel role for Dcr1-mediated transcription termination in genome maintenance and may account for widespread regulation of genome stability by nuclear RNAi in higher eukaryotes.
Saturday, November 29th
Telonis-Scott, M., Clemson, A. S., Johnson, T. K. and Sgro, C. M. (2014). Spatial analysis of gene regulation reveals new insights into the molecular basis of upper thermal limits. Mol Ecol [Epub ahead of print]. PubMed ID: 25401770
The cellular stress response has long been the primary model for studying the molecular basis of thermal adaptation, yet the link between gene expression, RNA metabolism and physiological responses to thermal stress remains largely unexplored. This was addressed by comparing the transcriptional and physiological responses of three geographically distinct populations of D. melanogaster from eastern Australia in response to, and recovery from, a severe heat stress with and without a pre-stress hardening treatment. Focus was placed on starvin (stv), recently identified as an important thermally responsive gene. Intriguingly, stv encodes seven transcripts from alternative transcription sites and alternative splicing, yet appears to be rapidly heat-inducible. First, genetic differences were shown in upper thermal limits of the populations tested. It was then demonstrated that the stv locus does not ubiquitously respond to thermal stress but is expressed as three distinct thermal and temporal RNA phenotypes (isoforms). The shorter transcript isoforms are rapidly up-regulated under stress in all populations and show similar molecular signatures to Heat shock proteins. Multiple stress exposures seem to generate a reserve of pre-mRNAs, effectively 'priming' the cells for subsequent stress. Remarkably, a bypass in the splicing blockade in these isoforms was demonstrated, suggesting an essential role for these transcripts under heat stress. Temporal profiles for the weakly heat responsive stv isoform subset show opposing patterns in the two most divergent populations. Innate and induced transcriptome responses to hyperthermia are complex, and warrant moving beyond gene-level analyses.
Bergland, A. O., Behrman, E. L., O'Brien, K. R., Schmidt, P. S. and Petrov, D. A. (2014). Genomic evidence of rapid and stable adaptive oscillations over seasonal time scales in Drosophila. PLoS Genet 10: e1004775. PubMed ID: 25375361
In many species, genomic data have revealed pervasive adaptive evolution indicated by the fixation of beneficial alleles. However, when selection pressures are highly variable along a species' range or through time adaptive alleles may persist at intermediate frequencies for long periods. So called 'balanced polymorphisms' have long been understood to be an important component of standing genetic variation, yet direct evidence of the strength of balancing selection and the stability and prevalence of balanced polymorphisms has remained elusive. It was hypothesized that environmental fluctuations among seasons in a North American orchard would impose temporally variable selection on Drosophila melanogaster that would drive repeatable adaptive oscillations at balanced polymorphisms. Hundreds of polymorphisms were identified whose frequency oscillates among seasons, and it was argued that these loci are subject to strong, temporally variable selection. This study shows that these polymorphisms respond to acute and persistent changes in climate and are associated in predictable ways with seasonally variable phenotypes. In addition, the results suggest that adaptively oscillating polymorphisms are likely millions of years old, with some possibly predating the divergence between D. melanogaster and D. simulans. Taken together, these results are consistent with a model of balancing selection wherein rapid temporal fluctuations in climate over generational time promotes adaptive genetic diversity at loci underlying polygenic variation in fitness related phenotypes.
sponses to hyperthermia are complex, and warrant moving beyond gene-level analyses.
Yang, H., He, B., Ma, H., Tsaur, S. C., Ma, C., Wu, Y., Ting, C. T. and Zhang, Y. E. (2014). Expression profile and gene age jointly shaped the genome-wide distribution of premature termination codons in a Drosophila melanogaster population. Mol Biol Evol . [Epub ahead of print]. PubMed ID: 25371429
Widespread premature termination codon mutations (PTCs) were recently observed in human and fly populations. This study took advantage of the population resequencing data in the Drosophila Genetic Reference Panel (DGRP) to investigate how the expression profile and the evolutionary age of genes shaped the allele frequency distribution of PTCs. After generating a high-quality dataset of PTCs, genes harboring PTCs were clustered into three categories: genes encoding low-frequency PTCs (</= 1.5%), moderate-frequency PTCs (1.5%-10%) and high-frequency PTCs (> 10%). All three groups show narrow transcription compared to PTC-free genes, with the moderate- and high-PTC frequency groups showing a pronounced pattern. Moreover, nearly half (42%) of the PTC-encoding genes are not expressed in any tissue. Interestingly, the moderate-frequency PTC group is strongly enriched for genes expressed in midgut, whereas genes harboring high-frequency PTCs tend to have sex-specific expression. It was further found that although young genes born in the last 60 million years (Myr) compose a mere 9% of the genome, they represent 16%, 30% and 50% of the genes containing low-, moderate- and high-frequency PTCs, respectively. Among DNA-based and RNA-based duplicated genes, the child copy is approximately twice as likely to contain PTCs as the parent copy, whereas young de novo genes are as likely to encode PTCs as DNA-based duplicated new genes. Based on these results, it is concluded that expression profile and gene age jointly shaped the landscape of PTC-mediated gene loss. Therefore, it is proposed that new genes may need a long time to become stably maintained after the origination.
Innocenti, P., Flis, I. and Morrow, E. H. (2014). Female responses to experimental removal of sexual selection components in Drosophila melanogaster. BMC Evol Biol 14: 239. PubMed ID: 25406540
Despite the common assumption that multiple mating should in general be favored in males, but not in females, to date there is no consensus on the general impact of multiple mating on female fitness. Notably, very little is known about the genetic and physiological features underlying the female response to sexual selection pressures. By combining an experimental evolution approach with genomic techniques, this study investigated the effects of single and multiple matings on female fecundity and gene expression. The opportunity for mating in replicate populations of Drosophila melanogaster was experimentally manipulated by removing components of sexual selection, with the aim of testing differences in short term post-mating effects of females evolved under different mating strategies. It was shown that monogamous females suffer decreased fecundity, a decrease that was partially recovered by experimentally reversing the selection pressure back to the ancestral state. The post-mating gene expression profiles of monogamous females differ significantly from promiscuous females, involving 9% of the genes tested (approximately 6% of total genes in D. melanogaster). These transcripts are active in several tissues, mainly ovaries, neural tissues and midgut, and are involved in metabolic processes, reproduction and signaling pathways. These results demonstrate how the female post-mating response can evolve under different mating systems, and provide novel insights into the genes targeted by sexual selection in females, by identifying a list of candidate genes responsible for the decrease in female fecundity in the absence of promiscuity.
Chen, X., Bracht, J. R., Goldman, A. D., Dolzhenko, E., Clay, D. M., Swart, E. C., Perlman, D. H., Doak, T. G., Stuart, A., Amemiya, C. T., Sebra, R. P. and Landweber, L. F. (2014). The architecture of a scrambled genome reveals massive levels of genomic rearrangement during development.Cell 158: 1187-1198. PubMed ID: 25171416
Programmed DNA rearrangements in the single-celled eukaryote Oxytricha trifallax completely rewire its germline into a somatic nucleus during development. This elaborate, RNA-mediated pathway eliminates noncoding DNA sequences that interrupt gene loci and reorganizes the remaining fragments by inversions and permutations to produce functional genes. This study reports the Oxytricha germline genome and compares it to the somatic genome to present a global view of its massive scale of genome rearrangements. The remarkably encrypted genome architecture contains >3,500 scrambled genes, as well as >800 predicted germline-limited genes expressed, and some posttranslationally modified, during genome rearrangements. Gene segments for different somatic loci often interweave with each other. Single gene segments can contribute to multiple, distinct somatic loci. Terminal precursor segments from neighboring somatic loci map extremely close to each other, often overlapping. This genome assembly provides a draft of a scrambled genome and a powerful model for studies of genome rearrangement.
Friday, November 28th
Pezeron, G., Millen, K., Boukhatmi, H. and Bray, S. (2014). Notch directly regulates cell morphogenesis genes, Reck, talin and trio, in adult muscle progenitors. J Cell Sci. PubMed ID: 25217625
There is growing evidence that Notch pathway activation can result in consequences on cell morphogenesis and behaviour, both during embryonic development and cancer progression. In general, Notch is proposed to co-ordinate these processes by regulating expression of key transcription factors. However, many Notch-regulated genes identified in genome-wide studies are involved in fundamental aspects of cell behaviour, suggesting a more direct influence on cellular properties. By testing the functions of 25 such genes it was confirmed that 12 are required in developing adult muscles consistent with roles downstream of Notch. Focusing on three, Reck, rhea/talin and trio, their expression was varified in adult muscle progenitors, and Notch-regulated enhancers in each were identified. Full activity of these enhancers requires functional binding sites for Su(H), the DNA-binding transcription factor in the Notch pathway, validating their direct regulation. Thus, besides its well-known roles in regulating the expression of cell-fate determining transcription factors, Notch signalling also has the potential to directly affect cell morphology/behaviour by modulating expression of genes such as Reck, rhea/talin and trio. This sheds new light on functional outputs of Notch activation in morphogenetic processes.
Haltom, A. R., Lee, T. V., Harvey, B. M., Leonardi, J., Chen, Y. J., Hong, Y., Haltiwanger, R. S. and Jafar-Nejad, H. (2014). The protein O-glucosyltransferase Rumi modifies Eyes shut to promote rhabdomere separation in Drosophila. PLoS Genet 10: e1004795. PubMed ID: 25412384
The protein O-glucosyltransferase Rumi/POGLUT1 regulates Drosophila Notch signaling by adding O-glucose residues to the Notch extracellular domain. Rumi has other predicted targets including Crumbs (Crb) and Eyes shut (Eys), both of which are involved in photoreceptor development. However, whether Rumi is required for the function of Crb and Eys remains unknown. This study reports that in the absence of Rumi or its enzymatic activity, several rhabdomeres in each ommatidium fail to separate from one another in a Notch-independent manner. Mass spectral analysis indicates the presence of O-glucose on Crb and Eys. However, mutating all O-glucosylation sites in a crb knock-in allele does not cause rhabdomere attachment, ruling out Crb as a biologically-relevant Rumi target in this process. In contrast, eys and rumi exhibit a dosage-sensitive genetic interaction. In addition, although in wild-type ommatidia most of the Eys protein is found in the inter-rhabdomeral space (IRS), in rumi mutants a significant fraction of Eys remains in the photoreceptor cells. The intracellular accumulation of Eys and the IRS defect worsen in rumi mutants raised at a higher temperature, and are accompanied by a approximately 50% decrease in the total level of Eys. Moreover, removing one copy of an endoplasmic reticulum chaperone enhances the rhabdomere attachment in rumi mutant animals. Altogether, these data suggest that O-glucosylation of Eys by Rumi ensures rhabdomere separation by promoting proper Eys folding and stability in a critical time window during the mid-pupal stage. Human EYS, which is mutated in patients with autosomal recessive retinitis pigmentosa, also harbors multiple Rumi target sites. Therefore, the role of O-glucose in regulating Eys may be conserved.
Shen, J., Lu, J., Sui, L., Wang, D., Yin, M., Hoffmann, I., Legler, A. and Pflugfelder, G. O. (2014). The orthologous Tbx transcription factors Omb and TBX2 induce epithelial cell migration and extrusion in vivo without involvement of matrix metalloproteinases. Oncotarget [Epub ahead of print]. PubMed ID: 25344916
The transcription factors TBX2 and TBX3 are overexpressed in various human cancers. This study investigated the effect of overexpressing the orthologous Tbx genes Drosophila optomotor-blind (omb) and human TBX2 in the epithelium of the Drosophila wing imaginal disc; two types of cell motility were observed. Omb/TBX2 overexpressing cells could move within the plane of the epithelium. Invasive cells migrated long-distance as single cells retaining or regaining normal cell shape and apico-basal polarity in spite of attenuated apical DE-cadherin concentration. Inappropriate levels of DE-cadherin were sufficient to drive cell migration in the wing disc epithelium. Omb/TBX2 overexpression and reduced DE-cadherin-dependent adhesion caused the formation of actin-rich lateral cell protrusions. Omb/TBX2 overexpressing cells could also delaminate basally, penetrating the basal lamina, however, without degradation of extracellular matrix. Expression of Timp, an inhibitor of matrix metalloproteases, blocked neither intraepithelial motility nor basal extrusion. These results reveal an MMP-independent mechanism of cell invasion and suggest a conserved role of Tbx2-related proteins in cell invasion and metastasis-related processes.
Buisson, N., Sirour, C., Moreau, N., Denker, E., Le Bouffant, R., Goullancourt, A., Darribere, T. and Bello, V. (2014). An adhesome comprising laminin, dystroglycan and myosin IIA is required during notochord development in Xenopus laevis. Development 141: 4569-4579. PubMed ID: 25359726
Dystroglycan (Dg; see Drosophila Dystroglycan) is a transmembrane receptor for laminin that must be expressed at the right time and place in order to be involved in notochord morphogenesis. The function of Dg was examined in Xenopus laevis embryos by knockdown of Dg and overexpression and replacement of the endogenous Dg with a mutated form of the protein. This analysis revealed that Dg is required for correct laminin assembly, for cell polarization during mediolateral intercalation and for proper differentiation of vacuoles. Using mutations in the cytoplasmic domain, two sites were identified that are involved in cell polarization and are required for mediolateral cell intercalation, and a site that is required for vacuolation. Furthermore, using a proteomic analysis, the cytoskeletal non-muscle myosin IIA has been identified for the first time as a molecular link between the Dg-cytoplasmic domain and cortical actin. The data allowed identification of the adhesome laminin-Dg-myosin IIA as being required to maintain the cortical actin cytoskeleton network during vacuolation, which is crucial to maintain the shape of notochordal cells.
Thursday, November 27th
Lebreton, S., Grabe, V., Omondi, A. B., Ignell, R., Becher, P. G., Hansson, B. S., Sachse, S. and Witzgall, P. (2014). Love makes smell blind: mating suppresses pheromone attraction in Drosophila females via Or65a olfactory neurons. Sci Rep 4: 7119. PubMed ID: 25406576
In Drosophila, the male sex pheromone cis-vaccenyl acetate (cVA) elicits aggregation and courtship, through the odorant receptor Or67d. Long-lasting exposure to cVA suppresses male courtship, via a second channel, Or65a. In females, the role of Or65a has not been studied. This study shows that, shortly after mating, Drosophila females are no longer attracted to cVA and that activation of olfactory sensory neurons (OSNs) expressing Or65a generates this behavioral switch: when silencing Or65a, mated females remain responsive to cVA. Neurons expressing Or67d converge into the DA1 glomerulus in the antennal lobe, where they synapse onto projection neurons (PNs), that connect to higher neural circuits generating the attraction response to cVA. Functional imaging of these PNs shows that the DA1 glomerulus is inhibited by simultaneous activation of Or65a OSNs, which leads to a suppression of the attraction response to cVA. The behavioral role of postmating cVA exposure is substantiated by the observation that matings with starved males, which produce less cVA, do not alter the female response. Moreover, exposure to synthetic cVA abolishes attraction and decreases sexual receptivity in unmated females. Taken together, Or65a mediates an aversive effect of cVA and may accordingly regulate remating, through concurrent behavioral modulation in males and females.
Vrailas-Mortimer, A. D., Ryan, S. M., Avey, M. J., Mortimer, N. T., Dowse, H. and Sanyal, S. (2014). p38 MAP Kinase Regulates Circadian Rhythms in Drosophila. J Biol Rhythms [Epub ahead of print]. PubMed ID: 25403440
The large repertoire of circadian rhythms in diverse organisms depends on oscillating central clock genes, input pathways for entrainment, and output pathways for controlling rhythmic behaviors. Stress-activated p38 MAP Kinases (p38K), although sparsely investigated in this context, show circadian rhythmicity in mammalian brains and are considered part of the circadian output machinery in Neurospora. This study found that Drosophila p38Kb is expressed in clock neurons, and mutants in p38Kb either are arrhythmic or have a longer free-running periodicity, especially as they age. Paradoxically, similar phenotypes are observed through either transgenic inhibition or activation of p38Kb in clock neurons, suggesting a requirement for optimal p38Kb function for normal free-running circadian rhythms. This study also found that p38Kb genetically interacts with multiple downstream targets to regulate circadian locomotor rhythms. More specifically, p38Kb interacts with the period gene to regulate period length and the strength of rhythmicity. In addition, p38Kb was shown to suppress the arrhythmic behavior associated with inhibition of a second p38Kb target, the transcription factor Mef2. Finally, manipulating p38K signaling in free-running conditions was found to alter the expression of another downstream target, MNK/Lk6, which has been shown to cycle with the clock and to play a role in regulating circadian rhythms. These data suggest that p38Kb may affect circadian locomotor rhythms through the regulation of multiple downstream pathways.
Gou, B., Liu, Y., Guntur, A. R., Stern, U. and Yang, C. H. (2014). Mechanosensitive neurons on the internal reproductive tract contribute to egg-laying-induced acetic acid attraction in Drosophila. Cell Rep 9: 522-530. PubMed ID: 25373900
Selecting a suitable site to deposit their eggs is an important reproductive need of Drosophila females. Although their choosiness toward egg-laying sites is well documented, the specific neural mechanism that activates females' search for attractive egg-laying sites is not known. This study shows that distention and contraction of females' internal reproductive tract triggered by egg delivery through the tract plays a critical role in activating such search. Females start to exhibit acetic acid (AA) attraction prior to depositing each egg but no attraction when they are not laying eggs. Artificially distending the reproductive tract triggers AA attraction in non-egg-laying females, whereas silencing the mechanosensitive neurons this study identified that can sense the contractile status of the tract eliminates such attraction. This work uncovers the circuit basis of an important reproductive need of Drosophila females and provides a simple model for dissecting the neural mechanism that underlies a reproductive need-induced behavioral modification.
Chadha, A. and Cook, B. (2014). The effect of stress on motor function in Drosophila. PLoS One 9: e112076. PubMed ID: 25375106
Exposure to unpredictable and uncontrollable conditions causes animals to perceive stress and change their behavior. It is unclear how the perception of stress modifies the motor components of behavior and which molecular pathways affect the behavioral change. In order to understand how stress affects motor function, an experimental platform was developed that quantifies walking motions in Drosophila. Stress induction using electrical shock was found to result in backwards motions of the forelegs at the end of walking strides. These leg retrogressions persisted during repeated stimulation, although they habituated substantially. The motions also continued for several strides after the end of the shock, indicating that stress induces a behavioral aftereffect. Such aftereffect could also be induced by restricting the motion of the flies via wing suspension. Further, the long-term effects could be amplified by combining either immobilization or electric shock with additional stressors. Thus, retrogression is a lingering form of response to a broad range of stressful conditions, which cause the fly to search for a foothold when it faces extreme and unexpected challenges. Mutants in the cAMP signaling pathway enhanced the stress response, indicating that this pathway regulates the behavioral response to stress. These findings identify the effect of stress on a specific motor component of behavior and define the role of cAMP signaling in this stress response.
Geurten, B. R., Jahde, P., Corthals, K. and Gopfert, M. C. (2014). Saccadic body turns in walking Drosophila. Front Behav Neurosci 8: 365. PubMed ID: 25386124
Drosophila melanogaster structures its optic flow during flight by interspersing translational movements with abrupt body rotations. Whether these 'body saccades' are accompanied by steering movements of the head is a matter of debate. By tracking single flies moving freely in an arena, it was discovered that walking Drosophila also perform saccades. Movement analysis revealed that the flies separate rotational from translational movements by quickly turning their bodies by 15 degrees within a tenth of a second. Although walking flies moved their heads by up to 20 degrees about their bodies, their heads moved with the bodies during saccadic turns. This saccadic strategy contrasts with the head saccades reported for e.g., blowflies and honeybees, presumably reflecting optical constraints: modeling revealed that head saccades as described for these latter insects would hardly affect the retinal input in Drosophila because of the lower acuity of its compound eye. The absence of head saccades in Drosophila was associated with the absence of haltere oscillations, which seem to guide head movements in other flies. In addition to adding new twists to Drosophila walking behavior, this analysis shows that Drosophila does not turn its head relative to its body when turning during walking.
Wednesday, November 26th
Menuz, K., Larter, N. K., Park, J. and Carlson, J. R. (2014). An RNA-seq screen of the Drosophila antenna identifies a transporter necessary for ammonia detection. PLoS Genet 10: e1004810. PubMed ID: 25412082
Many insect vectors of disease detect their hosts through olfactory cues, and thus it is of great interest to understand better how odors are encoded. However, little is known about the molecular underpinnings that support the unique function of coeloconic sensilla, an ancient and conserved class of sensilla that detect amines and acids, including components of human odor that are cues for many insect vectors. This study generated antennal transcriptome databases both for wild type Drosophila and for a mutant that lacks coeloconic sensilla. These resources were used to identify genes whose expression is highly enriched in coeloconic sensilla, including many genes not previously implicated in olfaction. Among them, an ammonium transporter gene [CG6499, renamed Ammonium transporter (Amt)] was identified that is essential for ammonia responses in a class of coeloconic olfactory receptor neurons (ORNs), but is not required for responses to other odorants. Surprisingly, the transporter is not expressed in ORNs, but rather in neighboring auxiliary cells. Thus, these data reveal an unexpected non-cell autonomous role for a component that is essential to the olfactory response to ammonia. The defective response observed in a Drosophila mutant of this gene is rescued by its Anopheles ortholog, and orthologs are found in virtually all insect species examined, suggesting that its role is conserved. Taken together, these results provide a quantitative analysis of gene expression in the primary olfactory organ of Drosophila, identify molecular components of an ancient class of olfactory sensilla, and reveal that auxiliary cells, and not simply ORNs, play an essential role in the coding of an odor that is a critical host cue for many insect vectors of human disease.
