What's hot today:
Current papers in developmental biology and gene function


Friday, December 29th

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Freda, P. J., Alex, J. T., Morgan, T. J. and Ragland, G. J. (2017). Genetic decoupling of thermal hardiness across metamorphosis in Drosophila melanogaster. Integr Comp Biol 57(5): 999-1009. PubMed ID: 29045669
Highly seasonal environments expose life stages to often drastically different thermal environments. To investigate whether developmental genetic correlations constrain the evolution thermal hardiness of Drosophila, this study applied quantitative genetic analyses to cold hardiness measured in both larvae and adults from isogenic lines of the Drosophila Genetic Reference Panel (DGRP), using survival at stressful low temperatures as the phenotypic metric. Quantitative genetic analyses revealed no significant genetic correlation for cold hardiness between life stages, suggesting complete genetic decoupling of thermal hardiness across the metamorphic boundary. Both quantitative genetic and GWA analyses suggested that polygenic variation underlies cold hardiness in both stages, and that associated loci largely affected one stage or the other, but not both. However, reciprocal enrichment tests and correlations between BSLMM parameters for each life stage support some shared physiological mechanisms that may reflect common cellular thermal response pathways. Overall, these results suggest no developmental genetic constraints on cold hardiness across metamorphosis in D. melanogaster, an important consideration in evolutionary models of responses to changing climates. Genetic correlations for environmental sensitivity across ontogeny remains largely unexplored in other organisms, thus assessing the generality of genetic decoupling will require further quantitative or population genetic analysis in additional species.
Bost, A., Franzenburg, S., Adair, K. L., Martinson, V. G., Loeb, G. and Douglas, A. E. (2017). How gut transcriptional function of Drosophila melanogaster varies with the presence and composition of the gut microbiota. Mol Ecol [Epub ahead of print]. PubMed ID: 29113026
Despite evidence from laboratory experiments that perturbation of the gut microbiota affects many traits of the animal host, understanding of the effect of variation in microbiota composition on animals in natural populations is very limited. This study used Drosophila to identify the impact of natural variation in the taxonomic composition of gut bacterial communities on host traits, with the gut transcriptome as a molecular index of microbiota-responsive host traits. Use of the gut transcriptome was validated by demonstrating significant transcriptional differences between the guts of laboratory flies colonized with bacteria and maintained under axenic conditions. Wild Drosophila from six field collections made over two years had gut bacterial communities of diverse composition, dominated to varying extents by Acetobacteraceae and Enterobacteriaceae. The gut transcriptomes also varied among collections and differed markedly from those of laboratory flies. However, no overall relationship between variation in the wild fly transcriptome and taxonomic composition of the gut microbiota was evident at all taxonomic scales of bacteria tested for both individual fly genes and functional categories in Gene Ontology. It is concluded that the interaction between microbiota composition and host functional traits may be confounded by uncontrolled variation in both ecological circumstance and host traits (e.g., genotype, age physiological condition) under natural conditions, and that microbiota effects on host traits identified in the laboratory should, therefore, be extrapolated to field population with great caution.
Bolukbasi, E., Khericha, M., Regan, J. C., Ivanov, D. K., Adcott, J., Dyson, M. C., Nespital, T., Thornton, J. M., Alic, N. and Partridge, L. (2017). Intestinal Fork head regulates nutrient absorption and promotes longevity. Cell Rep 21(3): 641-653. PubMed ID: 29045833
Reduced activity of nutrient-sensing signaling networks can extend organismal lifespan, yet the underlying biology remains unclear. This study shows that the anti-aging effects of rapamycin and reduced intestinal insulin/insulin growth factor (IGF) signaling (IIS) require the Drosophila FoxA transcription factor homolog Fork Head (FKH). Intestinal FKH induction extends lifespan, highlighting a role for the gut. FKH binds to and is phosphorylated by AKT and Target of Rapamycin. Gut-specific FKH upregulation improves gut barrier function in aged flies. Additionally, it increases the expression of nutrient transporters, as does lowered IIS. Evolutionary conservation of this effect of lowered IIS is suggested by the upregulation of related nutrient transporters in insulin receptor substrate 1 knockout mouse intestine. These studies highlights a critical role played by FKH in the gut in mediating anti-aging effects of reduced IIS. Malnutrition caused by poor intestinal absorption is a major problem in the elderly, and a better understanding of the mechanisms involved will have important therapeutic implications for human aging.