Chen, H., Zheng, X. and Zheng, Y. (2014). Age-associated loss of lamin-B leads to systemic inflammation and gut hyperplasia. Cell 159: 829-843. PubMed ID: 25417159
Aging of immune organs, termed as immunosenescence, is suspected to promote systemic inflammation and age-associated disease. The cause of immunosenescence and how it promotes disease, however, has remained unclear. This study reports the Drosophila fat body, a major immune organ, undergoes immunosenescence and mounts strong systemic inflammation that leads to deregulation of immune deficiency (IMD) signaling in the midgut of old animals. Inflamed old fat bodies secrete circulating peptidoglycan recognition proteins that repress IMD activity in the midgut, thereby promoting gut hyperplasia. Further, fat body immunosenecence is caused by age-associated lamin-B reduction specifically in fat body cells, which then contributes to heterochromatin loss and derepression of genes involved in immune responses. As lamin-associated heterochromatin domains are enriched for genes involved in immune response in both Drosophila and mammalian cells, these findings may provide insights into the cause and consequence of immunosenescence during mammalian aging.
Kairamkonda, S. and Nongthomba, U. (2014). Beadex function in the motor neurons is essential for female reproduction in Drosophila melanogaster. PLoS One 9: e113003. PubMed ID: 25396431
Drosophila melanogaster has served as an excellent model system for understanding the neuronal circuits and molecular mechanisms regulating complex behaviors. The Drosophila female reproductive circuits, in particular, are well studied and can be used as a tool to understand the role of novel genes in neuronal function in general and female reproduction in particular. In the present study, the role of Beadex, a transcription co-activator, in Drosophila female reproduction was assessed by generation of mutant and knock down studies. Null allele of Beadex was generated by transposase induced excision of P-element present within an intron of Beadex gene. The mutant showed highly compromised reproductive abilities as evaluated by reduced fecundity and fertility, abnormal oviposition and more importantly, the failure of sperm release from storage organs. However, no defect was found in the overall ovariole development. Tissue specific, targeted knock down of Beadex indicated that its function in neurons is important for efficient female reproduction, since its neuronal knock down led to compromised female reproductive abilities, similar to Beadex null females. Further, different neuronal class specific knock down studies revealed that Beadex function is required in motor neurons for normal fecundity and fertility of females. Thus, the present study attributes a novel and essential role for Beadex in female reproduction through neurons.
Thimgan, M. S., Seugnet, L., Turk, J. and Shaw, P. J. (2014). Identification of genes associated with resilience/vulnerability to sleep deprivation and starvation in Drosophila. Sleep [Epub ahead of print]. PubMed ID: 25409104
Flies mutant for the canonical clock protein cycle (cyc01) exhibit a sleep rebound that is approximately 10 times larger than wild-type flies and die after only 10 h of sleep deprivation. Surprisingly, when starved, cyc01 mutants can remain awake for 28 h without demonstrating negative outcomes. Thus, it was hypothesized that identifying transcripts that are differentially regulated between waking induced by sleep deprivation and waking induced by starvation would identify genes that underlie the deleterious effects of sleep deprivation and/or protect flies from the negative consequences of waking. Partial complementary DNA microarrays to identify transcripts that are differentially expressed between cyc01 mutants that had been sleep deprived or starved for 7 h. Genetics was used to determine whether disrupting genes involved in lipid metabolism would exhibit alterations in their response to sleep deprivation. 84 genes were identified with transcript levels that were differentially modulated by 7 h of sleep deprivation and starvation in cyc01 mutants and were confirmed in independent samples using quantitative polymerase chain reaction. Several of these genes were predicted to be lipid metabolism genes, including bubblegum, cueball, and CG4500, which based on data obtained in this study was renamed heimdall (hll). Using lipidomics it was confirmed that knockdown of hll using RNA interference significantly decreased lipid stores. Importantly, genetically modifying bubblegum, cueball, or hll resulted in sleep rebound alterations following sleep deprivation compared to genetic background controls. This study has identified a set of genes that may confer resilience/vulnerability to sleep deprivation and demonstrate that genes involved in lipid metabolism modulate sleep homeostasis.
Cooper, B. S., Hammad, L. A. and Montooth, K. L. (2014). Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster. Funct Ecol 28: 886-894. PubMed ID: 25382893
Changes in temperature disrupt the fluidity of cellular membranes, which can negatively impact membrane integrity and cellular processes. Many ectotherms, including Drosophila melanogaster, adjust the glycerophospholipid composition of their membranes to restore optimal fluidity when temperatures change, a type of trait plasticity termed homeoviscous adaptation. Existing data suggest that plasticity in the relative abundances of the glycerophospholipids phosphatidylethanolamine (PE) and phosphatidylcholine (PC) underlies cellular adaptation to temporal variability in the thermal environment. For example, laboratory populations of D. melanogaster evolved in the presence of temporally variable temperatures have greater developmental plasticity of the ratio of PE to PC (PE/PC) and greater fecundity than do populations evolved at constant temperatures. This work has been extended to natural populations of D. melanogaster by evaluating thermal plasticity of glycerophospholipid composition at different life stages, in genotypes isolated from Vermont, Indiana and North Carolina. The covariance was quantified between developmental and adult (reversible) plasticity, and between adult responses of the membrane to cool and warm thermal shifts. As predicted by physiological models of homeoviscous adaptation, flies from all populations decrease PE/PC and the degree of lipid unsaturation in response to warm temperatures. Furthermore, these populations have diverged in their degree of membrane plasticity. Flies from the most variable thermal environment (Vermont) decrease PE/PC to a greater extent than do other populations when developed at a warm temperature, a pattern that matches previous observation in laboratory-evolved populations. It was also found that developmental plasticity and adult plasticity of PE/PC covary across genotypes, but that adult responses to cool and warm thermal shifts do not. When combined with previous observations of laboratory-evolved populations, these findings implicate developmental plasticity of PE/PC as a mechanism of thermal adaptation in temporally variable environments. While little is known about the genetic bases of plastic responses to temperature, these observations suggest that both environmentally sensitive and environmentally specific alleles contribute to thermal adaptation of membranes, and that costs of plasticity may arise when the adult environment differs from that experienced during development.
Tuesday, November 25th
Menon, D. U., Coarfa, C., Xiao, W., Gunaratne, P. H. and Meller, V. H. (2014). siRNAs from an X-linked satellite repeat promote X-chromosome recognition in Drosophila melanogaster. Proc Natl Acad Sci U S A. PubMed ID: 25368194
Highly differentiated sex chromosomes create a lethal imbalance in gene expression in one sex. To accommodate hemizygosity of the X chromosome in male fruit flies, expression of X-linked genes increases twofold. This is achieved by the male- specific lethal (MSL) complex, which modifies chromatin to increase expression. Mutations that disrupt the X localization of this complex decrease the expression of X-linked genes and reduce male survival. The mechanism that restricts the MSL complex to X chromatin is not understood. The siRNA pathway has been shown to contribute to localization of the MSL complex, raising questions about the source of the siRNAs involved. The X-linked 1.688 g/cm3 satellite related repeats (1.688X repeats; 359-bp repeat unit) are restricted to the X chromosome and produce small RNA, making them an attractive candidate. RNA from these repeats was tested for a role in dosage compensation, and ectopic expression of single-stranded RNAs from 1.688X repeats was found to enhance the male lethality of mutants with defective X recognition. In contrast, expression of double-stranded hairpin RNA from a 1.688X repeat generated abundant siRNA and dramatically increased male survival. Consistent with improved survival, X localization of the MSL complex was largely restored in these males. The striking distribution of 1.688X repeats, which are nearly exclusive to the X chromosome, suggests that these are cis-acting elements contributing to identification of X chromatin.
Ghosh, R., Vegesna, S., Safi, R., Bao, H., Zhang, B., Marenda, D. R. and Liebl, F. L. (2014). Kismet positively regulates glutamate receptor localization and synaptic transmission at the Drosophila neuromuscular junction. PLoS One 9: e113494. PubMed ID: 25412171
The Drosophila neuromuscular junction (NMJ) is a glutamatergic synapse that is structurally and functionally similar to mammalian glutamatergic synapses. These synapses can, as a result of changes in activity, alter the strength of their connections via processes that require chromatin remodeling and changes in gene expression. The chromodomain helicase DNA binding (CHD) protein, Kismet (Kis), is expressed in both motor neuron nuclei and postsynaptic muscle nuclei of the Drosophila larvae. This study shows that Kis is important for motor neuron synaptic morphology, the localization and clustering of postsynaptic glutamate receptors, larval motor behavior, and synaptic transmission. The data suggest that Kis is part of the machinery that modulates the development and function of the NMJ. Kis is the homolog to human CHD7, which is mutated in CHARGE syndrome. Thus, the data suggest novel avenues of investigation for synaptic defects associated with CHARGE syndrome.
Chen, Z. H., Zhu, M., Yang, J., Liang, H., He, J., He, S., Wang, P., Kang, X., McNutt, M. A., Yin, Y. and Shen, W. H. (2014). PTEN interacts with histone H1 and controls chromatin condensation. Cell Rep 8: 2003-2014. PubMed ID: 25199838
Chromatin organization and dynamics are integral to global gene transcription. Histone modification influences chromatin status and gene expression. PTEN plays multiple roles in tumor suppression, development, and metabolism. This study, performed with HeLa cells, reports on the interplay of PTEN, histone H1, and chromatin. Loss of PTEN leads to dissociation of histone H1 from chromatin and decondensation of chromatin. PTEN deletion also results in elevation of histone H4 acetylation at lysine 16, an epigenetic marker for chromatin activation. PTEN and histone H1 physically interact through their C-terminal domains. Disruption of the PTEN C terminus promotes the chromatin association of MOF acetyltransferase and induces H4K16 acetylation. Hyperacetylation of H4K16 impairs the association of PTEN with histone H1, which constitutes regulatory feedback that may reduce chromatin stability. These results demonstrate that PTEN controls chromatin condensation, thus influencing gene expression. It is proposed that PTEN regulates global gene transcription profiling through histones and chromatin remodeling.
Frey, A., Listovsky, T., Guilbaud, G., Sarkies, P. and Sale, J. E. (2014). Histone H3.3 is required to maintain replication fork progression after UV damage. Curr Biol 24: 2195-2201. PubMed ID: 25201682
Unlike histone H3, which is present only in S phase, the variant histone H3.3 (see Drosophila Histone H3.3A) is expressed throughout the cell cycle and is incorporated into chromatin independent of replication. Recently, H3.3 has been implicated in the cellular response to ultraviolet (UV) light. Chicken DT40 cells completely lacking H3.3 are hypersensitive to UV light, a defect that epistasis analysis suggests may result from less-effective nucleotide excision repair. Unexpectedly, H3.3-deficient cells also exhibit a substantial defect in maintaining replication fork progression on UV-damaged DNA, which is independent of nucleotide excision repair, demonstrating a clear requirement for H3.3 during S phase. Both the UV hypersensitivity and replication fork slowing are reversed by expression of H3.3 and require the specific residues in the alpha2 helix that are responsible for H3.3 binding its dedicated chaperones. However, expression of an H3.3 mutant in which serine 31 is replaced with alanine, the equivalent residue in H3.2, restores normal fork progression but not UV resistance, suggesting that H3.3[S31A] may be incorporated at UV-damaged forks but is unable to help cells tolerate UV lesions. Similar behavior was observed with expression of H3.3 carrying mutations at K27 and G34, which have been reported in pediatric brain cancers. It is speculated that incorporation of H3.3 during replication may mark sites of lesion bypass and, possibly through an as-yet-unidentified function of the N-terminal tail, facilitate subsequent processing of the damage.
Monday, November 24th
Moris-Sanz, M., Estacio-Gomez, A., Alvarez-Rivero, J. and Diaz-Benjumea, F. J. (2014). Specification of neuronal subtypes by different levels of Hunchback. Development 141: 4366-4374. PubMed ID: 25344076
During the development of the central nervous system, neural progenitors generate an enormous number of distinct types of neuron and glial cells by asymmetric division. Intrinsic genetic programs define the combinations of transcription factors that determine the fate of each cell, but the precise mechanisms by which all these factors are integrated at the level of individual cells are poorly understood. This study analyzed the specification of the neurons in the ventral nerve cord of Drosophila that express Crustacean cardioactive peptide (CCAP). There are two types of CCAP neurons: interneurons and efferent neurons. Both were found to be specified during the Hunchback temporal window of neuroblast 3-5, but are not sibling cells. Further, this temporal window generates two ganglion mother cells that give rise to four neurons, which can be identified by the expression of empty spiracles. The expression of Hunchback in the neuroblast increases over time, and evidence is provided that the absolute levels of Hunchback expression specify the two different CCAP neuronal fates.
Ka, M., Condorelli, G., Woodgett, J. R. and Kim, W. Y. (2014). mTOR regulates brain morphogenesis by mediating GSK3 signaling. Development 141: 4076-4086. PubMed ID: 25273085
Balanced control of neural progenitor maintenance and neuron production is crucial in establishing functional neural circuits during brain development, and abnormalities in this process are implicated in many neurological diseases. However, the regulatory mechanisms of neural progenitor homeostasis remain poorly understood. This study shows that mammalian target of rapamycin (mTOR see Drosophila Tor) is required for maintaining neural progenitor pools and plays a key role in mediating glycogen synthase kinase 3 (GSK3; see Drosophila Shaggy) signaling during brain development. First, conditional mutant mice exhibiting deletion of mTOR were generated and characterized in neural progenitors and neurons in the developing brain using Nestin-cre and Nex-cre lines, respectively. The elimination of mTOR resulted in abnormal cell cycle progression of neural progenitors in the developing brain and thereby disruption of progenitor self-renewal. Accordingly, production of intermediate progenitors and postmitotic neurons were markedly suppressed. Next, it was discovered that GSK3, a master regulator of neural progenitors, interacts with mTOR and controls its activity in cortical progenitors. Finally, it was found that inactivation of mTOR activity suppresses the abnormal proliferation of neural progenitors induced by GSK3 deletion. These findings reveal that the interaction between mTOR and GSK3 signaling plays an essential role in dynamic homeostasis of neural progenitors during brain development.
Lavado, A., Ware, M., Pare, J. and Cao, X. (2014). The tumor suppressor Nf2 regulates corpus callosum development by inhibiting the transcriptional coactivator Yap. Development 141: 4182-4193. PubMed ID: 25336744
The corpus callosum connects cerebral hemispheres and is the largest axon tract in the mammalian brain. Callosal malformations are among the most common congenital brain anomalies and are associated with a wide range of neuropsychological deficits. Crossing of the midline by callosal axons relies on a proper midline environment that harbors guidepost cells emitting guidance cues to instruct callosal axon navigation. Little is known about what controls the formation of the midline environment. This study found that two components of the Hippo pathway, the tumor suppressor Nf2 (Merlin; see Drosophila Merlin) and the transcriptional coactivator Yap (Yap1; see Drosophila Yorkie), regulate guidepost development and expression of the guidance cue Slit2 in mouse. During normal brain development, Nf2 suppresses Yap activity in neural progenitor cells to promote guidepost cell differentiation and prevent ectopic Slit2 expression. Loss of Nf2 causes malformation of midline guideposts and Slit2 upregulation, resulting in callosal agenesis. Slit2 heterozygosity and Yap deletion both restore callosal formation in Nf2 mutants. Furthermore, selectively elevating Yap activity in midline neural progenitors is sufficient to disrupt guidepost formation, upregulate Slit2 and prevent midline crossing. The Hippo pathway is known for its role in controlling organ growth and tumorigenesis. This study identifies a novel role of this pathway in axon guidance. Moreover, by linking axon pathfinding and neural progenitor behaviors, these results provide an example of the intricate coordination between growth and wiring during brain development.
Norris, A. D., Sundararajan, L., Morgan, D. E., Roberts, Z. J. and Lundquist, E. A. (2014). The UNC-6/Netrin receptors UNC-40/DCC and UNC-5 inhibit growth cone filopodial protrusion via UNC-73/Trio, Rac-like GTPases and UNC-33/CRMP. Development 141: 4395-4405. PubMed ID: 25371370
UNC-6/Netrin is a conserved axon guidance cue that can mediate both attraction and repulsion. Previous studies have discovered that attractive UNC-40/DCC receptor signaling (see Drosophila Frazzled) stimulates growth cone filopodial protrusion and that repulsive UNC-40-UNC-5 heterodimers (see Drosophila Unc5) inhibit filopodial protrusion in C. elegans. This study identified cytoplasmic signaling molecules required for UNC-6-mediated inhibition of filopodial protrusion involved in axon repulsion. The Rac-like GTPases CED-10 and MIG-2, the Rac GTP exchange factor UNC-73/Trio, UNC-44/Ankyrin and UNC-33/CRMP act in inhibitory UNC-6 signaling. These molecules were required for the normal limitation of filopodial protrusion in developing growth cones and for inhibition of growth cone filopodial protrusion caused by activated MYR::UNC-40 and MYR::UNC-5 receptor signaling. Epistasis studies using activated CED-10 and MIG-2 indicated that UNC-44 and UNC-33 act downstream of the Rac-like GTPases in filopodial inhibition. UNC-73, UNC-33 and UNC-44 did not affect the accumulation of full-length UNC-5::GFP and UNC-40::GFP in growth cones, consistent with a model in which UNC-73, UNC-33 and UNC-44 influence cytoskeletal function during growth cone filopodial inhibition.
Sunday, November 23rd
Codelia, V. A., Sun, G. and Irvine, K. D. (2014). Regulation of YAP by mechanical strain through Jnk and Hippo signaling. Curr Biol 24: 2012-2017. PubMed ID: 25127217
Mechanical forces affect all the tissues of our bodies. Experiments conducted mainly on cultured cells have established that altering these forces influences cell behaviors, including migration, differentiation, apoptosis, and proliferation. The transcriptional coactivator YAP (see Drosophila Yorkie) has been identified as a nuclear relay of mechanical signals, but the molecular mechanisms that lead to YAP activation were not identified. YAP is the main transcriptional effector of the Hippo signaling pathway, a major growth regulatory pathway within metazoa, but at least in some instances, the influence of mechanical strain on YAP was reported to be independent of Hippo signaling. This study identified a molecular pathway that can promote the proliferation of cultured mammary epithelial cells in response to cyclic or static stretch. These mechanical stimuli are associated with increased activity of the transcriptional coactivator YAP, which is due at least in part to inhibition of Hippo pathway activity. Much of this influence on Hippo signaling can be accounted for by the activation of c-Jun N-terminal kinase (JNK; see Drosophila JNK) activity by mechanical strain and subsequent inhibition of Hippo signaling by JNK. LATS1 (see Drosophila Warts) is a key negative regulator of YAP within the Hippo pathway, and this study further shows that cyclic stretch is associated with a JNK-dependent increase in binding of a LATS inhibitor, LIMD1 (Drosophila homolog: Ajuba LIM protein), to the LATS1 kinase and that reduction of LIMD1 expression suppresses the activation of YAP by cyclic stretch. Together, these observations establish a pathway for mechanical regulation of cell proliferation via JNK-mediated inhibition of Hippo signaling.
Csibi, A., Lee, G., Yoon, S. O., Tong, H., Ilter, D., Elia, I., Fendt, S. M., Roberts, T. M. and Blenis, J. (2014). The mTORC1/S6K1 pathway regulates glutamine metabolism through the eIF4B-cependent control of c-Myc translation. Curr Biol 24: 2274-2280. PubMed ID: 25220053
Growth-promoting signaling molecules, including the mammalian target of rapamycin complex 1 (mTORC1; see Drosophila Tor), drive the metabolic reprogramming of cancer cells required to support their biosynthetic needs for rapid growth and proliferation. Glutamine is catabolyzed to alpha-ketoglutarate (alphaKG), a tricarboxylic acid (TCA) cycle intermediate, through two deamination reactions, the first requiring glutaminase (GLS) to generate glutamate and the second occurring via glutamate dehydrogenase (GDH) or transaminases. Activation of the mTORC1 pathway has been shown previously to promote the anaplerotic entry of glutamine to the TCA cycle via GDH. Moreover, mTORC1 activation also stimulates the uptake of glutamine, but the mechanism is unknown. It is generally thought that rates of glutamine utilization are limited by mitochondrial uptake via GLS, suggesting that, in addition to GDH, mTORC1 could regulate GLS. This study demonstrates that mTORC1 positively regulates GLS and glutamine flux through this enzyme. mTORC1 controls GLS levels through the S6K1-dependent (see Drosophila S6K) regulation of c-Myc (Myc; see Drosophila Myc). Molecularly, S6K1 enhances Myc translation efficiency by modulating the phosphorylation of eukaryotic initiation factor eIF4B (see Drosophila eIF4B), which is critical to unwind its structured 5' untranslated region (5'UTR). Finally, these data show that the pharmacological inhibition of GLS is a promising target in pancreatic cancers expressing low levels of PTEN.
Ellis, S. J., Lostchuck, E., Goult, B. T., Bouaouina, M., Fairchild, M. J., Lopez-Ceballos, P., Calderwood, D. A. and Tanentzapf, G. (2014). The Talin head domain reinforces integrin-mediated adhesion by promoting adhesion complex stability and clustering. PLoS Genet 10: e1004756. PubMed ID: 25393120
Talin serves an essential function during integrin-mediated adhesion in linking integrins to actin via the intracellular adhesion complex. In addition, the N-terminal head domain of talin regulates the affinity of integrins for their ECM-ligands, a process known as inside-out activation. Previous studies have shown that in Drosophila, mutating the integrin binding site in the talin head domain resulted in weakened adhesion to the ECM. Intriguingly, subsequent studies showed that canonical inside-out activation of integrin might not take place in flies. Consistent with this, a mutation in talin that specifically blocks its ability to activate mammalian integrins does not significantly impinge on talin function during fly development. This study describes results suggesting that the talin head domain reinforces and stabilizes the integrin adhesion complex by promoting integrin clustering distinct from its ability to support inside-out activation. Specifically, an allele of talin containing a mutation that disrupts intramolecular interactions within the talin head was shown to attenuate the assembly and reinforcement of the integrin adhesion complex. Importantly, evidence is provided that this mutation blocks integrin clustering in vivo. It is proposed that the talin head domain is essential for regulating integrin avidity in Drosophila and that this is crucial for integrin-mediated adhesion during animal development.
Huang, Y. C., Lu, Y. N., Wu, J. T., Chien, C. T. and Pi, H. (2014). The COP9 signalosome converts temporal hormone signaling to spatial restriction on neural competence. PLoS Genet 10: e1004760. PubMed ID: 25393278
During development, neural competence is conferred and maintained by integrating spatial and temporal regulations. The Drosophila sensory bristles that detect mechanical and chemical stimulations are arranged in stereotypical positions. The anterior wing margin (AWM) is arrayed with neuron-innervated sensory bristles, while posterior wing margin (PWM) bristles are non-innervated. This study found that the COP9 signalosome (CSN; see CSN5) suppresses the neural competence of non-innervated bristles at the PWM. In CSN mutants, PWM bristles are transformed into neuron-innervated, which is attributed to sustained expression of the neural-determining factor Senseless (Sens). The CSN suppresses Sens through repression of the ecdysone signaling target gene broad (br) that encodes the BR-Z1 transcription factor to activate sens expression. Strikingly, CSN suppression of BR-Z1 is initiated at the prepupa-to-pupa transition, leading to Sens downregulation, and termination of the neural competence of PWM bristles. The role of ecdysone signaling to repress br after the prepupa-to-pupa transition is distinct from its conventional role in activation, and requires CSN deneddylating activity and multiple cullins, the major substrates of deneddylation. Several CSN subunits physically associate with ecdysone receptors to represses br at the transcriptional level. A model is proposed in which nuclear hormone receptors cooperate with the deneddylation machinery to temporally shutdown downstream target gene expression, conferring a spatial restriction on neural competence at the PWM.
Saturday, November 22nd
Vachias, C., Fritsch, C., Pouchin, P., Bardot, O. and Mirouse, V. (2014). Tight coordination of growth and differentiation between germline and soma provides robustness for Drosophila egg development. Cell Rep 9: 531-541. PubMed ID: 25373901
Organs often need to coordinate the growth of distinct tissues during their development. This study analyzed the coordination between germline cysts and the surrounding follicular epithelium during Drosophila oogenesis. Genetic manipulations of the growth rate of both germline and somatic cells influence the growth of the other tissue accordingly. Growth coordination is therefore ensured by a precise, two-way, intrinsic communication. This coordination tends to maintain constant epithelial cell shape, ensuring tissue homeostasis. Moreover, this intrinsic growth coordination mechanism also provides cell differentiation synchronization. Among growth regulators, PI3-kinase and TORC1 also influence differentiation timing cell-autonomously. However, these two pathways are not regulated by the growth of the adjacent tissue, indicating that their function reflects an extrinsic and systemic influence. Altogether, these results reveal an integrated and particularly robust mechanism ensuring the spatial and temporal coordination of tissue size, cell size, and cell differentiation for the proper development of two adjacent tissues.
Armstrong, A. R., Laws, K. M. and Drummond-Barbosa, D. (2014). Adipocyte amino acid sensing controls adult germline stem cell number via the amino acid response pathway and independently of Target of Rapamycin signaling in Drosophila. Development 141: 4479-4488. PubMed ID: 25359724
How adipocytes contribute to the physiological control of stem cells is a critical question towards understanding the link between obesity and multiple diseases, including cancers. Previous studies have revealed that adult stem cells are influenced by whole-body physiology through multiple diet-dependent factors. For example, nutrient-dependent pathways acting within the Drosophila ovary control the number and proliferation of germline stem cells (GSCs). The potential role of nutrient sensing by adipocytes in modulating stem cells in other organs, however, remains largely unexplored. This study report that amino acid sensing by adult adipocytes specifically modulates the maintenance of GSCs through a Target of Rapamycin-independent mechanism. Instead, reduced amino acid levels and the consequent increase in uncoupled tRNAs trigger activation of the GCN2-dependent amino acid response pathway within adipocytes, causing increased rates of GSC loss. These studies reveal a new step in adipocyte-stem cell crosstalk.
Matsui, Y., Takehara, A., Tokitake, Y., Ikeda, M., Obara, Y., Morita-Fujimura, Y., Kimura, T. and Nakano, T. (2014). The majority of early primordial germ cells acquire pluripotency by AKT activation. Development 141: 4457-4467. PubMed ID: 25359722
Primordial germ cells (PGCs) are undifferentiated germ cells in embryos, the fate of which is to become gametes; however, mouse PGCs can easily be reprogrammed into pluripotent embryonic germ cells (EGCs) in culture in the presence of particular extracellular factors, such as combinations of Steel factor (KITL), LIF and bFGF (FGF2). Early PGCs form EGCs more readily than do later PGCs, and PGCs lose the ability to form EGCs by embryonic day (E) 15.5. This study examined the effects of activation of the serine/threonine kinase AKT in PGCs during EGC formation; notably, AKT activation, in combination with LIF and bFGF, enhanced EGC formation and caused approximately 60% of E10.5 PGCs to become EGCs. The results indicate that the majority of PGCs at E10.5 could acquire pluripotency with an activated AKT signaling pathway. Importantly, AKT activation did not fully substitute for bFGF and LIF, and AKT activation without both LIF and bFGF did not result in EGC formation. These findings indicate that AKT signal enhances and/or collaborates with signaling pathways of bFGF and of LIF in PGCs for the acquisition of pluripotency.
Garcia, T. X., Farmaha, J. K., Kow, S. and Hofmann, M. C. (2014). RBPJ in mouse Sertoli cells is required for proper regulation of the testis stem cell niche. Development 141: 4468-4478. PubMed ID: 25406395
Stem cells are influenced by their surrounding microenvironment, or niche. In the testis, Sertoli cells are the key niche cells directing the population size and differentiation fate of spermatogonial stem cells (SSCs). Failure to properly regulate SSCs leads to infertility or germ cell hyperplasia. Several Sertoli cell-expressed genes, such as Gdnf and Cyp26b1, have been identified as being indispensable for the proper maintenance of SSCs in their niche, but the pathways that modulate their expression have not been identified. Although it has been recently found that constitutively activating NOTCH signaling in Sertoli cells leads to premature differentiation of all prospermatogonia and sterility, suggesting that there is a crucial role for this pathway in the testis stem cell niche, a true physiological function of NOTCH signaling in Sertoli cells has not been demonstrated. To this end, recombination signal binding protein for immunoglobulin kappa J region (Rbpj), a crucial mediator of NOTCH signaling, was conditionally ablated in Sertoli cells using Amh-cre. Rbpj knockout mice had: significantly increased testis sizes; increased expression of niche factors, such as Gdnf and Cyp26b1; significant increases in the number of pre- and post-meiotic germ cells, including SSCs; and, in a significant proportion of mice, testicular failure and atrophy with tubule lithiasis, possibly due to these unsustainable increases in the number of germ cells. Germ cells were identified as the NOTCH ligand-expressing cells. It is concluded that NOTCH signaling in Sertoli cells is required for proper regulation of the testis stem cell niche and is a potential feedback mechanism, based on germ cell input, that governs the expression of factors that control SSC proliferation and differentiation.
Zhang, T., Murphy, M. W., Gearhart, M. D., Bardwell, V. J. and Zarkower, D. (2014). The mammalian Doublesex homolog DMRT6 coordinates the transition between mitotic and meiotic developmental programs during spermatogenesis. Development 141: 3662-3671. PubMed ID: 25249458
In mammals, a key transition in spermatogenesis is the exit from spermatogonial differentiation and mitotic proliferation and the entry into spermatocyte differentiation and meiosis. Although several genes that regulate this transition have been identified, how it is controlled and coordinated remains poorly understood. This study examined the role in male gametogenesis of the Doublesex-related gene Dmrt6 (Dmrtb1; see Drosophila Doublesex) in mice and find that Dmrt6 plays a crucial role in directing germ cells through the mitotic-to-meiotic germ cell transition. DMRT6 protein is expressed in late mitotic spermatogonia. In mice of the C57BL/6J strain, a null mutation in Dmrt6 disrupts spermatogonial differentiation, causing inappropriate expression of spermatogonial differentiation factors, including SOHLH1, SOHLH2 and DMRT1 as well as the meiotic initiation factor STRA8, and causing most late spermatogonia to undergo apoptosis. In mice of the 129Sv background, most Dmrt6 mutant germ cells can complete spermatogonial differentiation and enter meiosis, but they show defects in meiotic chromosome pairing, establishment of the XY body and processing of recombination foci, and they mainly arrest in mid-pachynema. mRNA profiling of Dmrt6 mutant testes together with DMRT6 chromatin immunoprecipitation sequencing suggest that DMRT6 represses genes involved in spermatogonial differentiation and activates genes required for meiotic prophase. These results indicate that Dmrt6 plays a key role in coordinating the transition in gametogenic programs from spermatogonial differentiation and mitosis to spermatocyte development and meiosis.
Friday, November 21st
Hasygar, K. and Hietakangas, V. (2014). p53- and ERK7-dependent ribosome surveillance response regulates Drosophila Insulin-Like peptide secretion. PLoS Genet 10: e1004764. PubMed ID: 25393288
Insulin-like signalling is a conserved mechanism that coordinates animal growth and metabolism with nutrient status. In Drosophila, insulin-producing median neurosecretory cells (IPCs) regulate larval growth by secreting insulin-like peptides (dILPs) in a diet-dependent manner. Previous studies have shown that nutrition affects dILP secretion through humoral signals derived from the fat body. This study uncovered a novel mechanism that operates cell autonomously in the IPCs to regulate dILP secretion. Impairment of ribosome biogenesis specifically in the IPCs was shown to strongly inhibits dILP secretion, consequently leading to reduced body size and a delay in larval development. This response is dependent on p53, a known surveillance factor for ribosome biogenesis. A downstream effector of this growth inhibitory response is an atypical MAP kinase ERK7 (ERK8/MAPK15), which is upregulated in the IPCs following impaired ribosome biogenesis as well as starvation. ERK7 is sufficient and essential to inhibit dILP secretion upon impaired ribosome biogenesis, and it acts epistatically to p53. Moreover, evidence is provided that p53 and ERK7 contribute to the inhibition of dILP secretion upon starvation. Thus, it is concluded that a cell autonomous ribosome surveillance response, which leads to upregulation of ERK7, inhibits dILP secretion to impede tissue growth under limiting dietary conditions.
Ham, H., Woolery, A. R., Tracy, C., Stenesen, D., Kramer, H. and Orth, K. (2014). Unfolded protein response-regulated dFic reversibly AMPylates BiP during endoplasmic reticulum homeostasis. J Biol Chem. PubMed ID: 25395623
Drosophila Fic (dFic) mediates AMPylation, a covalent attachment of adenosine monophosphate (AMP) from ATP to hydroxyl side chains of protein substrates. This study identified the endoplasmic reticulum (ER) chaperone BiP (Heat shock 70-kDa protein cognate 3) as a substrate for dFic; the modification site was mapped to Thr366 within the ATPase domain. The level of AMPylated BiP in Drosophila S2 cells is high during homeostasis, whereas the level of AMPylated BiP decreases upon the accumulation of misfolded proteins in the ER. Both dFic and BiP are transcriptionally activated upon ER stress, supporting the role of dFic in the unfolded protein response (UPR) pathway. The inactive conformation of BiP is the preferred substrate for dFic, thus endorsing a model whereby AMPylation regulates the function of BiP as a chaperone, allowing acute activation of BiP by deAMPylation during an ER stress response. These findings not only present the first substrate of eukaryotic AMPylator but also provide a target for regulating the UPR, an emerging avenue for cancer therapy.
Loubery, S., Seum, C., Moraleda, A., Daeden, A., Furthauer, M. and Gonzalez-Gaitan, M. (2014). Uninflatable and Notch control the targeting of sara endosomes during asymmetric division. Curr Biol 24: 2142-2148. PubMed ID: 25155514
Cell fate decision during asymmetric division is mediated by the biased partition of cell fate determinants during mitosis. In the case of the asymmetric division of the fly sensory organ precursor cells, directed Notch signaling from pIIb to the pIIa daughter endows pIIa with its distinct fate. Previous studies have shown that Notch/Delta molecules internalized in the mother cell traffic through Sara endosomes and are directed to the pIIa daughter. This study shows that the receptor Notch itself is required during the asymmetric targeting of the Sara endosomes to pIIa. Notch binds Uninflatable, and both traffic together through Sara endosomes, which is essential to direct asymmetric endosomes motility and Notch-dependent cell fate assignation. The data uncover a part of the core machinery required for the asymmetric motility of a vesicular structure that is essential for the directed dispatch of Notch signaling molecules during asymmetric mitosis.
Oberhofer, G., Grossmann, D., Siemanowski, J. L., Beissbarth, T. and Bucher, G. (2014). Wnt/β-catenin signaling integrates patterning and metabolism of the insect growth zone. Development [Epub ahead of print]. PubMed ID: 25395458
Wnt/β-catenin and Hedgehog (Hh) signaling are essential for transmitting signals across cell membranes in animal embryos. Early patterning of the principal insect model, Drosophila melanogaster, occurs in the syncytial blastoderm, where diffusion of transcription factors obviates the need for signaling pathways. However, in the cellularized growth zone of typical short germ insect embryos, signaling pathways are predicted to play a more fundamental role. Indeed, the Wnt/β-catenin pathway is required for posterior elongation in most arthropods, although which target genes are activated in this context remains elusive. This study used the short germ beetle Tribolium castaneum to investigate two Wnt and Hh signaling centers located in the head anlagen and in the growth zone of early embryos. Wnt/β-catenin signaling was found to act upstream of Hh in the growth zone, whereas the opposite interaction occurs in the head. The target gene sets of the Wnt/β-catenin and Hh pathways were determined; the growth zone signaling center activates a much greater number of genes and the Wnt and Hh target gene sets are essentially non-overlapping. The Wnt pathway activates key genes of all three germ layers, including pair-rule genes, and Tc-caudal (see Drosophila caudal) and Tc-twist (see Drosophila twist). Furthermore, the Wnt pathway is required for hindgut development and Tc-senseless (see Drosophila senseless) as a novel hindgut patterning gene required in the early growth zone. At the same time, Wnt acts on growth zone metabolism and cell division, thereby integrating growth with patterning. Posterior Hh signaling activates several genes potentially involved in a proteinase cascade of unknown function.
Thursday, November 20th
Jumbo-Lucioni, P., Parkinson, W. and Broadie, K. (2014). Altered synaptic architecture and glycosylated synaptomatrix composition in a Drosophila classic galactosemia disease model. Dis Model Mech [Epub ahead of print]. PubMed ID: 25326312
Classic galactosemia (CG) is an autosomal recessive disorder resulting from loss of galactose-1-phosphate uridyltransferase (GALT) that catalyzes conversion of galactose-1-phosphate and uridine diphosphate (UDP)-glucose to glucose-1-phosphate and UDP-galactose, immediately upstream of UDP-N-acetylgalactosamine and UDP-N-acetylglucosamine synthesis. These four UDP-sugars are essential donors driving synthesis of glycoproteins and glycolipids that heavily decorate cell surfaces and extracellular spaces. In addition to acute, potentially lethal neonatal symptoms, maturing CG patients develop striking neurodevelopmental, motor and cognitive impairments. Previous studies suggest association of neurological symptoms with glycosylation defects, with CG recently described as a Congenital Disorder of Glycosylation (CDG) showing defects in both N- and O-linked glycans. This study characterized behavioral traits, synaptic development and glycosylated synaptomatrix formation in a GALT-deficient Drosophila disease model. Loss of Drosophila GALT (dGALT) greatly impairs coordinated movement and results in structural overelaboration and architectural abnormalities at the neuromuscular junction (NMJ). Dietary galactose and mutation of galactokinase (dGALK) or UDP-glucose dehydrogenase (sugarless) genes are identified as critical environmental and genetic modifiers, respectively, of behavioral and cellular defects. Assaying the NMJ extracellular synaptomatrix with a broad panel of lectin probes reveals profound alterations in dGALT mutants, including depletion of galactosyl/N-acetylgalactosamine and fucosylated horseradish peroxidase (HRP) moieties, which are differentially corrected by dGALK co-removal and sugarless overexpression. Synaptogenesis relies on trans-synaptic signals modulated by this synaptomatrix carbohydrate environment, and dGALT null NMJs display striking changes in heparan sulfate proteoglycan (HSPG) co-receptor and Wnt ligand levels, which are also corrected by dGALK co-removal and sugarless overexpression. These results reveal synaptomatrix glycosylation losses, altered trans-synaptic signaling pathway components, defective synaptogenesis and impaired coordinated movement in a CG neurological disease model.
Bou Dib, P., Gnagi, B., Daly, F., Sabado, V., Tas, D., Glauser, D. A., Meister, P. and Nagoshi, E. (2014). A conserved role for p48 homologs in protecting Dopaminergic neurons from oxidative stress. PLoS Genet 10: e1004718. PubMed ID: 25340742
Parkinson's disease (PD) is the most common neurodegenerative movement disorder characterized by the progressive loss of dopaminergic (DA) neurons. Both environmental and genetic factors are thought to contribute to the pathogenesis of PD. Although several genes linked to rare familial PD have been identified, endogenous risk factors for sporadic PD, which account for the majority of PD cases, remain largely unknown. Genome-wide association studies have identified many single nucleotide polymorphisms associated with sporadic PD in neurodevelopmental genes including the transcription factor p48/ptf1a. This study investigatee whether p48 plays a role in the survival of DA neurons in Drosophila melanogaster and Caenorhabditis elegans. A Drosophila p48 homolog, 48-related-2 (Fer2), is expressed in and required for the development and survival of DA neurons in the protocerebral anterior medial (PAM) cluster. Loss of Fer2 expression in adulthood causes progressive PAM neuron degeneration in aging flies along with mitochondrial dysfunction and elevated reactive oxygen species (ROS) production, leading to the progressive locomotor deficits. The oxidative stress challenge upregulates Fer2 expression and exacerbates the PAM neuron degeneration in Fer2 loss-of-function mutants. hlh-13, the worm homolog of p48, is also expressed in DA neurons. Unlike the fly counterpart, hlh-13 loss-of-function does not impair development or survival of DA neurons under normal growth conditions. Yet, similar to Fer2, hlh-13 expression is upregulated upon an acute oxidative challenge and is required for the survival of DA neurons under oxidative stress in adult worms. Taken together, these results indicate that p48 homologs share a role in protecting DA neurons from oxidative stress and degeneration, and suggest that loss-of-function of p48 homologs in flies and worms provides novel tools to study gene-environmental interactions affecting DA neuron survival.
Tsai, P. I., Course, M. M., Lovas, J. R., Hsieh, C. H., Babic, M., Zinsmaier, K. E. and Wang, X. (2014). PINK1-mediated phosphorylation of Miro inhibits synaptic growth and protects dopaminergic neurons in Drosophila. Sci Rep 4: 6962. PubMed ID: 25376463
Mutations in the mitochondrial Ser/Thr kinase PINK1 cause Parkinson's disease. One of the substrates of PINK1 is the outer mitochondrial membrane protein Miro, which regulates mitochondrial transport. This study uncovered novel physiological functions of PINK1-mediated phosphorylation of Miro, using Drosophila as a model. Endogenous Drosophila Miro (DMiro) was replaced with transgenically expressed wildtype, or mutant DMiro predicted to resist PINK1-mediated phosphorylation. The expression of phospho-resistant DMiro in a DMiro null mutant background was found to phenocopy a subset of phenotypes of PINK1 null. Specifically, phospho-resistant DMiro increased mitochondrial movement and synaptic growth at larval neuromuscular junctions, and decreased the number of dopaminergic neurons in adult brains. Therefore, PINK1 may inhibit synaptic growth and protect dopaminergic neurons by phosphorylating DMiro. Furthermore, muscle degeneration, swollen mitochondria and locomotor defects found in PINK1 null flies were not observed in phospho-resistant DMiro flies. Thus, this study established an in vivo platform to define functional consequences of PINK1-mediated phosphorylation of its substrates.
Luchtenborg, A. M. and Katanaev, V. L. (2014). Lack of evidence of the interaction of the Ass peptide with the Wnt signaling cascade in Drosophila models of Alzheimer's disease. Mol Brain 7: 81. PubMed ID: 25387847
Alzheimer's disease (AD) is the leading form of dementia worldwide. The Ass-peptide (see Drosophila Appl) is believed to be the major pathogenic compound of the disease. Since several years it is hypothesized that Ass impacts the Wnt signaling cascade and therefore activation of this signaling pathway is proposed to rescue the neurotoxic effect of Ass. Expression of the human Ass42 in the Drosophila nervous system leads to a drastically shortened life span. The action of Ass42 specifically in the glutamatergic motoneurons is responsible for the reduced survival. However, this study found that the morphology of the glutamatergic larval neuromuscular junctions, which are widely used as the model for mammalian central nervous system synapses, is not affected by Ass42 expression. It was furthermore demonstrated that genetic activation of the Wnt signal transduction pathway in the nervous system is not able to rescue the shortened life span or a rough eye phenotype in Drosophila.
These data confirm that the life span is a useful readout of Ass42 induced neurotoxicity in Drosophila; the neuromuscular junction seems however not to be an appropriate model to study AD in flies. Additionally, these results challenge the hypothesis that Wnt signaling might be implicated in Ass42 toxicity and might serve as a drug target against AD.
Logan-Garbisch, T., Bortolazzo, A., Luu, P., Ford, A., Do, D., Khodabakhshi, P. and French, R. L. (2014). Developmental ethanol exposure leads to dysregulation of lipid metabolism and oxidative stress in Drosophila. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 25387828
Ethanol exposure during development causes an array of developmental abnormalities, both physiological and behavioral. In mammals, these abnormalities are collectively known as Fetal Alcohol Effects (FAE) or Fetal Alcohol Spectrum Disorder (FASD). This study has established a Drosophila melanogaster model of FASD; it was previously shown that developmental ethanol exposure in flies leads to reduced expression of insulin like peptides (dILPs; see Insulin-related peptide) and their receptor. This work linked that observation to dysregulation of fatty acid metabolism and lipid accumulation. Further, it was shown that developmental ethanol exposure in Drosophila causes oxidative stress, that this stress is a primary cause of the developmental lethality and delay associated with ethanol exposure, and, finally, that one of the mechanisms by which ethanol increases oxidative stress is through abnormal fatty acid metabolism. These data suggest a previously uncharacterized mechanism by which ethanol causes the symptoms associated with FASD.