Dacanay, F. N. D., Ladra, M., Junio, H. A. and Nellas, R. B. (2017). Molecular affinity of Mabolo extracts to an Octopamine receptor of a fruit fly. Molecules 22(10). PubMed ID: 29064449
Essential oils extracted from plants are composed of volatile organic compounds that can affect insect behavior. Identifying the active components of the essential oils to their biochemical target is necessary to design novel biopesticides. In this study, essential oils extracted from Diospyros discolor (Willd.) were analyzed using gas chromatography mass spectroscopy (GC-MS) to create an untargeted metabolite profile. Subsequently, a conformational ensemble of the Drosophila melanogaster Octopamine receptor in mushroom bodies (OAMB) was created from a molecular dynamics simulation to resemble a flexible receptor for docking studies. GC-MS analysis revealed the presence of several metabolites, i.e. mostly aromatic esters. Interestingly, these aromatic esters were found to exhibit relatively higher binding affinities to OAMB than the receptor's natural agonist, octopamine. The molecular origin of this observed enhanced affinity is the pi -stacking interaction between the aromatic moieties of the residues and ligands. This strategy, computational inspection in tandem with untargeted metabolomics, may provide insights in screening the essential oils as potential OAMB inhibitors.
Doran, M. L., Knee, J. M., Wang, N., Rzezniczak, T. Z., Parkes, T. L., Li, L. and Merritt, T. J. S. (2017). Metabolomic analysis of oxidative stress: Superoxide dismutase mutation and paraquat induced stress in Drosophila melanogaster. Free Radic Biol Med 113: 323-334. PubMed ID: 29031835
To determine the broad metabolic effects of oxidative stress, this study has quantified the response in Drosophila to both oxidative stress. Flies were challenged with loss of Superoxide dismutase activity or exposure to paraquat. Metabolic changes were then quantified using a LC-MS platform. Wide spread changes were found in both submetabolomes in response to all three types of stresses including: changes to the urea cycle, tryptophan metabolism, porphyrin metabolism, as well as a series of metabolic pathways involved in glutathione synthesis. Strikingly, while there are commonalities across the conditions, all three resulted in different metabolomic responses, with the greatest difference between the genetic and environmental responses. Genetic oxidative stress resulted in substantially more widespread effects, both in terms of the percent of the metabolome altered, and the magnitude of changes in individual metabolites. Chronic and acute environmental stress resulted in more similar responses although both were distinct from genetic stress. Overall, these results indicate that the metabolomic response to oxidative stress is complex, reaching across multiple metabolic pathways, with some shared features but with more features unique to different, specific stressors.
Erkosar, B., Kolly, S., van der Meer, J. R. and Kawecki, T. J. (2017). Adaptation to chronic nutritional stress leads to reduced dependence on microbiota in Drosophila melanogaster. MBio 8(5). PubMed ID: 29066546
Numerous studies have shown that animal nutrition is tightly linked to gut microbiota, especially under nutritional stress. In Drosophila, microbiota are known to promote juvenile growth, development, and survival on poor diets, mainly through enhanced digestion leading to changes in hormonal signaling. This study shows that this reliance on microbiota is greatly reduced in replicated Drosophila populations that became genetically adapted to a poor larval diet in the course of over 170 generations of experimental evolution. Protein and polysaccharide digestion in these poor-diet-adapted populations became much less dependent on colonization with microbiota. This was accompanied by changes in expression levels of dFOXO transcription factor, a key regulator of cell growth and survival, and many of its targets. These evolutionary changes in the expression of dFOXO targets to a large degree mimic the response of the same genes to microbiota, suggesting that the evolutionary adaptation to poor diet acted on mechanisms that normally mediate the response to microbiota. This study suggests that some metazoans have retained the evolutionary potential to adapt their physiology such that association with microbiota may become optional rather than essential.