Wednesday, November 19th
Delabaere, L., Orsi, G. A., Sapey-Triomphe, L., Horard, B., Couble, P. and Loppin, B. (2014). The spartan ortholog maternal haploid is required for paternal chromosome integrity in the Drosophila zygote. Curr Biol 24: 2281-2287. PubMed ID: 25242033Summary:
The animal sperm nucleus is characterized by an extremely compacted organization of its DNA after the global replacement of histones with sperm-specific nuclear basic proteins, such as protamines. In the absence of DNA repair activity in the mature gamete, the integrity of the paternal genome is potentially challenged by the unique topological constraints exerted on sperm DNA. In addition, the maintenance of paternal DNA integrity during the rapid remodeling of sperm chromatin at fertilization has long been regarded as a maternal trait. However, little is known about the nature of the egg proteins involved in this essential aspect of zygote formation. Previous work has characterized the unique phenotype of the classical Drosophila maternal effect mutant maternal haploid (mh), which specifically affects the integration of paternal chromosomes in the zygote. This study shows that MH is the fly ortholog of the recently identified human DVC1/Spartan protein, a conserved regulator of DNA damage tolerance. Like Spartan, MH protein is involved in the resistance to UV radiation and recruits the p97/TER94 segregase to stalled DNA replication forks in somatic cells. In the zygote, mh phenotype was found to be consistent with perturbed or incomplete paternal DNA replication. Remarkably, however, the specific accumulation of MH in the male pronucleus before the first S phase suggests that this maternal protein is required to maintain paternal DNA integrity during nuclear decondensation or to set the paternal chromatin landscape in preparation of the first zygotic cycle.
Fahmy, K., Akber, M., Cai, X., Koul, A., Hayder, A. and Baumgartner, S. (2014). αTubulin 67C and Ncd are essential for establishing a cortical microtubular network and formation of the Bicoid mRNA gradient in Drosophila. PLoS One 9: e112053. PubMed ID: 25390693
The Bicoid (Bcd) protein gradient in Drosophila serves as a paradigm for gradient formation in textbooks. To explain the generation of the gradient, the ARTS (active RNA transport and synthesis) model, which is based on the observation of a bcd mRNA gradient, proposes that the bcd mRNA, localizes at the anterior pole at fertilization, migrates along microtubules (MTs) at the cortex to the posterior to form a bcd mRNA gradient which is translated to form a protein gradient. To fulfill the criteria of the ARTS model, an early cortical MT network is thus a prerequisite. This study reports hitherto undiscovered MT activities in the early embryo important for bcd mRNA transport: (1) an early and omnidirectional MT network exclusively at the anterior cortex of early nuclear cycle embryos showing activity during metaphase and anaphase only, (2) long MTs up to 50 microm extending into the yolk at blastoderm stage to enable basal-apical transport. The cortical MT network is not anchored to the actin cytoskeleton. The posterior transport of the mRNA via the cortical MT network critically depends on maternally-expressed αTubulin67C and the minus-end motor Ncd. In either mutant, cortical transport of the alphaTubulin67C mRNA does not take place and the mRNA migrates along another yet undisclosed interior MT network, instead. These data strongly corroborate the ARTS model and explain the occurrence of the alphaTubulin67C mRNA gradient.
Ozuak, O., Buchta, T., Roth, S. and Lynch, J. A. (2014). Dorsoventral polarity of the Nasonia embryo primarily relies on a BMP gradient formed without input from Toll. Curr Biol [Epub ahead of print]. PubMed ID: 25308075
In Drosophila, Toll signaling leads to a gradient of nuclear uptake of Dorsal with a peak at the ventral egg pole and is the source for dorsoventral (DV) patterning and polarity of the embryo. In contrast, Toll signaling plays no role in embryonic patterning in mosit animals, while BMP signaling plays the major role. In order to understand the origin of the novelty of the Drosophila system, thss study has examined DV patterning in Nasonia vitripennis (Nv), a representative of the Hymenoptera and thus the most ancient branch points within the Holometabola. It has been previously shown that while the expression of several conserved DV patterning genes is almost identical in Nasonia and Drosophila embryos at the onset of gastrulation, the ways these patterns evolve in early embryogenesis are very different from what is seen in Drosophila or the beetle Tribolium. In contrast to Drosophila or Tribolium, this study found that wasp Toll has a very limited ventral role, whereas BMP is required for almost all DV polarity of the embryo, and these two signaling systems act independently of each other to generate DV polarity. This result gives insights into how the Toll pathway could have usurped a BMP-based DV patterning system in insects. In addition, this work strongly suggests that a novel system for BMP activity gradient formation must be employed in the wasp, since orthologs of crucial components of the fly system are either missing entirely or lack function in the embryo.
Pare, A. C., Vichas, A., Fincher, C. T., Mirman, Z., Farrell, D. L., Mainieri, A. and Zallen, J. A. (2014). A positional Toll receptor code directs convergent extension in Drosophila. Nature [Epub ahead of print]. PubMed ID: 25363762
Elongation of the head-to-tail body axis by convergent extension is a conserved developmental process throughout metazoans. In Drosophila, patterns of transcription factor expression provide spatial cues that induce systematically oriented cell movements and promote tissue elongation. However, the mechanisms by which patterned transcriptional inputs control cell polarity and behaviour have long been elusive. This study demonstrates that three Toll family receptors, Toll-2 (18 wheeler), Toll-6 and Toll-8, are expressed in overlapping transverse stripes along the anterior-posterior axis and act in combination to direct planar polarity and polarized cell rearrangements during convergent extension. Simultaneous disruption of all three receptors strongly reduces actomyosin-driven junctional remodelling and axis elongation, and an ectopic stripe of Toll receptor expression is sufficient to induce planar polarized actomyosin contractility. These results demonstrate that tissue-level patterns of Toll receptor expression provide spatial signals that link positional information from the anterior-posterior patterning system to the essential cell behaviours that drive convergent extension.
Tuesday, November 18th
Poukkula, M., Hakala, M., Pentinmikko, N., Sweeney, M. O., Jansen, S., Mattila, J., Hietakangas, V., Goode, B. L. and Lappalainen, P. (2014). GMF promotes leading-edge dynamics and collective cell migration in vivo. Curr Biol. PubMed ID: 25308079
Lamellipodia are dynamic actin-rich cellular extensions that drive advancement of the leading edge during cell migration. Lamellipodia undergo periodic extension and retraction cycles, but the molecular mechanisms underlying these dynamics and their role in cell migration have remained obscure. This study shows that glia-maturation factor (GMF), which is an Arp2/3 (see Arpc1) complex inhibitor and actin filament debranching factor, regulates lamellipodial protrusion dynamics in living cells. In cultured S2R+ cells, GMF silencing resulted in an increase in the width of lamellipodial actin filament arrays. Importantly, live-cell imaging of mutant Drosophila egg chambers revealed that the dynamics of actin-rich protrusions in migrating border cells is diminished in the absence of GMF. Consequently, velocity of border cell clusters undergoing guided migration was reduced in GMF mutant flies. Furthermore, genetic studies demonstrated that GMF cooperates with the Drosophila homolog of Aip1 (flare) in promoting disassembly of Arp2/3-nucleated actin filament networks and driving border cell migration. These data suggest that GMF functions in vivo to promote the disassembly of Arp2/3-nucleated actin filament arrays, making an important contribution to cell migration within a 3D tissue environment.
Mukherjea, M., Ali, M. Y., Kikuti, C., Safer, D., Yang, Z., Sirkia, H., Ropars, V., Houdusse, A., Warshaw, D. M. and Sweeney, H. L. (2014). Myosin VI must dimerize and deploy its unusual lever arm in order to perform its cellular roles. Cell Rep 8: 1522-1532. PubMed ID: 25159143
It is unclear whether the reverse-direction myosin (myosin VI; see Drosophla Jaguar) functions as a monomer or dimer in cells and how it generates large movements on actin. This study deleted a stable, single-alpha-helix (SAH) domain that has been proposed to function as part of a lever arm to amplify movements without impact on in vitro movement or in vivo functions. A myosin VI construct that used this SAH domain as part of its lever arm was able to take large steps in vitro but did not rescue in vivo functions. It was necessary for myosin VI to internally dimerize, triggering unfolding of a three-helix bundle and calmodulin binding in order to step normally in vitro and rescue endocytosis and Golgi morphology in myosin VI-null fibroblasts. A model for myosin VI emerges in which cargo binding triggers dimerization and unfolds the three-helix bundle to create a lever arm essential for in vivo functions.
Khanna, M. R., Mattie, F. J., Browder, K. C., Radyk, M. D., Crilly, S. E., Bakerink, K. J., Harper, S. L., Speicher, D. W. and Thomas, G. H. (2014). Spectrin tetramer formation is not required for viable development in Drosophila. J Biol Chem [Epub ahead of print]. PubMed ID: 25381248
The dominant paradigm for spectrin function is that (αβ)2-spectrin tetramers or higher order oligomers form membrane associated two-dimensional networks in association with F-actin to reinforce the plasma membrane. Tetramerization is an essential event in such structures. This study characterized the tetramerization interaction between α-spectrin and β-spectrins in Drosophila. Wild-type α-spectrin binds to both β- and βH-chains with high affinity, resembling other non-erythroid spectrins. However, α-specR22S, a tetramerization site mutant homologous to the pathological α-specR28S allele in humans, eliminates detectable binding to β-spectrin and reduces binding to βH-spectrin ~1000 fold. Even though spectrins are essential proteins, α-specR22S rescues α-spectrin mutants to adulthood with only minor phenotypes indicating that tetramerization, and thus conventional network formation, is not the essential function of non-erythroid spectrin. These data provide the first rigorous test for the general requirement for tetramer-based non-erythroid spectrin networks throughout an organism and find that they have very limited roles, in direct contrast to the current paradigm.
Schlager, M. A., Serra-Marques, A., Grigoriev, I., Gumy, L. F., Esteves da Silva, M., Wulf, P. S., Akhmanova, A. and Hoogenraad, C. C. (2014). Bicaudal D family adaptor proteins control the velocity of Dynein-based movements. Cell Rep 8: 1248-1256. PubMed ID: 25176647
Cargo transport along microtubules is driven by the collective function of microtubule plus- and minus-end-directed motors (kinesins and dyneins). How the velocity of cargo transport is driven by opposing teams of motors is still poorly understood. This study, carried out in primary hippocampal cultures, combined inducible recruitment of motors and adaptors to Rab6 secretory vesicles with detailed tracking of vesicle movements to investigate how changes in the transport machinery affect vesicle motility. The velocities of kinesin-based vesicle movements were found to be slower and more homogeneous than those of dynein-based movements. It was also found that Bicaudal D (BICD) adaptor proteins can regulate dynein-based vesicle motility. BICD-related protein 1 (BICDR-1) accelerates minus-end-directed vesicle movements and affects Rab6 vesicle distribution. These changes are accompanied by reduced axonal outgrowth in neurons, supporting their physiological importance. This study suggests that adaptor proteins can modulate the velocity of dynein-based motility and thereby control the distribution of transport carriers.
Monday, November 17th
Olds, W. H. and Xu, T. (2014). Regulation of food intake by mechanosensory ion channels in enteric neurons. Elife 3. PubMed ID: 25285450
Regulation of food intake is fundamental to energy homeostasis in animals. The contribution of non-nutritive and metabolic signals in regulating feeding is unclear. This study shows that enteric neurons play a major role in regulating feeding through specialized mechanosensory ion channels in Drosophila. Modulating activities of a specific subset of enteric neurons, the posterior enteric neurons (PENs), results in 6-fold changes in food intake. Deficiency of the mechanosensory ion channel PPK1 gene (pickpocket) or RNAi knockdown of its expression in the PENS result in a similar increase in food intake, which can be rescued by expression of wild-type PPK1 in the same neurons. Finally, pharmacological inhibition of the mechanosensory ion channel phenocopies the result of genetic interrogation. Together, this study provides the first molecular genetic evidence that mechanosensory ion channels in the enteric neurons are involved in regulating feeding, offering an enticing alternative to current therapeutic strategy for weight control.
Chng, W. B., Sleiman, M. S., Schupfer, F. and Lemaitre, B. (2014). Transforming growth factor beta/Activin signaling functions as a sugar-sensing feedback loop to regulate digestive enzyme expression. Cell Rep 9: 336-348. PubMed ID: 25284780
Organisms need to assess their nutritional state and adapt their digestive capacity to the demands for various nutrients. Modulation of digestive enzyme production represents a rational step to regulate nutriment uptake. However, the role of digestion in nutrient homeostasis has been largely neglected. This study analyzed the mechanism underlying glucose repression of digestive enzymes in the adult Drosophila midgut. Glucose represses the expression of many carbohydrases and lipases. The data reveal that the consumption of nutritious sugars stimulates the secretion of the transforming growth factor beta (TGF-beta) ligand, Dawdle, from the fat body. Dawdle then acts via circulation to activate TGF-beta/Activin signaling in the midgut, culminating in the repression of digestive enzymes that are highly expressed during starvation. Thus, this study not only identifies a mechanism that couples sugar sensing with digestive enzyme expression but points to an important role of TGF-beta/Activin signaling in sugar metabolism.
Mackey, A. M., Sarkes, D. A., Bettencourt, I., Asara, J. M. and Rameh, L. E. (2014). PIP4kgamma is a substrate for mTORC1 that maintains basal mTORC1 signaling during starvation. Sci Signal 7: ra104. PubMed ID: 25372051
Phosphatidylinositol-5-phosphate 4-kinases (PIP4ks) are a family of lipid kinases that specifically use phosphatidylinositol 5-monophosphate (PI-5-P) as a substrate to synthesize phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]. Suppression of PIP4k function in Drosophila results in smaller cells and reduced target of rapamycin complex 1 (TORC1) signaling. This study shows that the gamma isoform of PIP4k stimulates signaling through mammalian TORC1 (mTORC1). Knockdown of PIP4kgamma reduced cell mass in cells in which mTORC1 is constitutively activated by Tsc2 (see Drosophila Gigas) deficiency. In Tsc2 null cells, mTORC1 activation was partially independent of amino acids or glucose and glutamine. PIP4kgamma knockdown inhibited the nutrient-independent activation of mTORC1 in Tsc2 knockdown cells and reduced basal mTORC1 signaling in wild-type cells. PIP4kgamma was phosphorylated by mTORC1 and associated with the complex. Phosphorylated PIP4kgamma was enriched in light microsomal vesicles, whereas the unphosphorylated form was enriched in heavy microsomal vesicles associated with the Golgi. Furthermore, basal mTORC1 signaling was enhanced by overexpression of unphosphorylated wild-type PIP4kgamma or a phosphorylation-defective mutant and decreased by overexpression of a phosphorylation-mimetic mutant. Together, these results demonstrate that PIP4kgamma and mTORC1 interact in a self-regulated feedback loop to maintain low and tightly regulated mTORC1 activation during starvation.
Kubrak, O. I., Kucerova, L., Theopold, U. and Nassel, D. R. (2014). The sleeping beauty: How reproductive diapause affects hormone signaling, Metabolism, immune response and somatic naintenance in Drosophila melanogaster. PLoS One 9: e113051. PubMed ID: 25393614
Some organisms can adapt to seasonal and other environmental challenges by entering a state of dormancy, diapause. Thus, insects exposed to decreased temperature and short photoperiod enter a state of arrested development, lowered metabolism, and increased stress resistance. Drosophila melanogaster females can enter a shallow reproductive diapause in the adult stage, which drastically reduces organismal senescence, but little is known about the physiology and endocrinology associated with this dormancy, and the genes involved in its regulation. Diapause was induced in D. melanogaster and effects were monitored over 12 weeks on dynamics of ovary development, carbohydrate and lipid metabolism, as well as expression of genes involved in endocrine signaling, metabolism and innate immunity. During diapause food intake diminishes drastically, but circulating and stored carbohydrates and lipids are elevated. Gene transcripts of glucagon- and insulin-like peptides increase, and expression of several target genes of these peptides also change. Four key genes in innate immunity can be induced by infection in diapausing flies, and two of these, Drosomycin and Cecropin A1, are upregulated by diapause independently of infection. Diapausing flies display very low mortality, extended lifespan and decreased aging of the intestinal epithelium. Many phenotypes induced by diapause are reversed after one week of recovery from diapause conditions. Furthermore, mutant flies lacking specific insulin-like peptides (dilp5 and dilp2-3) display increased diapause incidence. This study provides a first comprehensive characterization of reproductive diapause in D. melanogaster, and evidence that glucagon- and insulin-like signaling are among the key regulators of the altered physiology during this dormancy.
Sunday, November 16th
Dzhindzhev, N. S., Tzolovsky, G., Lipinszki, Z., Schneider, S., Lattao, R., Fu, J., Debski, J., Dadlez, M. and Glover, D. M. (2014). Plk4 phosphorylates Ana2 to trigger Sas6 recruitment and procentriole formation. Curr Biol [Epub ahead of print]. PubMed ID: 25264260
Centrioles are 9-fold symmetrical structures at the core of centrosomes and base of cilia whose dysfunction has been linked to a wide range of inherited diseases and cancer. Their duplication is regulated by a protein kinase of conserved structure, the C. elegans ZYG-1 or its Polo-like kinase 4 (Plk4; see Drosophila SAK) counterpart in other organisms. Although Plk4's centriolar partners and mechanisms that regulate its stability are known, its crucial substrates for centriole duplication have never been identified. This study shows that Drosophila Plk4 phosphorylates four conserved serines in the STAN motif of the core centriole protein Ana2 to enable it to bind and recruit its Sas6 partner. Ana2 and Sas6 normally load onto both mother and daughter centrioles immediately after their disengagement toward the end of mitosis to seed procentriole formation. Nonphosphorylatable Ana2 still localizes to the centriole but can no longer recruit Sas6 and centriole duplication fails. Thus, following centriole disengagement, recruitment of Ana2 and its phosphorylation by Plk4 are the earliest known events in centriole duplication to recruit Sas6 and thereby establish the architecture of the new procentriole engaged with its parent.
Carmena, M., Lombardia, M. O., Ogawa, H. and Earnshaw, W. C. (2014). Polo kinase regulates the localization and activity of the chromosomal passenger complex in meiosis and mitosis in Drosophila melanogaster. Open Biol 4. PubMed ID: 25376909
Cell cycle progression is regulated by members of the cyclin-dependent kinase (CDK), Polo and Aurora families of protein kinases. The levels of expression and localization of the key regulatory kinases are themselves subject to very tight control. There is increasing evidence that crosstalk between the mitotic kinases provides for an additional level of regulation. Previous work has shown that Aurora B activates Polo kinase at the centromere in mitosis, and that the interaction between Polo and the chromosomal passenger complex (CPC) component INCENP is essential in this activation. This report shows that Polo kinase is required for the correct localization and activity of the CPC in meiosis and mitosis. Study of the phenotype of different polo allele combinations compared to the effect of chemical inhibition revealed significant differences in the localization and activity of the CPC in diploid tissues. These results shed new light on the mechanisms that control the activity of Aurora B in meiosis and mitosis.
Kachaner, D., Pinson, X., El Kadhi, K. B., Normandin, K., Talje, L., Lavoie, H., Lepine, G., Carreno, S., Kwok, B. H., Hickson, G. R. and Archambault, V. (2014). Interdomain allosteric regulation of Polo kinase by Aurora B and Map205 is required for cytokinesis. J Cell Biol 207: 201-211. PubMed ID: 25332165
Drosophila Polo and its human orthologue Polo-like kinase 1 fulfill essential roles during cell division. Members of the Polo-like kinase (Plk) family contain an N-terminal kinase domain (KD) and a C-terminal Polo-Box domain (PBD), which mediates protein interactions. How Plks are regulated in cytokinesis is poorly understood. This study shows that phosphorylation of Polo by Aurora B is required for cytokinesis. This phosphorylation in the activation loop of the KD promotes the dissociation of Polo from the PBD-bound microtubule-associated protein Map205, which acts as an allosteric inhibitor of Polo kinase activity. This mechanism allows the release of active Polo from microtubules of the central spindle and its recruitment to the site of cytokinesis. Failure in Polo phosphorylation results in both early and late cytokinesis defects. Importantly, the antagonistic regulation of Polo by Aurora B and Map205 in cytokinesis reveals that interdomain allosteric mechanisms can play important roles in controlling the cellular functions of Plks.
Rosic, S., Kohler, F. and Erhardt, S. (2014). Repetitive centromeric satellite RNA is essential for kinetochore formation and cell division. J Cell Biol [Epub ahead of print]. PubMed ID: 25365994
Chromosome segregation requires centromeres on every sister chromatid to correctly form and attach the microtubule spindle during cell division. Even though centromeres are essential for genome stability, the underlying centromeric DNA is highly variable in sequence and evolves quickly. Epigenetic mechanisms are therefore thought to regulate centromeres. This study shows that the 359-bp repeat satellite III (SAT III), which spans megabases on the X chromosome of Drosophila melanogaster, produces a long noncoding RNA that localizes to centromeric regions of all major chromosomes. Depletion of SAT III RNA causes mitotic defects, not only of the sex chromosome but also in trans of all autosomes. It was furthermore found that SAT III RNA binds to the kinetochore component CENP-C, and is required for correct localization of the centromere-defining proteins CENP-A and CENP-C, as well as outer kinetochore proteins. In conclusion, these data reveal that SAT III RNA is an integral part of centromere identity, adding RNA to the complex epigenetic mark at centromeres in flies.
Ladurner, R., Bhaskara, V., Huis In 't Veld, P. J., Davidson, I. F., Kreidl, E., Petzold, G. and Peters, J. M. (2014). Cohesin's ATPase Activity Couples Cohesin Loading onto DNA with Smc3 Acetylation. Curr Biol 24: 2228-2237. PubMed ID: 25220052
Cohesin mediates sister chromatid cohesion by topologically entrapping sister DNA molecules inside its ring structure. Cohesin is loaded onto DNA by the Scc2/NIPBL-Scc4/MAU2-loading complex (see Drosophila Nipped B) in a manner that depends on the adenosine triphosphatase (ATPase) activity of cohesin's Smc1 and Smc3 subunits (see Ocampo-Hafalla and Frank Uhlmann, 2011; Cohesin loading and sliding). Subsequent cohesion establishment during DNA replication depends on Smc3 acetylation by Esco1 and Esco2 and on recruitment of sororin, which 'locks' cohesin on DNA by inactivating the cohesin release factor Wapl. Human cohesin ATPase mutants associate transiently with DNA in a manner that depends on the loading complex but cannot be stabilized on chromatin by depletion of Wapl. These mutants cannot be acetylated, fail to interact with sororin, and do not mediate cohesion. The absence of Smc3 acetylation in the ATPase mutants is not a consequence of their transient association with DNA but is directly caused by their inability to hydrolyze ATP because acetylation of wild-type cohesin also depends on ATP hydrolysis. These data indicate that cohesion establishment involves the following steps. First, cohesin transiently associates with DNA in a manner that depends on the loading complex. Subsequently, ATP hydrolysis by cohesin leads to entrapment of DNA and converts Smc3 into a state that can be acetylated. Finally, Smc3 acetylation leads to recruitment of sororin, inhibition of Wapl, and stabilization of cohesin on DNA. The finding that cohesin's ATPase activity is required for both cohesin loading and Smc3 acetylation raises the possibility that cohesion establishment is directly coupled to the reaction in which cohesin entraps DNA.
Saturday, November 15th
Demagny, H., Araki, T. and De Robertis, E. M. (2014). The tumor suppressor Smad4/DPC4 is regulated by phosphorylations that integrate FGF, Wnt, and TGF-beta signaling. Cell Rep 9: 688-700. PubMed ID: 25373906
Smad4 (see Drosophila Medea) is a major tumor suppressor currently thought to function constitutively in the transforming growth factor beta (TGF-beta)-signaling pathway. This study reports that Smad4 activity is directly regulated by the Wnt and fibroblast growth factor (FGF) pathways through GSK3 (see Drosophila Shaggy) and mitogen-activated protein kinase (MAPK; see Drosophila Rolled) phosphorylation sites. FGF activates MAPK, which primes three sequential GSK3 phosphorylations that generate a Wnt-regulated phosphodegron bound by the ubiquitin E3 ligase beta-TrCP (see Drosophila Slmb). In the presence of FGF, Wnt potentiates TGF-beta signaling by preventing Smad4 GSK3 phosphorylations that inhibit a transcriptional activation domain located in the linker region. When MAPK is not activated, the Wnt and TGF-beta signaling pathways remain insulated from each other. In Xenopus embryos, these Smad4 phosphorylations regulate germ-layer specification and Spemann organizer formation. The results show that three major signaling pathways critical in development and cancer are integrated at the level of Smad4.
Gujral, T. S., Chan, M., Peshkin, L., Sorger, P. K., Kirschner, M. W., and MacBeath, G. (2014). A noncanonical Frizzled2 pathway regulates epithelial-mesenchymal transition and metastasis. Cell 159: 844–856. Abstract
Wnt signaling plays a critical role in embryonic development, and genetic aberrations in this network have been broadly implicated in colorectal cancer. This study found that the Wnt receptor Frizzled2 (Fzd2; see Drosophila Frizzled) and its ligands Wnt5a/b (see Drosophila Wingless) are elevated in metastatic liver, lung, colon, and breast cancer cell lines and in high-grade tumors and that their expression correlates with markers of epithelial-mesenchymal transition (EMT). Pharmacologic and genetic perturbations reveal that Fzd2 drives EMT and cell migration through a previously unrecognized, noncanonical pathway that includes Fyn and Stat3 (see Drosophila Src42A and Stat92E). A gene signature regulated by this pathway predicts metastasis and overall survival in patients. An antibody was developed to Fzd2 that reduces cell migration and invasion and inhibits tumor growth and metastasis in xenografts. It is proposed that targeting this pathway could provide benefit for patients with tumors expressing high levels of Fzd2 and Wnt5a/b.
Shimizu, N., Ishitani, S., Sato, A., Shibuya, H. and Ishitani, T. (2014). Hipk2 and PP1c cooperate to maintain Dvl protein levels required for Wnt signal transduction. Cell Rep 8: 13904. PubMed ID: 25159144
The phosphoprotein Dishevelled (Dvl) is a common essential component of Wnt/beta-catenin and Wnt/planar cell polarity (PCP) signaling pathways. However, the regulation and significance of Dvl phosphorylation are not fully understood. This study shows that homeodomain-interacting protein kinase 2 (Hipk2; see Drosophila Hipk) facilitates protein phosphatase 1 catalytic subunit (PP1c)-mediated dephosphorylation of Dvl via its C-terminal domain and that this dephosphorylation blocks ubiquitination and consequent degradation mediated by the E3 ubiquitin ligase Itch, which targets the phosphorylated form of Dvl proteins. Inhibition of Hipk2 or PP1c function reduces Dvl protein levels and suppresses Wnt/beta-catenin and Wnt/PCP pathway-dependent events in mammalian cells and zebrafish embryos, suggesting that Hipk2 and PP1c are essential for maintaining Dvl protein levels that are sufficient to activate Wnt signaling. It was also shown that Wnt-3a, a Wnt/beta-catenin ligand, induces dissociation of the Dvl-Hipk2-PP1c complex and Dvl degradation under high-cell-density conditions. This regulation may be a negative feedback mechanism that fine-tunes Wnt/beta-catenin signaling.
Schieber, M. and Chandel, N. S. (2014). TOR signaling couples oxygen sensing to lifespan in C. elegans. Cell Rep 9: 9-15. PubMed ID: 25284791
Metazoans adapt to a low-oxygen environment (hypoxia) through activation of stress-response pathways. This study reports that transient hypoxia exposure extends lifespan in C. elegans through mitochondrial reactive oxygen species (ROS)-dependent regulation of the nutrient-sensing kinase target of rapamycin (TOR; see Drosophila Tor) and its upstream activator, RHEB-1 (see Drosophila Rheb). The increase in lifespan during hypoxia requires the intestinal GATA-type transcription factor ELT-2 (see Drosophila GATAe) downstream of TOR signaling. Using RNA sequencing (RNA-seq), this study describes an ELT-2-dependent hypoxia response that includes an intestinal glutathione S-transferase, GSTO-1, and it was uncovered that GSTO-1 is required for lifespan under hypoxia. These results indicate mitochondrial ROS-dependent TOR signaling integrates metabolic adaptations in order to confer survival under hypoxia.
Friday, November 14th
Lai, S. L. and Doe, C. Q. (2014). Transient nuclear Prospero induces neural progenitor quiescence. Elife 3 [Epub ahead of print]. PubMed ID: 25354199
Stem cells can self-renew, differentiate, or enter quiescence. Understanding how stem cells switch between these states is highly relevant for stem cell-based therapeutics. Drosophila neural progenitors (neuroblasts) have been an excellent model for studying self-renewal and differentiation, but quiescence remains poorly understood. This study shows that when neuroblasts enter quiescence, the differentiation factor Prospero is transiently detected in the neuroblast nucleus, followed by the establishment of a unique molecular profile lacking most progenitor and differentiation markers. The pulse of low level nuclear Prospero precedes entry into neuroblast quiescence even when the timing of quiescence is advanced or delayed by changing temporal identity factors. Furthermore, loss of Prospero prevents entry into quiescence, whereas a pulse of low level nuclear Prospero can drive proliferating larval neuroblasts into quiescence. It is proposed that Prospero levels distinguish three progenitor fates: absent for self-renewal, low for quiescence, and high for differentiation.
Wu, Y., Helt, J. C., Wexler, E., Petrova, I. M., Noordermeer, J. N., Fradkin, L. G. and Hing, H. (2014). Wnt5 and drl/ryk gradients pattern the Drosophila olfactory dendritic map. J Neurosci 34: 1496972. PubMed ID: 25378162
During development, dendrites migrate to their correct locations in response to environmental cues. The mechanisms of dendritic guidance are poorly understood. Recent work has shown that the Drosophila olfactory map is initially formed by the spatial segregation of the projection neuron (PN) dendrites in the developing antennal lobe (AL). This study reports that between 16 and 30 h after puparium formation, the PN dendrites undergo dramatic rotational reordering to achieve their final glomerular positions. During this period, a novel set of AL-extrinsic neurons express high levels of the Wnt5 protein and are tightly associated with the dorsolateral edge of the AL. Wnt5 forms a dorsolateral-high to ventromedial-low pattern in the antennal lobe neuropil. Loss of Wnt5 prevents the ventral targeting of the dendrites, whereas Wnt5 overexpression disrupts dendritic patterning. Drl/Ryk, a known Wnt5 receptor, is expressed in a dorsolateral-to-ventromedial (DL > VM) gradient by the PN dendrites. Loss of Drl in the PNs results in the aberrant ventromedial targeting of the dendrites, a defect that is suppressed by reduction in Wnt5 gene dosage. Conversely, overexpression of Drl in the PNs results in the dorsolateral targeting of their dendrites, an effect that requires Drl's cytoplasmic domain. It is proposed that Wnt5 acts as a repulsive guidance cue for the PN dendrites, whereas Drl signaling in the dendrites inhibits Wnt5 signaling. In this way, the precise expression patterns of Wnt5 and Drl orient the PN dendrites allowing them to target their final glomerular positions.
Caballero, I. M., Manuel, M. N., Molinek, M., Quintana-Urzainqui, I., Mi, D., Shimogori, T. and Price, D. J. (2014). Cell-autonomous repression of Shh by transcription factor Pax6 regulates diencephalic patterning by controlling the central diencephalic organizer. Cell Rep 8: 1405-1418. PubMed ID: 25176648
During development, region-specific patterns of regulatory gene expression are controlled by signaling centers that release morphogens providing positional information to surrounding cells. Regulation of signaling centers themselves is therefore critical. The size and the influence of a Shh-producing forebrain organizer (see Drosophila Hedgehog), the zona limitans intrathalamica (ZLI), are limited by Pax6 (see Drosophila Eyeless). By studying mouse chimeras, this study found that Pax6 acts cell autonomously to block Shh expression in cells around the ZLI. Immunoprecipitation and luciferase assays indicate that Pax6 can bind the Shh promoter and repress its function. An analysis of chimeras suggests that many of the regional gene expression pattern defects that occur in Pax6-/- diencephalic cells result from a non-cell-autonomous position-dependent defect of local intercellular signaling. Blocking Shh signaling in Pax6-/- mutants reverses major diencephalic patterning defects. It is concluded that Pax6's cell-autonomous repression of Shh expression around the ZLI is critical for many aspects of normal diencephalic patterning.
Wolff, T., Iyer, N. A. and Rubin, G. M. (2014). Neuroarchitecture and neuroanatomy of the Drosophila central complex: A GAL4-based dissection of protocerebral bridge neurons and circuits. J Comp Neurol [Epub ahead of print]. PubMed ID: 25380328
Insects exhibit an elaborate repertoire of behaviors in response to environmental stimuli. The central complex plays a key role in combining various modalities of sensory information with an insect's internal state and past experience to select appropriate responses. Progress has been made in understanding the broad spectrum of outputs from the central complex neuropils and circuits involved in numerous behaviors. Many resident neurons have also been identified. However, the specific roles of these intricate structures and the functional connections between them remain largely obscure. Significant gains rely on obtaining a comprehensive catalogue of the neurons and associated GAL4 lines that arborize within these brain regions, and on mapping neuronal pathways connecting these structures. Toward this end, small populations of neurons in the Drosophila melanogaster central complex were stochastically labeled using the multicolor flip-out technique and a catalogue was created of the neurons, their morphologies, trajectories, relative arrangements and corresponding GAL4 lines. This report focuses on one structure of the central complex, the protocerebral bridge, and identifies just 17 morphologically distinct cell types that arborize in this structure. This work also provides new insights into the anatomical structure of the four components of the central complex and its accessory neuropils. Most strikingly, the protocerebral bridge was found to contain 18 glomeruli, not 16, as previously believed. Revised wiring diagrams that take into account this updated architectural design are presented. This updated map of the Drosophila central complex will facilitate a deeper behavioral and physiological dissection of this sophisticated set of structures.
Thursday, November 13th
Mohammed, J., Siepel, A. and Lai, E. C. (2014). Diverse modes of evolutionary emergence and flux of conserved microRNA clusters. RNA. PubMed ID: 25332374
Many animal miRNA loci reside in genomic clusters that generate multicistronic primary-miRNA transcripts. While clusters that contain copies of the same miRNA hairpin are clearly products of local duplications, the evolutionary provenance of clusters with disparate members is less clear. Recently, it was proposed that essentially all such clusters in Drosophila derived from de novo formation of miRNA-like hairpins within existing miRNA transcripts, and that the maintenance of multiple miRNAs in such clusters was due to evolutionary hitchhiking on a major cluster member. However, this model seems at odds with the fact that many such miRNA clusters are composed of well-conserved miRNAs. In an effort to trace the birth and expansion of miRNA clusters that are presently well-conserved across Drosophilids, a broad swath of metazoan species was analyzed, with particular emphasis on arthropod evolution. Beyond duplication and de novo birth, this study highlighted a diversity of modes that contribute to miRNA evolution, including neofunctionalization of miRNA copies, fissioning of locally duplicated miRNA clusters, miRNA deletion, and miRNA cluster expansion via the acquisition and/or neofunctionalization of miRNA copies from elsewhere in the genome. In particular, it is suggested that miRNA clustering by acquisition represents an expedient strategy to bring cohorts of target genes under coordinate control by miRNAs that had already been individually selected for regulatory impact on the transcriptome.
Foronda, D., Weng, R., Verma, P., Chen, Y. W. and Cohen, S. M. (2014). Coordination of insulin and Notch pathway activities by microRNA miR-305 mediates adaptive homeostasis in the intestinal stem cells of the Drosophila gut. Genes Dev 28: 2421-2431. PubMed ID: 25367037
Homeostasis of the intestine is maintained by dynamic regulation of a pool of intestinal stem cells. The balance between stem cell self-renewal and differentiation is regulated by the Notch and insulin signaling pathways. Dependence on the insulin pathway places the stem cell pool under nutritional control, allowing gut homeostasis to adapt to environmental conditions. This study presents evidence that miR-305 is required for adaptive homeostasis of the gut. miR-305 regulates the Notch and insulin pathways in the intestinal stem cells. Notably, miR-305 expression in the stem cells is itself under nutritional control via the insulin pathway. This link places regulation of Notch pathway activity under nutritional control. These findings provide a mechanism through which the insulin pathway controls the balance between stem cell self-renewal and differentiation that is required for adaptive homeostasis in the gut in response to changing environmental conditions.
Choi, Y. J., Lai, W. S., Fedic, R., Stumpo, D. J., Huang, W., Li, L., Perera, L., Brewer, B. Y., Wilson, G. M., Mason, J. M. and Blackshear, P. J. (2014). The Drosophila Tis11 protein and its effects on mRNA expression in flies. J Biol Chem [Epub ahead of print]. PubMed ID: 25342740
Members of the mammalian tristetraprolin (TTP) family of CCCH tandem zinc finger proteins can bind to certain AU-rich elements (AREs) in mRNAs, leading to their deadenylation and destabilization. Mammals express three or four members of this family, but Drosophila melanogaster and other insects appear to contain a single gene, Tis11. Recombinant Drosophila Tis11 protein was found to bind to ARE-containing RNA oligonucleotides with low nanomolar affinity. Remarkably, co-expression in mammalian cells with target RNAs demonstrated that Tis11 could promote destabilization of ARE-containing mRNAs, and that this was partially dependent on a conserved C-terminal sequence resembling the mammalian NOT1 binding domain. Drosophila Tis11 promoted both deadenylation and decay of a target transcript in this heterologous cell system. Chromosome deletion/duplication and P element insertion were used to produce two types of Tis11 deficiency in adult flies, both of which were viable and fertile. To address the hypothesis that Tis11 deficiency would lead to the abnormal accumulation of potential target transcripts, gene expressionin adult flies was analyzed by deep mRNA sequencing. 69 transcripts from 56 genes were identified that were significantly up-regulated more than 1.5-fold in both types of Tis11-deficient flies. Ten of the up-regulated transcripts encoded probable proteases, but many other functional classes of proteins were represented. Many of the up-regulated transcripts contained potential binding sites for TTP family member proteins that were conserved in other Drosophila species. Tis11 is thus an ARE-binding, mRNA-destabilizing protein that may play a role in post-transcriptional gene expression in Drosophila and other insects.
Zearfoss, N. R., Deveau, L. M., Clingman, C. C., Schmidt, E., Johnson, E. S., Massi, F. and Ryder, S. P. (2014). A conserved three-nucleotide core motif defines Musashi RNA-binding specificity. J Biol Chem. PubMed ID: 25368328
Musashi (MSI) family proteins control cell proliferation and differentiation in many biological systems. They are over-expressed in tumors of several origins, and their expression level correlates with poor prognosis. MSI proteins control gene expression by binding RNA and regulating its translation. They contain two RNA recognition motif (RRM) domains, which recognize a defined sequence element. The relative contribution of each nucleotide to the binding affinity and specificity is unknown. This study analyzed the binding specificity of three MSI family RRM domains using a quantitative fluorescence anisotropy assay. It was found that the core element driving recognition is the sequence UAG. Nucleotides outside of this motif have a limited contribution to binding free energy. For mouse MSI1, recognition is determined by the first of the two RRM domains. The second RRM adds affinity but does not contribute to binding specificity. In contrast, the recognition element for Drosophila Msi is more extensive than the mouse homolog, suggesting functional divergence. The short nature of the binding determinant suggests that protein-RNA affinity alone is insufficient to drive target selection by MSI family proteins.
Nagarajan, S. and Grewal, S. S. (2014). An investigation of nutrient-dependent mRNA translation in Drosophila larvae. Biol Open [Epub ahead of print]. PubMed ID: 25305039
The larval period of the Drosophila life cycle is characterized by immense growth. In nutrient rich conditions, larvae increase in mass approximately two hundred-fold in five days. However, upon nutrient deprivation, growth is arrested. The prevailing view is that dietary amino acids drive this larval growth by activating the conserved insulin/PI3 kinase and Target of rapamycin (TOR) pathways and promoting anabolic metabolism. One key anabolic process is protein synthesis. However, few studies have attempted to measure mRNA translation during larval development or examine the signaling requirements for nutrient-dependent regulation. This work addresses this issue. Using polysome analyses, it was observed that starvation rapidly (within thirty minutes) decreased larval mRNA translation, with a maximal decrease at 6-18 hours. By analyzing individual genes, it was observed that nutrient-deprivation led to a general reduction in mRNA translation, regardless of any starvation-mediated changes (increase or decrease) in total transcript levels. Although sugars and amino acids are key regulators of translation in animal cells and are the major macronutrients in the larval diet, this study found that they alone were not sufficient to maintain mRNA translation in larvae. The insulin/PI3 kinase and TOR pathways are widely proposed as the main link between nutrients and mRNA translation in animal cells. However, this study found that genetic activation of PI3K and TOR signaling, or regulation of two effectors - 4EBP and S6K - could not prevent the starvation-mediated translation inhibition. Similarly, it was shown that the nutrient stress-activated eIF2α kinases, GCN2 and PERK, were not required for starvation-induced inhibition of translation in larvae. These findings indicate that nutrient control of mRNA translation in larvae is more complex than simply amino acid activation of insulin and TOR signaling.
Wednesday, November 12th
Liu, Z., Merkurjev, D., Yang, F., Li, W., Oh, S., Friedman, M. J., Song, X., Zhang, F., Ma, Q., Ohgi, K. A., Krones, A. and Rosenfeld, M. G. (2014). Enhancer activation requires trans-recruitment of a mega transcription factor complex. Cell 159: 358-373. PubMed ID: 25303530
Enhancers provide critical information directing cell-type-specific transcriptional programs, regulated by binding of signal-dependent transcription factors and their associated cofactors. This study reports that the most strongly activated estrogen (E2)-responsive enhancers are characterized by trans-recruitment and in situ assembly of a large 1-2 MDa complex of diverse DNA-binding transcription factors by ERalpha at ERE-containing enhancers. Enhancers recruiting these factors are referred to as mega transcription factor-bound in trans (MegaTrans) enhancers. The MegaTrans complex is a signature of the most potent functional enhancers and is required for activation of enhancer RNA transcription and recruitment of coactivators, including p300 (see Drosophila CBP) and Med1. The MegaTrans complex functions, in part, by recruiting specific enzymatic machinery, exemplified by DNA-dependent protein kinase. Thus, MegaTrans-containing enhancers represent a cohort of functional enhancers that mediate a broad and important transcriptional program and provide a molecular explanation for transcription factor clustering and hotspots noted in the genome.
Dowen, J. M., Fan, Z. P., Hnisz, D., Ren, G., Abraham, B. J., Zhang, L. N., Weintraub, A. S., Schuijers, J., Lee, T. I., Zhao, K. and Young, R. A. (2014). Control of cell identity genes occurs in insulated neighborhoods in Mammalian chromosomes. Cell 159: 374-387. PubMed ID: 25303531
The pluripotent state of embryonic stem cells (ESCs) is produced by active transcription of genes that control cell identity and repression of genes encoding lineage-specifying developmental regulators. This study uses ESC cohesin ChIA-PET data to identify the local chromosomal structures at both active and repressed genes across the genome. The results produce a map of enhancer-promoter interactions and reveal that super-enhancer-driven genes generally occur within chromosome structures that are formed by the looping of two interacting CTCF sites co-occupied by cohesin (see Drosophila Cohesin). These looped structures form insulated neighborhoods whose integrity is important for proper expression of local genes. It was also found that repressed genes encoding lineage-specifying developmental regulators occur within insulated neighborhoods. These results provide insights into the relationship between transcriptional control of cell identity genes and control of local chromosome structure.
Magbanua, J. P., Runneburger, E., Russell, S. and White, R. (2014). A variably occupied CTCF binding site in the Ultrabithorax gene in the Drosophila Bithorax Complex. Mol Cell Biol [Epub ahead of print]. PubMed ID: 25368383
Although the majority of genomic binding sites for the insulator protein CTCF are constitutively occupied, a subset show variably occupancy. Such variable sites provide an opportunity to assess context-specific CTCF functions in gene regulation. This study has identified a variably occupied CTCF site in the Drosophila Ultrabithorax (Ubx) gene. This site is occupied in tissues where Ubx is active (third thoracic leg imaginal disc) but is not bound in tissues where the Ubx gene is repressed (first thoracic leg imaginal disc). Using chromatin conformation capture this site was shown to preferentially interact with the Ubx promoter region in the active state. The site lies close to Ubx enhancer elements and is also close to the locations of several gypsy transposon insertions that disrupt Ubx expression, leading to the bx mutant phenotype. Gypsy insertions carry the Su(Hw)-dependent gypsy insulator and were found to affect both CTCF binding at the variable site and the chromatin topology. This suggests that insertion of the gypsy insulator in this region interferes with CTCF function and supports a model for the normal function of the variable CTCF site as a chromatin loop facilitator, promoting interaction between Ubx enhancers and the Ubx transcription start site.
Podsiadlo, A., Wrzesien, M., Paja, W., Rudnicki, W. and Wilczynski, B. (2013). Active enhancer positions can be accurately predicted from chromatin marks and collective sequence motif data. BMC Syst Biol 7 Suppl 6: S16. PubMed ID: 24565409
Transcriptional regulation in multi-cellular organisms is a complex process involving multiple modular regulatory elements for each gene. Building whole-genome models of transcriptional networks requires mapping all relevant enhancers and then linking them to target genes. Previous methods of enhancer identification based either on sequence information or on epigenetic marks have different limitations stemming from incompleteness of each of these datasets taken separately. This work presents a new approach for discovery of regulatory elements based on the combination of sequence motifs and epigenetic marks measured with ChIP-Seq. The method uses supervised learning approaches to train a model describing the dependence of enhancer activity on sequence features and histone marks. The results indicate that using combination of features provides superior results to previous approaches based on either one of the datasets. While histone modifications remain the dominant feature for accurate predictions, the models based on sequence motifs have advantages in their general applicability to different tissues. Additionally, the relevance of different sequence motifs in prediction accuracy was assessed showing that even tissue-specific enhancer activity depends on multiple motifs. Based on these results, it is concluded that it is worthwhile to include sequence motif data into computational approaches to active enhancer prediction and also that classifiers trained on a specific set of enhancers can generalize with significant accuracy beyond the training set.
Tuesday, November 11th
Ma, H., Groth, R. D., Cohen, S. M., Emery, J. F., Li, B., Hoedt, E., Zhang, G., Neubert, T. A. and Tsien, R. W. (2014). γCaMKII shuttles Ca(2+)/CaM to the nucleus to trigger CREB phosphorylation and gene expression. Cell 159: 281-294. PubMed ID: 25303525
Activity-dependent CREB (see Drosophila CrebB) phosphorylation and gene expression are critical for long-term neuronal plasticity. Local signaling at voltage gated CaV1 channels triggers these events, but how information is relayed onward to the nucleus remains unclear. This study reports a mechanism that mediates long-distance communication within cells: a shuttle that transports Ca(2+)/calmodulin (see Drosophila Calmodulin) from the surface membrane to the nucleus. This study shows that the shuttle protein is γCaMKII (see Drosophila CaMKII), its phosphorylation at Thr287 by βCaMKII protects the Ca(2+)/CaM signal, and CaN (see Drosophila Calcineurin) triggers its nuclear translocation. Both betaCaMKII and CaN act in close proximity to CaV1 channels, supporting their dominance, whereas γCaMKII operates as a carrier, not as a kinase. Upon arrival within the nucleus, Ca(2+)/CaM activates CaMKK and its substrate CaMKIV, the CREB kinase. This mechanism resolves long-standing puzzles about CaM/CaMK-dependent signaling to the nucleus. The significance of the mechanism is emphasized by dysregulation of CaV1, γCaMKII, βCaMKII, and CaN in multiple neuropsychiatric disorders.
Peng, M., Yin, N. and Li, M. O. (2014). Sestrins function as guanine nucleotide dissociation inhibitors for Rag GTPases to control mTORC1 signaling. Cell 159: 122-133. PubMed ID: 25259925
Mechanistic target of rapamycin complex 1 (mTORC1) (see Drosophila Tor) integrates diverse environmental signals to control cellular growth and organismal homeostasis. In response to nutrients, Rag GTPases (see Drosophila RagA-B) recruit mTORC1 to the lysosome to be activated, but how Rags are regulated remains incompletely understood. This study shows that Sestrins (see Drosophila Sestrin) bind to the heterodimeric RagA/B-RagC/D GTPases, and function as guanine nucleotide dissociation inhibitors (GDIs) for RagA/B. Sestrin overexpression inhibits amino-acid-induced Rag guanine nucleotide exchange and mTORC1 translocation to the lysosome. Mutation of the conserved GDI motif creates a dominant-negative form of Sestrin that renders mTORC1 activation insensitive to amino acid deprivation, whereas a cell-permeable peptide containing the GDI motif inhibits mTORC1 signaling. Mice deficient in all Sestrins exhibit reduced postnatal survival associated with defective mTORC1 inactivation in multiple organs during neonatal fasting. These findings reveal a nonredundant mechanism by which the Sestrin family of GDIs regulates the nutrient-sensing Rag GTPases to control mTORC1 signaling.
Siegelbaum, S. A. (2014). Reelin signaling specifies the molecular identity of the pyramidal neuron distal dendritic compartment. Cell 158: 1335-1347. PubMed ID: 25201528
The apical dendrites of many neurons contain proximal and distal compartments that receive synaptic inputs from different brain regions. These compartments also contain distinct complements of ion channels that enable the differential processing of their respective synaptic inputs, making them functionally distinct. At present, the molecular mechanisms that specify dendritic compartments are not well understood. This study reports that the extracellular matrix protein Reelin, acting through its downstream, intracellular Dab1 (see Drosophila Dab) and Src family (see Drosophila Src64B) tyrosine kinase signaling cascade, is essential for establishing and maintaining the molecular identity of the distal dendritic compartment of cortical pyramidal neurons. Reelin signaling is required for the striking enrichment of HCN1 and GIRK1 channels in the distal tuft dendrites of both hippocampal CA1 and neocortical layer 5 pyramidal neurons, where the channels actively filter inputs targeted to these dendritic domains.
Doherty, J., Sheehan, A. E., Bradshaw, R., Fox, A. N., Lu, T. Y. and Freeman, M. R. (2014). PI3K signaling and Stat92E converge to modulate glial responsiveness to axonal injury. PLoS Biol 12: e1001985. PubMed ID: 25369313
Glial cells are exquisitely sensitive to neuronal injury but mechanisms by which glia establish competence to respond to injury, continuously gauge neuronal health, and rapidly activate reactive responses remain poorly defined. This study shows glial PI3K signaling in the uninjured brain regulates baseline levels of Draper, a receptor essential for Drosophila glia to sense and respond to axonal injury. After injury, Draper levels are up-regulated through a Stat92E-modulated, injury-responsive enhancer element within the draper gene. Surprisingly, canonical JAK/STAT signaling does not regulate draper expression. Rather, injury-induced draper activation is downstream of the Draper/Src42a/Shark/Rac1 engulfment signaling pathway. Thus, PI3K signaling and Stat92E are critical in vivo regulators of glial responsiveness to axonal injury. Evidence is provided for a positive auto-regulatory mechanism whereby signaling through the injury-responsive Draper receptor leads to Stat92E-dependent, transcriptional activation of the draper gene. It is proposed that Drosophila glia use this auto-regulatory loop as a mechanism to adjust their reactive state following injury.
Monday, November 10th
Das, R., Sebo, Z., Pence, L. and Dobens, L. L. (2014). Drosophila Tribbles antagonizes insulin signaling-mediated growth and metabolism via interactions with Akt kinase. PLoS One 9: e109530. PubMed ID: 25329475
Drosophila Tribbles (Trbl) is the founding member of the Trib family of kinase-like docking proteins that modulate cell signaling during proliferation, migration and growth. In a wing misexpression screen for Trbl interacting proteins, the Ser/Thr protein kinase Akt1. Given the central role of Akt1 in insulin signaling, the function of Trbl was tested in larval fat body, a tissue where rapid increases in size are exquisitely sensitive to insulin/insulin-like growth factor levels. Consistent with a role in antagonizing insulin-mediated growth, trbl RNAi knockdown in the fat body increased cell size, advanced the timing of pupation and increased levels of circulating triglyceride. Complementarily, overexpression of Trbl reduced fat body cell size, decreased overall larval size, delayed maturation and lowered levels of triglycerides, while circulating glucose levels increased. The conserved Trbl kinase domain is required for function in vivo and for interaction with Akt in a yeast two-hybrid assay. Consistent with direct regulation of Akt, overexpression of Trbl in the fat body decreased levels of activated Akt (pSer505-Akt) while misexpression of trbl RNAi increased phospho-Akt levels, and neither treatment affected total Akt levels. Trbl misexpression effectively suppressed Akt-mediated wing and muscle cell size increases and reduced phosphorylation of the Akt target FoxO (pSer256-FoxO). Taken together, these data show that Drosophila Trbl has a conserved role to bind Akt and block Akt-mediated insulin signaling, and implicate Trib proteins as novel sites of signaling pathway integration that link nutrient availability with cell growth and proliferation.
Jiang, K., Liu, Y., Fan, J., Epperly, G., Gao, T., Jiang, J. and Jia, J. (2014). Hedgehog-regulated atypical PKC promotes phosphorylation and activation of Smoothened and Cubitus interruptus in Drosophila. Proc Natl Acad Sci U S A. PubMed ID: 25349414
Smoothened (Smo) is essential for transduction of the Hedgehog (Hh) signal in both insects and vertebrates. Cell surface/cilium accumulation of Smo is thought to play an important role in Hh signaling, but how the localization of Smo is controlled remains poorly understood. This study demonstrates that atypical PKC (aPKC) regulates Smo phosphorylation and basolateral accumulation in Drosophila wings. Inactivation of aPKC by either RNAi or a mutation inhibits Smo basolateral accumulation and attenuates Hh target gene expression. In contrast, expression of constitutively active aPKC elevates basolateral accumulation of Smo and promotes Hh signaling. The aPKC-mediated phosphorylation of Smo at Ser680 promotes Ser683 phosphorylation by casein kinase 1 (CK1), and these phosphorylation events elevate Smo activity in vivo. Moreover, aPKC has an additional positive role in Hh signaling by regulating the activity of Cubitus interruptus (Ci) through phosphorylation of the Zn finger DNA-binding domain. Finally, the expression of aPKC is up-regulated by Hh signaling in a Ci-dependent manner. These findings indicate a direct involvement of aPKC in Hh signaling beyond its role in cell polarity.
Ishio, A., Sasamura, T., Ayukawa, T., Kuroda, J., Ishikawa, H. O., Aoyama, N., Matsumoto, K., Gushiken, T., Okajima, T., Yamakawa, T. and Matsuno, K. (2014). O-fucose monosaccharide of Drosophila Notch has a temperature-sensitive function and cooperates with O-glucose glycan in Notch transport and Notch signaling activation. J Biol Chem [Epub ahead of print]. PubMed ID: 25378397
Notch (N) is a transmembrane receptor that mediates the cell-cell interactions necessary for many cell-fate decisions. N has many epidermal growth factor-like repeats that are O-fucosylated by the protein O-fucosyltransferase 1 (O-fut1), and the O-fut1 gene is essential for N signaling. However, the role of the monosaccharide O-fucose on N is unclear, because O-fut1 also appears to have O-fucosyltransferase activity-independent functions, including as an N-specific chaperon. Such an enzymatic activity-independent function could account for the essential role of O-fut1 in N signaling. To evaluate the role of the monosaccharide O-fucose modification in N signaling, this study generated a knock-in mutant of O-fut1 (O-fut1R245A knock-in), which expresses a mutant protein that lacks O-fucosyltransferase activity, but maintains the N-specific chaperon activity. Using O-fut1R245A knock-in and other gene mutations that abolish the O-fucosylation of N, it was found that the monosaccharide O-fucose modification of N has a temperature-sensitive function that is essential for N signaling. The O-fucose monosaccharide and O-glucose glycan modification, catalyzed by Rumi, function redundantly in the activation of N signaling. It was also showm that the redundant function of these two modifications is responsible for the presence of N at the cell surface. These findings elucidate how different forms of glycosylation on a protein can influence the protein's functions.
Aleman, A., Rios, M., Juarez, M., Lee, D., Chen, A. and Eivers, E. (2014). Mad linker phosphorylations control the intensity and range of the BMP-activity gradient in developing Drosophila tissues. Sci Rep 4: 6927. PubMed ID: 25377173L
The BMP ligand Dpp, operates as a long range morphogen to control many important functions during Drosophila development from tissue patterning to growth. The BMP signal is transduced intracellularly via C-terminal phosphorylation of the BMP transcription factor Mad, which forms an activity gradient in developing embryonic tissues. This study shows that Cyclin dependent kinase 8 and Shaggy phosphorylate three Mad linker serines. Linker phosphorylations control the peak intensity and range of the BMP signal across rapidly developing embryonic tissues. Shaggy knockdown broadened the range of the BMP-activity gradient and increased high threshold target gene expression in the early embryo, while expression of a Mad linker mutant in the wing disc resulted in enhanced levels of C-terminally phosphorylated Mad, a 30% increase in wing tissue, and elevated BMP target genes. In conclusion, these results describe how Mad linker phosphorylations work to control the peak intensity and range of the BMP signal in rapidly developing Drosophila tissues.
Hugosson, F., Sjogren, C., Birve, A., Hedlund, L., Eriksson, T. and Palmer, R. H. (2014). The Drosophila Midkine/Pleiotrophin homologues Miple1 and Miple2 affect adult lifespan but are dispensable for Alk signaling during embryonic gut formation. PLoS One 9: e112250. PubMed ID: 25380037
Midkine (MDK) and Pleiotrophin (PTN) are small heparin-binding cytokines with closely related structures. The Drosophila genome harbours two genes encoding members of the MDK/PTN family of proteins, known as miple1 and miple2. The role of Miple proteins was investigated in vivo, in particular with regard to their proposed role as ligands for the Alk receptor tyrosine kinase (RTK). This study shows that Miple proteins are neither required to drive Alk signaling during Drosophila embryogenesis, nor are they essential for development in the fruit fly. Additionally it was shown that neither MDK nor PTN can activate hALK in vivo when ectopically co-expressed in the fly. In conclusion, the data suggest that Alk is not activated by MDK/PTN related growth factors Miple1 and Miple 2 in vivo.
Sunday, November 9th
Mosca, T. J. and Luo, L. (2014). Synaptic organization of the Drosophila antennal lobe and its regulation by the Teneurins. Elife 3. PubMed ID: 25310239
Understanding information flow through neuronal circuits requires knowledge of their synaptic organization. This study utilized fluorescent pre- and postsynaptic markers to map synaptic organization in the Drosophila antennal lobe, the first olfactory processing center. Olfactory receptor neurons (ORNs) produce a constant synaptic density across different glomeruli. Each ORN within a class contributes nearly identical active zone number. Active zones from ORNs, projection neurons (PNs), and local interneurons have distinct subglomerular and subcellular distributions. The correct number of ORN active zones and PN acetylcholine receptor clusters requires the Teneurins, conserved transmembrane proteins involved in neuromuscular synapse organization and synaptic partner matching. Ten-a acts in ORNs to organize presynaptic active zones via the spectrin cytoskeleton. Ten-m acts in PNs autonomously to regulate acetylcholine receptor cluster number and transsynaptically to regulate ORN active zone number. These studies advanced the ability to assess synaptic architecture in complex CNS circuits and their underlying molecular mechanisms.
Nakayama, M., Matsushita, F. and Hama, C. (2014). The matrix protein Hikaru genki localizes to cholinergic synaptic clefts and regulates postsynaptic organization in the Drosophila brain. J Neurosci 34: 13872-13877. PubMed ID: 25319684
The synaptic cleft, a crucial space involved in neurotransmission, is filled with extracellular matrix that serves as a scaffold for synaptic differentiation. However, little is known about the proteins present in the matrix and their functions in synaptogenesis, especially in the CNS. This study reports that Hikaru genki (Hig), a secreted protein with an Ig motif and complement control protein domains, localizes specifically to the synaptic clefts of cholinergic synapses in the Drosophila CNS. The data indicate that this specific localization is achieved by capture of secreted Hig in synaptic clefts, even when it is ectopically expressed in glia. In the absence of Hig, the cytoskeletal scaffold protein DLG accumulats abnormally in cholinergic postsynapses, and the synaptic distribution of acetylcholine receptor (AchR) subunits Dalpha6 and Dalpha7 significantly decreased. hig mutant flies consistently exhibited resistance to the AchR agonist spinosad, which causes lethality by specifically activating the Dalpha6 subunit, suggesting that loss of Hig compromises the cholinergic synaptic activity mediated by Dalpha6. These results indicate that Hig is a specific component of the synaptic cleft matrix of cholinergic synapses and regulates their postsynaptic organization in the CNS.
Sakuma, C., Kawauchi, T., Haraguchi, S., Shikanai, M., Yamaguchi, Y., Gelfand, V. I., Luo, L., Miura, M. and Chihara, T. (2014). Drosophila Strip serves as a platform for early endosome organization during axon elongation. Nat Commun 5: 5180. PubMed ID: 25312435
Early endosomes are essential for regulating cell signalling and controlling the amount of cell surface molecules during neuronal morphogenesis. Early endosomes undergo retrograde transport (clustering) before their homotypic fusion. Small GTPase Rab5 is known to promote early endosomal fusion, but the mechanism linking the transport/clustering with Rab5 activity is unclear. This study showed that Drosophila Strip is a key regulator for neuronal morphogenesis. Strip knockdown disturbs the early endosome clustering, and Rab5-positive early endosomes become smaller and scattered. Strip genetically and biochemically interacts with both Glued (the regulator of dynein-dependent transport) and Sprint (the guanine nucleotide exchange factor for Rab5), suggesting that Strip is a molecular linker between retrograde transport and Rab5 activation. Overexpression of an active form of Rab5 in strip-mutant neurons suppresses the axon elongation defects. Thus, Strip acts as a molecular platform for the early endosome organization that has important roles in neuronal morphogenesis.
Kunduri, G., Yuan, C., Parthibane, V., Nyswaner, K. M., Kanwar, R., Nagashima, K., Britt, S. G., Mehta, N., Kotu, V., Porterfield, M., Tiemeyer, M., Dolph, P. J., Acharya, U. and Acharya, J. K. (2014). Phosphatidic acid phospholipase A1 mediates ER-Golgi transit of a family of G protein-coupled receptors. J Cell Biol 206: 79-95. PubMed ID: 25002678
The coat protein II (COPII)-coated vesicular system transports newly synthesized secretory and membrane proteins from the endoplasmic reticulum (ER) to the Golgi complex. Recruitment of cargo into COPII vesicles requires an interaction of COPII proteins either with the cargo molecules directly or with cargo receptors for anterograde trafficking. This study shows that cytosolic phosphatidic acid phospholipase A1 (PAPLA1) interacts with COPII protein family members and is required for the transport of Rh1 (rhodopsin 1), an N-glycosylated G protein-coupled receptor (GPCR), from the ER to the Golgi complex. In papla1 mutants, in the absence of transport to the Golgi, Rh1 is aberrantly glycosylated and is mislocalized. These defects lead to decreased levels of the protein and decreased sensitivity of the photoreceptors to light. Several GPCRs, including other rhodopsins and Bride of sevenless, are similarly affected. These findings show that a cytosolic protein is necessary for transit of selective transmembrane receptor cargo by the COPII coat for anterograde trafficking.
Rosenbaum, E. E., Vasiljevic, E., Cleland, S. C., Flores, C. and Colley, N. J. (2014). The Gos28 SNARE mediates intra-golgi transport of Rhodopsin and is required for photoreceptor survival. J Biol Chem . [Epub ahead of print]. PubMed ID: 25261468
SNARE proteins play indispensable roles in membrane fusion events in many cellular processes, including synaptic transmission and protein trafficking. This study characterize the Golgi SNARE protein, Gos28, and its role in Rhodopsin (Rh1) transport through Drosophila photoreceptors. Mutations in gos28 lead to defective Rh1 trafficking and retinal degeneration. This study has pinpointed a role for Gos28 in the intra-Golgi transport of Rh1, downstream from alpha-mannosidase-II in the medial-Golgi. The necessity of key residues in the Gos28 SNARE motif was confirmed, and it was demonstrated that its transmembrane domain is not required for vesicle fusion, consistent with Gos28 functioning as a t-SNARE for Rh1 transport. Finally, human Gos28 was shown to rescue both the Rh1 trafficking defects and retinal degeneration in Drosophila gos28 mutants, demonstrating the functional conservation of these proteins. These results identify Gos28 as an essential SNARE protein in Drosophila photoreceptors and provide mechanistic insights into the role of SNAREs in neurodegenerative disease.
Saturday, November 8th
Cordoba, S. and Estella, C. (2014). The bHLH-PAS transcription factor Dysfusion regulates tarsal joint formation in response to Notch activity during Drosophila leg development. PLoS Genet 10: e1004621. PubMed ID: 25329825
A characteristic of all arthropods is the presence of flexible structures called joints that connect all leg segments. Drosophila legs include two types of joints: the proximal or 'true' joints that are motile due to the presence of muscle attachment and the distal joints that lack musculature. These joints are not only morphologically, functionally and evolutionarily different, but also the morphogenetic program that forms them is distinct. Development of both proximal and distal joints requires Notch activity; however, it is still unknown how this pathway can control the development of such homologous although distinct structures. This study shows that the bHLH-PAS transcription factor encoded by the gene dysfusion (dys), is expressed and absolutely required for tarsal joint development while it is dispensable for proximal joints. In the presumptive tarsal joints, Dys regulates the expression of the pro-apoptotic genes reaper and head involution defective and the expression of the RhoGTPases modulators, RhoGEf2 and RhoGap71E, thus directing key morphogenetic events required for tarsal joint development. When ectopically expressed, dys is able to induce some aspects of the morphogenetic program necessary for distal joint development such as fold formation and programmed cell death. This novel Dys function depends on its obligated partner Tango to activate the transcription of target genes. A dedicated dys cis-regulatory module was identified that regulates dys expression in the tarsal presumptive leg joints through direct Su(H) binding. All these data place dys as a key player downstream of Notch, directing distal versus proximal joint morphogenesis.
Chanut-Delalande, H., Hashimoto, Y., Pelissier-Monier, A., Spokony, R., Dib, A., Kondo, T., Bohere, J., Niimi, K., Latapie, Y., Inagaki, S., Dubois, L., Valenti, P., Polesello, C., Kobayashi, S., Moussian, B., White, K. P., Plaza, S., Kageyama, Y. and Payre, F. (2014). Pri peptides are mediators of ecdysone for the temporal control of development. Nat Cell Biol 16: 1035-1044. PubMed ID: 25344753
Animal development fundamentally relies on the precise control, in space and time, of genome expression. Whereas a wealth of information is available about spatial patterning, the mechanisms underlying temporal control remain poorly understood. This study shows that Pri peptides (see Tarsal-less), encoded by small open reading frames, are direct mediators of the steroid hormone ecdysone for the timing of developmental programs in Drosophila. A previously uncharacterized enzyme of ecdysone biosynthesis, Glutathione S transferase E14 (GstE14), was identified, and ecdysone was found to trigger pri expression to define the onset of epidermal trichome development, through post-translational control of the Shavenbaby transcription factor. Manipulating pri expression is sufficient to either put on hold or induce premature differentiation of trichomes. Furthermore, it was found that ecdysone-dependent regulation of pri is not restricted to epidermis and occurs over various tissues and times. Together, these findings provide a molecular framework to explain how systemic hormonal control coordinates specific programs of differentiation with developmental timing.
Morishita, K., Ozasa, F., Eguchi, K., Yoshioka, Y., Yoshida, H., Hiai, H. and Yamaguchi, M. (2014). Drosophila DOCK family protein Sponge regulates the JNK pathway during thorax development. Cell Struct Funct. PubMed ID: 25311449
The dedicator of cytokinesis (DOCK) family proteins that are conserved in a wide variety of species are known as DOCK1-DOCK11 in mammals. Sponge (Spg) is a Drosophila counterpart to the mammalian DOCK3. Specific knockdown of spg by pannier-GAL4 or apterous-GAL4 driver in wing discs induced split thorax phenotype in adults. Reduction of the Drosophila c-Jun N-terminal kinase (JNK), basket (bsk) gene dose enhanced the spg knockdown-induced phenotype. Conversely, overexpression of bsk suppressed the split thorax phenotype. Monitoring JNK activity in the wing imaginal discs by immunostaining with anti-phosphorylated JNK (anti-pJNK) antibody together with examination of lacZ expression in a puckered-lacZ enhancer trap line revealed the strong reduction of the JNK activity in the spg knockdown clones. This was further confirmed by Western immunoblot analysis of extracts from wing discs of spg knockdown fly with anti-pJNK antibody. Furthermore, the Duolink in situ Proximity Ligation Assay method detected interaction signals between Spg and Rac1 in the wing discs. Taken together, these results indicate Spg positively regulates JNK pathway that is required for thorax development and the regulation is mediated by interaction with Rac1.
Hua, Z. L., Chang, H., Wang, Y., Smallwood, P. M. and Nathans, J. (2014). Partial interchangeability of Fz3 and Fz6 in tissue polarity signaling for epithelial orientation and axon growth and guidance. Development 141: 3944-3954. PubMed ID: 25294940
In mammals, a set of anatomically diverse polarity processes - including axon growth and guidance, hair follicle orientation, and stereociliary bundle orientation in inner ear sensory hair cells - appear to be mechanistically related, as judged by their dependence on vertebrate homologues of core tissue polarity/planar cell polarity (PCP) genes in Drosophila. To explore more deeply the mechanistic similarities between different polarity processes, this study has determined the extent to which frizzled 3 (Fz3; see Drosophila Frizzled) can rescue the hair follicle and Merkel cell polarity defects in frizzled 6-null (Fz6-/-) mice, and, reciprocally, the extent to which Fz6 can rescue the axon growth and guidance defects in Fz3-/- mice. These experiments reveal full rescue of the Fz6-/- phenotype by Fz3 and partial rescue of the Fz3-/- phenotype by Fz6, implying that these two proteins are likely to act in a conserved manner in these two contexts. Stimulated by these observations, additional anatomical structures were sought that exhibit macroscopic polarity and that might plausibly use Fz3 and/or Fz6 signaling. This search has revealed a hitherto unappreciated pattern of papillae on the dorsal surface of the tongue that depends, at least in part, on redundant signaling by Fz3 and Fz6. Taken together, these experiments provide compelling evidence for a close mechanistic relationship between multiple anatomically diverse polarity processes.
Friday, November 7th
Li, X. Y., Harrison, M. M., Villalta, J. E., Kaplan, T. and Eisen, M. B. (2014). Establishment of regions of genomic activity during the maternal to zygotic transition. Elife 3 [Epub ahead of print]. PubMed ID: 25313869
This study describes the genome-wide distributions and temporal dynamics of nucleosomes and post-translational histone modifications throughout the maternal-to-zygotic transition in embryos of Drosophila melanogaster. At mitotic cycle 8, when few zygotic genes are being transcribed, embryonic chromatin is in a relatively simple state: there are few nucleosome free regions, undetectable levels of the histone methylation marks characteristic of mature chromatin, and low levels of histone acetylation at a relatively small number of loci. Histone acetylation increases by cycle 12, but it is not until cycle 14 that nucleosome free regions and domains of histone methylation become widespread. Early histone acetylation is strongly associated with regions that were previously shown to be bound in early embryos by the maternally deposited transcription factor Zelda, suggesting that Zelda triggers a cascade of events, including the accumulation of specific histone modifications, that plays a role in the subsequent activation of these sequences.
Maksimenko, O., Bartkuhn, M., Stakhov, V., Herold, M., Zolotarev, N., Jox, T., Buxa, M. K., Kirsch, R., Bonchuk, A., Fedotova, A., Kyrchanova, O., Renkawitz, R. and Georgiev, P. (2014). Two new insulator proteins, Pita and ZIPIC, target CP190 to chromatin. Genome Res [Epub ahead of print]. PubMed ID: 25342723
Insulators are multi-protein - DNA complexes that regulate the nuclear architecture. The Drosophila CP190 protein is a cofactor for the DNA-binding insulator proteins Su(Hw), CTCF, and BEAF-32. The fact that CP190 has been found at genomic sites devoid of either of the known insulator factors has until now been unexplained. This study has identified two DNA-binding zinc-finger proteins, Pita and a new factor named ZIPIC (CG7928), that interact with CP190 in vivo and in vitro at specific interaction domains. Genomic binding sites for these proteins are clustered with CP190 as well as with CTCF and BEAF-32. Model binding sites for Pita or ZIPIC demonstrate a partial enhancer-blocking activity and protect gene expression from PRE-mediated silencing. The function of the CTCF-bound MCP insulator sequence requires binding of Pita. These results identify two new insulator proteins and emphasize the unifying function of CP190, which can be recruited by many DNA-binding insulator proteins.
Hughes, S. E. and Hawley, R. S. (2014). Topoisomerase II is required for the proper separation of heterochromatic eegions during Drosophila melanogaster female meiosis. PLoS Genet 10: e1004650. PubMed ID: 25340780
Heterochromatic homology ensures the segregation of achiasmate chromosomes during meiosis I in Drosophila melanogaster females, perhaps as a consequence of the heterochromatic threads that connect achiasmate homologs during prometaphase I. This study asked how these threads, and other possible heterochromatic entanglements, are resolved prior to anaphase I. The knockdown of Topoisomerase II (Top2) by RNAi in the later stages of meiosis results in a specific defect in the separation of heterochromatic regions after spindle assembly. In Top2 RNAi-expressing oocytes, heterochromatic regions of both achiasmate and chiasmate chromosomes often failed to separate during prometaphase I and metaphase I. Heterochromatic regions were stretched into long, abnormal projections with centromeres localizing near the tips of the projections in some oocytes. Despite these anomalies, bipolar spindles were observed in most Top2 RNAi-expressing oocytes, although the obligately achiasmate 4th chromosomes exhibited a near complete failure to move toward the spindle poles during prometaphase I. Both achiasmate and chiasmate chromosomes displayed defects in biorientation. Given that euchromatic regions separate much earlier in prophase, no defects were expected or observed in the ability of euchromatic regions to separate during late prophase upon knockdown of Top2 at mid-prophase. Finally, embryos from Top2 RNAi-expressing females frequently failed to initiate mitotic divisions. These data suggest both that Topoisomerase II is involved in the resolution of heterochromatic DNA entanglements during meiosis I and that these entanglements must be resolved in order to complete meiosis.
Mengoli, V., Bucciarelli, E., Lattao, R., Piergentili, R., Gatti, M. and Bonaccorsi, S. (2014). The analysis of mutant alleles of different strength reveals multiple functions of topoisomerase 2 in regulation of Drosophila chromosome structure. PLoS Genet 10: e1004739. PubMed ID: 25340516
Topoisomerase II is a major component of mitotic chromosomes but its role in the assembly and structural maintenance of chromosomes is rather controversial, as different chromosomal phenotypes have been observed in various organisms and in different studies on the same organism. In contrast to vertebrates that harbor two partially redundant Topo II isoforms, Drosophila and yeasts have a single Topo II enzyme. In addition, fly chromosomes, unlike those of yeast, are morphologically comparable to vertebrate chromosomes. Thus, Drosophila is a highly suitable system to address the role of Topo II in the assembly and structural maintenance of chromosomes. This study shows that modulation of Top2 function in living flies by means of mutant alleles of different strength and in vivo RNAi results in multiple cytological phenotypes. In weak Top2 mutants, meiotic chromosomes of males exhibit strong morphological abnormalities and dramatic segregation defects, while mitotic chromosomes of larval brain cells are not affected. In mutants of moderate strength, mitotic chromosome organization is normal, but anaphases display frequent chromatin bridges that result in chromosome breaks and rearrangements involving specific regions of the Y chromosome and 3L heterochromatin. Severe Top2 depletion resulted in many aneuploid and polyploid mitotic metaphases with poorly condensed heterochromatin and broken chromosomes. Finally, in the almost complete absence of Top2, mitosis in larval brains was virtually suppressed and in the rare mitotic figures observed chromosome morphology was disrupted. These results indicate that different residual levels of Top2 in mutant cells can result in different chromosomal phenotypes, and that the effect of a strong Top2 depletion can mask the effects of milder Top2 reductions. Thus, these results suggest that the previously observed discrepancies in the chromosomal phenotypes elicited by Topo II downregulation in vertebrates might depend on slight differences in Topo II concentration and/or activity.
Oh, Y., Yoon, S. E., Zhang, Q., Chae, H. S., Daubnerova, I., Shafer, O. T., Choe, J. and Kim, Y. J. (2014). A homeostatic sleep-stabilizing pathway in Drosophila composed of the Sex Peptide receptor and its ligand, the myoinhibitory peptide. PLoS Biol 12: e1001974. PubMed ID: 25333796
Sleep, a reversible quiescent state found in both invertebrate and vertebrate animals, disconnects animals from their environment and is highly regulated for coordination with wakeful activities, such as reproduction. The fruit fly, Drosophila melanogaster, has proven to be a valuable model for studying the regulation of sleep by circadian clock and homeostatic mechanisms. This study demonstrates that the Sex peptide receptor (SPR) of Drosophila, known for its role in female reproduction, is also important in stabilizing sleep in both males and females. Mutants lacking either the SPR or its central ligand, myoinhibitory peptide (MIP), fall asleep normally, but have difficulty in maintaining a sleep-like state. This analyses have mapped the SPR sleep function to pigment dispersing factor (pdf) neurons, an arousal center in the insect brain. MIP downregulates intracellular cAMP levels in pdf neurons through the SPR. MIP is released centrally before and during night-time sleep, when the sleep drive is elevated. Sleep deprivation during the night facilitates MIP secretion from specific brain neurons innervating pdf neurons. Moreover, flies lacking either SPR or MIP cannot recover sleep after the night-time sleep deprivation. These results delineate a central neuropeptide circuit that stabilizes the sleep state by feeding a slow-acting inhibitory input into the arousal system and plays an important role in sleep homeostasis.
Xu, S., Wilf, R., Menon, T., Panikker, P., Sarthi, J. and Elefant, F. (2014). Epigenetic control of learning and memory in Drosophila by Tip60 HAT action. Genetics [Epub ahead of print]. PubMed ID: 25326235
Disruption of epigenetic gene control mechanisms in the brain causes significant cognitive impairment that is a debilitating hallmark of most neurodegenerative disorders including Alzheimer's disease (AD). Histone acetylation is one of the best characterized of these epigenetic mechanisms that is critical for regulating learning and memory associated gene expression profiles, yet the specific histone acetyltransferases (HATs) that mediate these effects have yet to be fully characterized. This study investigated an epigenetic role for the HAT Tip60 in learning and memory formation using the Drosophila CNS mushroom body (MB) as a well-characterized cognition model. Tip60 is endogenously expressed in the Kenyon cells, the intrinsic neurons of the MB and in the MB axonal lobes. Targeted loss of Tip60 HAT activity in the MB causes thinner and shorter axonal lobes while increasing Tip60 HAT levels cause no morphological defects. Functional consequences of both loss and gain of Tip60 HAT levels in the MB are evidenced by defects in immediate recall memory. ChIP-Seq analysis reveals that Tip60 target genes are enriched for functions in cognitive processes and accordingly, key genes representing these pathways are misregulated in the Tip60 HAT mutant fly brain. Remarkably, it was found that both learning and immediate recall memory deficits that occur under AD associated amyloid precursor protein (APP) induced neurodegenerative conditions can be effectively rescued by increasing Tip60 HAT levels specifically in the MB. Together, these findings uncover an epigenetic transcriptional regulatory role for Tip60 in cognitive function and highlight the potential of HAT activators as a therapeutic option for neurodegenerative disorders.
Li, Y., Fink, C., El-Kholy, S. and Roeder, T. (2014). The octopamine receptor octβ2R is essential for ovulation and fertilization in the fruit fly Drosophila melanogaster. Arch Insect Biochem Physiol. PubMed ID: 25353988
The biogenic monoamine octopamine is essential for ovulation and fertilization in insects. Release of this hormone from neurons in the thoracoabdominal ganglion triggers ovulation and sperm release from the spermathecae. This study shows that the effects of octopamine on ovulation are mediated by at least two different octopamine receptors. In addition to the Oamb receptor that is present in the epithelium of the oviduct, the octβ2R receptor is essential for ovulation and fertilization. Octβ2R is widely expressed in the female reproductive tract. Most prominent is expression in the oviduct muscle and the spermathecae. Animals deficient in expression of the receptor show a severe egg-laying defect. The corresponding females have a much larger ovary that is caused by egg retention in the ovary. Moreover, the very few laid eggs are not fertilized, indicating problems in the process of sperm delivery. It is assumed that octβ2R acts in a similar way as ss2-adrenoreceptors in smooth muscles, were activation of this receptor induces an increase in cAMP levels that lead to relaxation of the muscle. Taken together, these findings show that octopaminergic control of ovulation and fertilization is more complex than anticipated and that various receptors located in different cells act together to enable a well-orchestrated activity of the female reproductive system in response to copulation.
Mendes, C. S., Rajendren, S. V., Bartos, I., Marka, S. and Mann, R. S. (2014). Kinematic responses to changes in walking orientation and gravitational load in Drosophila melanogaster. PLoS One 9: e109204. PubMed ID: 25350743
Walking behavior is context-dependent, resulting from the integration of internal and external influences by specialized motor and pre-motor centers. Neuronal programs must be sufficiently flexible to the locomotive challenges inherent in different environments. Although insect studies have contributed substantially to the identification of the components and rules that determine locomotion, understanding of how multi-jointed walking insects respond to changes in walking orientation and direction and strength of the gravitational force is still not understood. In order to answer these questions the kinematic properties of untethered Drosophila was measured with high temporal and spatial resolution during inverted and vertical walking. In addition, the kinematic responses to increases in gravitational load were measured. Animals were found to be capable of shifting their step, spatial and inter-leg parameters in order to cope with more challenging walking conditions. For example, flies walking in an inverted orientation decreased the duration of their swing phase leading to increased contact with the substrate and, as a result, greater stability. It was also found that when flies carry additional weight, thereby increasing their gravitational load, some changes in step parameters vary over time, providing evidence for adaptation. However, above a threshold that is between 1 and 2 times their body weight flies display locomotion parameters that suggest they are no longer capable of walking in a coordinated manner. Finally, it was found that functional chordotonal organs are required for flies to cope with additional weight, as animals deficient in these proprioceptors display increased sensitivity to load bearing as well as other locomotive defects.
Wednesday, November 5th
Chai, G., Zhou, L., Manto, M., Helmbacher, F., Clotman, F., Goffinet, A. M. and Tissir, F. (2014). Celsr3 is required in motor neurons to steer their axons in the hindlimb. Nat Neurosci 17: 1171-1179. PubMed ID: 25108913
The cadherin Celsr3, homolog of Drosophila Flamingo, regulates the directional growth and targeting of axons in the CNS, but whether it acts in collaboration with or in parallel to other guidance cues is unknown. Furthermore, the function of Celsr3 in the peripheral nervous system is still largely unexplored. This study shows that Celsr3 mediates pathfinding of motor axons innervating the hindlimb. In mice, Celsr3-deficient axons of the peroneal nerve segregate from those of the tibial nerve but fail to extend dorsally, and they stall near the branch point. Mutant axons respond to repulsive ephrinA-EphA forward signaling and glial cell-derived neurotrophic factor (GDNF). However, they are insensitive to attractive EphA-ephrinA reverse signaling. In transfected cells, Celsr3 immunoprecipitates with ephrinA2, ephrinA5, Ret, GDNF family receptor alpha1 (GFRalpha1) and Frizzled3 (Fzd3). The function of Celsr3 is Fzd3 dependent but Vangl2 independent. These results provide evidence that the Celsr3-Fzd3 pathway interacts with EphA-ephrinA reverse signaling to guide motor axons in the hindlimb.
Samuel, M. A., Voinescu, P. E., Lilley, B. N., de Cabo, R., Foretz, M., Viollet, B., Pawlyk, B., Sandberg, M. A., Vavvas, D. G. and Sanes, J. R. (2014). LKB1 and AMPK regulate synaptic remodeling in old age. Nat. Neurosci 17: 1190-1197. PubMed ID: 25086610
Age-related decreases in neural function result in part from alterations in synapses. To identify molecular defects that lead to such changes, this study focused on the outer retina, in which synapses are markedly altered in old rodents and humans. The serine/threonine kinase LKB1 (see Drosophila Lkb1) and one of its substrates, AMPK (see Drosophila Ampk), regulate this process. In old mice, synaptic remodeling was accompanied by specific decreases in the levels of total LKB1 and active (phosphorylated) AMPK. In the absence of either kinase, young adult mice developed retinal defects similar to those that occurred in old wild-type animals. LKB1 and AMPK function in rod photoreceptors where their loss leads to aberrant axonal retraction, the extension of postsynaptic dendrites and the formation of ectopic synapses. Conversely, increasing AMPK activity genetically or pharmacologically attenuates and may reverse age-related synaptic alterations. Together, these results identify molecular determinants of age-related synaptic remodeling and suggest strategies for attenuating these changes.
Finci, L. I., Kruger, N., Sun, X., Zhang, J., Chegkazi, M., Wu, Y., Schenk, G., Mertens, H. D., Svergun, D. I., Zhang, Y., Wang, J. H. and Meijers, R. (2014). The crystal structure of netrin-1 in complex with DCC reveals the bifunctionality of netrin-1 as a guidance cue. Neuron 83: 839-849. PubMed ID: 25123307
Netrin-1 (see Drosophila Netrins) is a guidance cue that can trigger either attraction or repulsion effects on migrating axons of neurons, depending on the repertoire of receptors available on the growth cone. How a single chemotropic molecule can act in such contradictory ways has long been a puzzle at the molecular level. This study presents the crystal structure of netrin-1 in complex with the Deleted in Colorectal Cancer (DCC; see Drosophila Frazzled) receptor.One netrin-1 molecule can simultaneously bind to two DCC molecules through a DCC-specific site and through a unique generic receptor binding site, where sulfate ions staple together positively charged patches on both DCC and netrin-1. Furthermore, this study demonstrates that UNC5A (See Drosophila Unc5) can replace DCC on the generic receptor binding site to switch the response from attraction to repulsion. It is proposed that the modularity of binding allows for the association of other netrin receptors at the generic binding site, eliciting alternative turning responses.
Magnusson, J. P., Goritz, C., Tatarishvili, J., Dias, D. O., Smith, E. M., Lindvall, O., Kokaia, Z. and Frisen, J. (2014). A latent neurogenic program in astrocytes regulated by Notch signaling in the mouse. Science 346: 237-241. PubMed ID: 25301628
Neurogenesis is restricted in the adult mammalian brain; most neurons are neither exchanged during normal life nor replaced in pathological situations. This study reports that stroke elicits a latent neurogenic program in striatal astrocytes in mice. Notch1 (see Drosophila Notch) signaling is reduced in astrocytes after stroke, and attenuated Notch1 signaling is necessary for neurogenesis by striatal astrocytes. Blocking Notch signaling triggers astrocytes in the striatum and the medial cortex to enter a neurogenic program, even in the absence of stroke, resulting in ~850 new neurons in a mouse striatum. Thus, under Notch signaling regulation, astrocytes in the adult mouse brain parenchyma carry a latent neurogenic program that may potentially be useful for neuronal replacement strategies.
Joy, T., Hirono, K. and Doe, C. Q. (2014). The RanGEF Bj1 promotes Prospero nuclear export and neuroblast self-renewal. Dev Neurobiol [Epub ahead of print]. PubMed ID: 25312250
Drosophila larval neuroblasts are a model system for studying stem cell self-renewal and differentiation. This study report a novel role for the Drosophila gene Bj1 in promoting larval neuroblast self-renewal. Bj1 is the guanine-nucleotide exchange factor for Ran GTPase, which regulates nuclear import/export. Bj1 transcripts are highly enriched in larval brain neuroblasts (in both central brain and optic lobe), while Bj1 protein is detected in both neuroblasts and their neuronal progeny. Loss of Bj1 using both mutants or RNAi causes a progressive loss of larval neuroblasts, showing that Bj1 is required to maintain neuroblast numbers. Loss of Bj1 does not result in neuroblast apoptosis, but rather leads to abnormal nuclear accumulation of the differentiation factor Prospero, and premature neuroblast differentiation. It is concluded that the Bj1 RanGEF promotes Prospero nuclear export and neuroblast self-renewal.
Tuesday, November 4th
Patil, V. S., Anand, A., Chakrabarti, A. and Kai, T. (2014). The Tudor domain protein Tapas, a homolog of the vertebrate Tdrd7, functions in piRNA pathway to regulate retrotransposons in germline of Drosophila melanogaster. BMC Biol 12: 61. PubMed ID: 25287931
Piwi-associated RNAs (piRNAs) are a special class of small RNAs that provide defense against transposable elements (TEs) in animal germline cells. In Drosophila, germline piRNAs are thought to be processed at a unique perinuclear structure, nuage, which houses piRNA pathway proteins including the Piwi clade of Argonaute family proteins, along with several Tudor domain proteins, RNA helicases and nucleases. Tudor domain protein Tejas (Tej), an ortholog of vertebrate Tdrd5, is an important component of the piRNA pathway. The current study identified the paralog of Drosophila tej gene, tapas (tap), which is an ortholog of vertebrate Tdrd7. Like Tej, Tap is localized at the perinuclear structure in germline cells called nuage. The tap loss alone leads to a mild increase in transposon expression and decrease in piRNAs targeting transposons expressed in the germline. tap genetically interacts with other piRNA pathway genes, and Tap physically interacts with piRNA pathway components, such as Piwi family proteins Aubergine (Aub) and Argonaute3 (Ago3) and the RNA helicases Vasa (Vas) and Spindle-E (SpnE). tap together with tej is required for survival of germline cells during early stages and for polarity formation. It was further observed that loss of tej and tap together results in more severe defects in piRNA pathway in germline cells compared to single mutants: the double mutant ovaries exhibit mislocalization of piRNA pathway components and significantly greater reduction of piRNAs against transposons predominantly expressed in germline compared to single mutants. The single or double mutants did not have any reduction in piRNAs mapping to transposons predominantly expressed in gonadal somatic cells and those derived from unidirectional clusters such as flamenco. Consistently, the loss of both tej and tap function results in mislocalization of Piwi in germline cells, while Piwi remains localized to the nucleus in somatic cells. These data suggest that Tej and Tap work together for germline maintenance and piRNA production in germline cells. These observations suggest that tej and tap work together for the germline maintenance. tej and tap also function in a synergistic manner to maintain examined piRNA components at the perinuclear nuage and for piRNA production in Drosophila germline.
Kang, Y. S., Kurano, M. and Stumph, W. E. (2014). The Myb domain of the largest subunit of SNAPc adopts different architectural configurations on U1 and U6 snRNA gene promoter sequences. Nucleic Acids Res. PubMed ID: 25324315
The small nuclear RNA (snRNA) activating protein complex (SNAPc) is essential for transcription of genes that encode the snRNAs. Drosophila melanogaster SNAPc (DmSNAPc) consists of three subunits (DmSNAP190, DmSNAP50 and DmSNAP43) that form a stable complex that recognizes an snRNA gene promoter element called the PSEA. Although all three subunits are required for sequence-specific DNA binding activity, only DmSNAP190 possesses a canonical DNA binding domain consisting of 4.5 tandem Myb repeats homologous to the Myb repeats in the DNA binding domain of the Myb oncoprotein. This study used site-specific protein-DNA photo-cross-linking technology followed by site-specific protein cleavage to map domains of DmSNAP190 that interact with specific phosphate positions in the U6 PSEA. The results indicate that at least two DmSNAP190 Myb repeats contact the DNA in a significantly different manner when DmSNAPc binds to a U6 PSEA versus a U1PSEA, even though the two PSEA sequences differ at only 5 of 21 nucleotide positions. The results are consistent with a model in which the specific DNA sequences of the U1 and U6 PSEAs differentially alter the conformation of DmSNAPc, leading to the subsequent recruitment of different RNA polymerases to the U1 and U6 gene promoters.
Gandhi, S. G., Bag, I., Sengupta, S., Pal-Bhadra, M. and Bhadra, U. (2014). Drosophila oncogene Gas41 is RNAi modulator that intersects heterochromatin and siRNA pathway. FEBS J. PubMed ID: 25323651
Glioma amplified sequence41 (Gas41) is a highly conserved putative transcription factor that is frequently abundant in human gliomas. Gas41 shows oncogenic activity by promoting cell growth and viability. This study shows Gas41 is required for proper functioning of RNAi machinery in the nuclei, though three basic structural domains of RNAi components PAZ, PIWI and dsRNA binding are absent in the structural sequences. Variations of structural domains are highly conversed among prokaryotes and eukaryotes. Gas41 interacts with cytological RNase III enzyme Dicer1 both biochemically and genetically. However, Drosophila Gas41 functions as chromatin remodeler and interacts with different heterochromatin markers and repeat induced transgene silencing by modulating PEV. This study also shows that transcriptional inactive Gas41 mutant interferes the functional assembly of heterochromatin associated proteins, H3K9me2 and HP1 in developing embryos. A reduction of heterochromatic markers is accompanied with mini-w promoter sequence in Gas41 mutants. These findings suggest that, Drosophila Gas41 guides the repeat associated gene silencing, and Dicer1 interaction thereby depicting a new role of the Gas41. It is a critical RNAi component. In Drosophila, Gas41 plays a dual role. In one hand, it seems to participate with Dicer 1 in the RNAi pathway and alternatively also participate in repeat-induced gene silencing by accumulating heterochromatin proteins at the mw array promoters. Therefore, it proposes an intriguing and seemingly paradoxical new finding in RNA technology in the process of heterochromatin gene silencing.
Monday, November 3rd
Ranieri, N., Therond, P. P. and Ruel, L. (2014). Switch of PKA substrates from Cubitus interruptus to Smoothened in the Hedgehog signalosome complex. Nat Commun 5: 5034. PubMed ID: 25289679
Hedgehog (Hh) signalling is crucial for developmental patterning and tissue homeostasis. In Drosophila, Hh signalling is mediated by a bifunctional transcriptional mediator, called Cubitus interruptus (Ci). Protein Kinase A (PKA)-dependent phosphorylation of the serpentine protein Smoothened (Smo) leads to Ci activation, whereas PKA-dependent phosphorylation of Ci leads to the formation of Ci repressor form. The mechanism that switches PKA from an activator to a repressor is not known. This study shows that Hh signalling activation causes PKA to switch its substrates from Ci to Smo within the Hh signalling complex (HSC). In particular, Hh signalling increases the level of Smo, which then outcompetes Ci for association with PKA and causes a switch in PKA substrate recognition. A new model is proposed in which the PKA is constitutively present and active within the HSC, and in which the relative levels of Ci and Smo within the HSC determine differential activation and cellular response to Hh signalling.
Sopko, R., Foos, M., Vinayagam, A., Zhai, B., Binari, R., Hu, Y., Randklev, S., Perkins, L. A., Gygi, S. P. and Perrimon, N. (2014). Combining genetic perturbations and proteomics to examine kinase-phosphatase networks in Drosophila embryos. Dev Cell 31: 114-127. PubMed ID: 25284370
Connecting phosphorylation events to kinases and phosphatases is key to understanding the molecular organization and signaling dynamics of networks. This study has generated a validated set of transgenic RNA-interference reagents for knockdown and characterization of all protein kinases and phosphatases present during early Drosophila melanogaster development. These genetic tools enable collection of sufficient quantities of embryos depleted of single gene products for proteomics. As a demonstration of an application of the collection, multiplexed isobaric labeling was used for quantitative proteomics to derive global phosphorylation signatures associated with kinase-depleted embryos to systematically link phosphosites with relevant kinases. This strategy uncovers kinase consensus motifs and prioritizes phosphoproteins for kinase target validation. This approach was validated by providing auxiliary evidence for Wee kinase-directed regulation of the chromatin regulator Stonewall. Further, it was shown how correlative phosphorylation at the site level can indicate function, as exemplified by Sterile20-like kinase-dependent regulation of Stat92E.
Mishra, A. K., Sachan, N., Mutsuddi, M. and Mukherjee, A. (2014). TRAF6 is a novel regulator of Notch signaling in Drosophila melanogaster. Cell Signal 26: 3016-3026. PubMed ID: 25280943
Notch signaling pathway unravels a fundamental cellular communication system that plays an elemental role in development. It is evident from different studies that the outcome of Notch signaling depends on signal strength, timing, cell type, and cellular context. Since Notch signaling affects a spectrum of cellular activity at various developmental stages by reorganizing itself in more than one way to produce different intensities in the signaling output, it is important to understand the context dependent complexity of Notch signaling and different routes of its regulation. This study identified TRAF6 (Drosophila homolog of mammalian TRAF6) as an interacting partner of Notch intracellular domain (Notch-ICD). TRAF6 genetically interacts with Notch pathway components in trans-heterozygous combinations. Immunocytochemical analysis shows that TRAF6 co-localizes with Notch in Drosophila third instar larval tissues. The genetic interaction data suggests that the loss-of-function of TRAF6 leads to the rescue of previously identified Kurtz-Deltex mediated wing notching phenotype and enhances Notch protein survival. Co-expression of TRAF6 and Deltex results in depletion of Notch in the larval wing discs and down-regulates Notch targets, Wingless and Cut. Taken together, these results suggest that TRAF6 may function as a negative regulator of Notch signaling.
Hu, L., Huang, H., Li, J., Yin, M. X., Lu, Y., Wu, W., Zeng, R., Jiang, J., Zhao, Y. and Zhang, L. (2014). Drosophila CK2 promotes Wts to suppress Yki activity for growth control. J Biol Chem. PubMed ID: 25320084
Drosophila Hippo signaling regulates Wts activity to phosphorylate and inhibit Yki in order to control tissue growth. CK2 is widely expressed and involved in a variety of signaling pathways. This study reports that Drosophila CK2 promotes Wts activity to phosphorylate and inhibit Yki activity, which is independent of Hpo induced Wts promotion. In vivo, CK2 overexpression suppresses hpo mutant induced Ex upregulation and overgrowth phenotype while it cannot affect wts mutant. Consistent with this, knockdown of CK2 upregulates Hpo pathway target expression. It was also found that Drosophila CK2 is essential for tissue growth as a cell death inhibitor as knockdown of CK2 in developing discs induces severe growth defect as well as caspase3 (Drice) signal. Taken together, these results uncover a dual role of CK2: while its major role is promoting cell survive, it may potentially be a growth inhibitor as well.
Sunday, November 2nd
Paaby, A. B., Bergland, A. O., Behrman, E. L. and Schmidt, P. S. (2014). A highly pleiotropic amino acid polymorphism in the Drosophila insulin receptor contributes to life history adaptation. Evolution [Epub ahead of print]. PubMed ID: 25319083
Finding the specific nucleotides that underlie adaptive variation is a major goal in evolutionary biology, but polygenic traits pose a challenge because the complex genotype-phenotype relationship can obscure the effects of individual alleles. However, natural selection working in large wild populations can shift allele frequencies and indicate functional regions of the genome. Previous studies have shown that the two most common alleles of a complex amino acid insertion-deletion polymorphism in the Drosophila Insulin receptor show independent, parallel clines in frequency across the North American and Australian continents. This study reports that the cline is stable over at least a five-year period and that the polymorphism also demonstrates temporal shifts in allele frequency concurrent with seasonal change. The alleles were tested for effects on levels of insulin signaling, fecundity, development time, body size, stress tolerance, and lifespan. The alleles are associated with predictable differences in these traits, consistent with patterns of Drosophila life history variation across geography that likely reflect adaptation to the heterogeneous climatic environment. These results implicate insulin signaling as a major mediator of life history adaptation in Drosophila, and suggest that life history tradeoffs can be explained by extensive pleiotropy at a single locus.
Saisawang, C. and Ketterman, A. J. (2014). Micro-plasticity of genomes as illustrated by the evolution of glutathione transferases in 12 Drosophila species. PLoS One 9: e109518. PubMed ID: 25310450
Glutathione transferases (GST) are an ancient superfamily comprising a large number of paralogous proteins in a single organism. This multiplicity of GSTs has allowed the copies to diverge for neofunctionalization with proposed roles ranging from detoxication and oxidative stress response to involvement in signal transduction cascades. A comparative genomic analysis was performed using FlyBase annotations and Drosophila melanogaster GST sequences as templates to further annotate the GST orthologs in the 12 Drosophila sequenced genomes (see Micro-Plasticity of Genomes As Illustrated by the Evolution of Glutathione Transferases in 12 Drosophila Species). This study found that GST genes in the Drosophila subgenera have undergone repeated local duplications followed by transposition, inversion, and micro-rearrangements of these copies. The colinearity and orientations of the orthologous GST genes appear to be unique in many of the species which suggests that genomic rearrangement events have occurred multiple times during speciation. The high micro-plasticity of the genomes appears to have a functional contribution utilized for evolution of this gene family.
Williams, C. M., Watanabe, M., Guarracino, M. R., Ferraro, M. B., Edison, A. S., Morgan, T. J., Boroujerdi, A. F. and Hahn, D. A. (2014). Cold adaptation shapes the robustness of metabolic networks in Drosophila melanogaster. Evolution [Epub ahead of print]. PubMed ID: 25308124
When ectotherms are exposed to low temperatures, they enter a cold-induced coma (chill coma) that prevents resource acquisition, mating, oviposition, and escape from predation. There is substantial variation in time taken to recover from chill coma both within and among species, and this variation is correlated with habitat temperatures such that insects from cold environments recover more quickly. This suggests an adaptive response, but the mechanisms underlying variation in recovery times are unknown, making it difficult to decisively test adaptive hypotheses. This study used replicated lines of Drosophila melanogaster selected in the laboratory for fast (hardy) or slow (susceptible) chill-coma recovery times to investigate modifications to metabolic profiles associated with cold adaptation. Metabolite concentrations of flies before, during, and after cold exposure were measured using NMR spectroscopy to test the hypotheses that hardy flies maintain metabolic homeostasis better during cold exposure and recovery, and that their metabolic networks are more robust to cold-induced perturbations. The metabolites of cold-hardy flies were less cold responsive and their metabolic networks during cold exposure were more robust, supporting the hypotheses. Metabolites involved in membrane lipid synthesis, tryptophan metabolism, oxidative stress, energy balance, and proline metabolism were altered by selection on cold tolerance. The potential significance of these alterations are discussed
Saturday, November 1st
Shmueli, M. D., Schnaider, L., Herzog, G., Gazit, E. and Segal, D. (2014). Computational and experimental characterization of dVHL establish a Drosophila model of VHL syndrome. PLoS One 9: e109864. PubMed ID: 25310726
The von Hippel-Lindau (VHL) cancer syndrome is associated with mutations in the VHL gene. The pVHL protein is involved in response to changes in oxygen availability as part of an E3-ligase that targets the Hypoxia-Inducible Factor (see Drosophila Similar) for degradation. pVHL has a molten globule configuration with marginal thermodynamic stability. The cancer-associated mutations further destabilize it. The Drosophila homolog, dVHL, has relatively low sequence similarity to pVHL, and is also involved in regulating HIF1-α. Using in silico, in vitro and in vivo approaches this study demonstrates high similarity between the structure and function of dVHL and pVHL. These proteins have a similar fold, secondary and tertiary structures, as well as thermodynamic stability. Key functional residues in dVHL are evolutionary conserved. This structural homology underlies functional similarity of both proteins, evident by their ability to bind their reciprocal partner proteins, and by the observation that transgenic pVHL can fully maintain normal dVHL-HIF1-alpha downstream pathways in flies. This novel transgenic Drosophila model is thus useful for studying the VHL syndrome, and for testing drug candidates to treat it.
Breda, C., Nugent, M. L., Estranero, J. G., Kyriacou, C. P., Outeiro, T. F., Steinert, J. R. and Giorgini, F. (2014). Rab11 modulates alpha-synuclein mediated defects in synaptic transmission and behaviour. Hum Mol Genet [Epub ahead of print]. PubMed ID: 25305083
A central pathological hallmark of Parkinson's disease (PD) is the presence of proteinaceous depositions known as Lewy bodies, which consist largely of the protein α-synuclein (αSyn). Mutations, multiplications, and polymorphisms in the gene encoding αSyn are associated with familial forms of PD and susceptibility to idiopathic PD. Alterations in αSyn impair neuronal vesicle formation/transport, and likely contribute to PD pathogenesis by neuronal dysfunction and degeneration. αSyn is functionally associated with several Rab family GTPases, which perform various roles in vesicle trafficking. This study explored the role of the endosomal recycling factor Rab11 in the pathogenesis of PD using Drosophila models of aSyn toxicity. αSyn induces synaptic potentiation at the larval neuromuscular junction by increasing synaptic vesicle size, and that these alterations are reversed by Rab11 overexpression. Furthermore, Rab11 decreases aSyn aggregation and ameliorates several αSyn-dependent phenotypes in both larvae and adult fruit flies, including locomotor activity, degeneration of dopaminergic neurons, and shortened lifespan. This work highlights the importance of Rab11 in the modulation of synaptic vesicle size and consequent enhancement of synaptic function. The results suggest that targeting Rab11 activity could have therapeutic value in PD.
Barbaro, B. A., Lukacsovich, T., Agrawal, N., Burke, J., Bornemann, D. J., Purcell, J. M., Worthge, S. A., Caricasole, A., Weiss, A., Song, W., Morozova, O. A., Colby, D. W. and Marsh, J. L. (2014). Comparative study of naturally occurring huntingtin fragments in Drosophila points to exon 1 as the most pathogenic species in Huntington's disease. Hum Mol Genet [Epub ahead of print]. PubMed ID: 25305076
Although Huntington's disease is caused by the expansion of a CAG triplet repeat within the context of the 3144-amino acid huntingtin protein (HTT), studies reveal that N-terminal fragments of HTT containing the expanded PolyQ region can be produced by proteolytic processing and/or aberrant splicing. N-terminal HTT fragments are also prevalent in postmortem tissue, and expression of some of these fragments in model organisms can cause pathology. This has led to the hypothesis that N-terminal peptides may be critical modulators of disease pathology, raising the possibility that targeting aberrant splicing or proteolytic processing may present attractive therapeutic targets. However, many factors can contribute to pathology, including genetic background and differential expression of transgenes, in addition to intrinsic differences between fragments and their cellular effects. This study used Drosophila as a model system to determine the relative toxicities of different naturally occurring huntingtin fragments in a system in which genetic background, transgene expression levels and post-translational proteolytic processing can be controlled. These studies reveal that among the naturally occurring N-terminal HTT peptides, the exon 1 peptide is exceptionally pathogenic and exhibits unique structural and biophysical behaviors that do not appear to be incremental changes compared with other fragments. If this proves correct, efforts to specifically reduce the levels of exon 1 peptides or to target toxicity-influencing post-translational modifications that occur with the exon 1 context are likely to have the greatest impact on pathology.
Hess-Homeier, D. L., Fan, C. Y., Gupta, T., Chiang, A. S. and Certel, S. J. (2014). Astrocyte-specific regulation of hMeCP2 expression in Drosophila. Biol Open [Epub ahead of print]. PubMed ID: 25305037
Alterations in the expression of Methyl-CpG-binding protein 2 (MeCP2) either by mutations or gene duplication leads to a wide spectrum of neurodevelopmental disorders including Rett Syndrome and MeCP2 duplication disorder. Common features of Rett Syndrome (RTT), MeCP2 duplication disorder, and neuropsychiatric disorders indicate that even moderate changes in MeCP2 protein levels result in functional and structural cell abnormalities. This study investigated two areas of MeCP2 pathophysiology using Drosophila as a model system: the effects of MeCP2 glial gain-of-function activity on circuits controlling sleep behavior, and the cell-type specific regulation of MeCP2 expression. This study examined the effects of elevated MeCP2 levels on microcircuits by expressing human MeCP2 (hMeCP2) in astrocytes and distinct subsets of amine neurons including dopamine and octopamine (OA) neurons. Depending on the cell-type, hMeCP2 expression reduced sleep levels, altered daytime/nighttime sleep patterns, and generated sleep maintenance deficits. A 498 base pair region of the MeCP2e2 isoform was identified that is targeted for regulation in distinct subsets of astrocytes. Levels of the full-length hMeCP2e2 and mutant RTT R106W protein decreased in astrocytes in a temporally and spatially regulated manner. In contrast, expression of the deletion Delta166 hMeCP2 protein was not altered in the entire astrocyte population. qPCR experiments revealed a reduction in full-length hMeCP2e2 transcript levels suggesting transgenic hMeCP2 expression is regulated at the transcriptional level. Given the phenotypic complexities that are caused by alterations in MeCP2 levels, these results provide insight into distinct cellular mechanisms that control MeCP2 expression and link microcircuit abnormalities with defined behavioral deficits.
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