Thursday, December 28th

Varadkar, P., Abbasi, F., Takeda, K., Dyson, J. J. and McCright, B. (2017). PP2A-B56gamma is required for an efficient spindle assembly checkpoint. Cell Cycle 16(12): 1210-1219. PubMed ID: 28562161
Evolutionary Homolog Study
The Spindle Assembly Checkpoint (SAC) is part of a complex feedback system designed to ensure that cells do not proceed through mitosis unless all chromosomal kinetochores have attached to spindle microtubules. The formation of the kinetochore complex and the implementation of the SAC are regulated by multiple kinases and phosphatases. BubR1 is a phosphoprotein that is part of the Cdc20 containing mitotic checkpoint complex that inhibits the APC/C so that Cyclin B1 and Securin are not degraded, thus preventing cells going into anaphase. In this study, it was found that PP2A in association with its B56gamma regulatory subunit, are needed for the stability of BubR1 during nocodazole induced cell cycle arrest. In primary cells that lack B56gamma, BubR1 is prematurely degraded and the cells proceed through mitosis. The reduced SAC efficiency results in cells with abnormal chromosomal segregation, a hallmark of transformed cells. Previous studies on PP2A's role in the SAC and kinetochore formation were done using siRNAs to all 5 of the B56 family members. This study shows that inactivation of only the PP2A-B56gamma subunit can affect the efficiency of the SAC. Data is provided that show the intracellular locations of the B56 subunits varies between family members, which is consistent with the hypothesis that they are not completely functionally redundant.
Li, Z., Cui, Q., Xu, J., Cheng, D., Wang, X., Li, B., Lee, J. M., Xia, Q., Kusakabe, T. and Zhao, P. (2017). SUMOylation regulates the localization and activity of Polo-like kinase 1 during cell cycle in the silkworm, Bombyx mori. Sci Rep 7(1): 15536. PubMed ID: 29138491
Polo-like kinase 1 (Plk1) is a crucial cell cycle regulator by its specific localization and activity during cell cycle. It has been shown that the phosphorylation and ubiquitylation of Plk1 are required for its own activation and localization. This study reports that SUMOylation regulates the activity of Plk1 in the lepidopteran insect of Bombyx mori. In the absence of SUMOylation, it causes the lost localization of Plk1 on centrosomes and kinetochores, as well as an uneven distribution in midzone. It was further identified that the putative SUMOylation site of Bombyx Plk1 at lysine 466 is required for its localization on centrosomes, and K466 mutation in Plk1 could influence its interaction with Smt3/Ubc9 complex. These findings are also confirmed by Drosophila Polo and human Plk1, which together reveals a conserved role of Plk1 SUMOylation in mammals. Moreover, conjugation of Smt3 to Plk1 SUMOylation mutant promotes its localization on centrosomes and kinetochores, and rescues functional defects of chromosome alignment in cells depleted of endogenous Plk1. Altogether, the present data indicate that the SUMOylation of Plk1 could participate in proper chromosome alignment and segregation during mitosis, and provides a novel layer for the regulation of Plk1 localization and activity throughout cell cycle.
Henze, R., Dittrich, P. and Ibrahim, B. (2017). A Dynamical Model for Activating and Silencing the Mitotic Checkpoint. Sci Rep 7(1): 3865. PubMed ID: 28634351
Evolutionary Homolog Study
The spindle assembly checkpoint (SAC) is an evolutionarily conserved mechanism, exclusively sensitive to the states of kinetochores attached to microtubules. During metaphase, the anaphase-promoting complex/cyclosome (APC/C) is inhibited by the SAC but it rapidly switches to its active form following proper attachment of the final spindle. It had been thought that APC/C activity is an all-or-nothing response, but recent findings have demonstrated that it switches steadily. This study develop a detailed mathematical model that considers all 92 human kinetochores and all major proteins involved in SAC activation and silencing. Deterministic and spatially-stochastic simulations were developed and it was found that certain spatial properties do not play significant roles. Furthermore, it was shown that this model is consistent with in-vitro mutation experiments of crucial proteins as well as the recently-suggested rheostat switch behavior, measured by Securin (see Drosophila Pimples) or CyclinB concentration. Considering an autocatalytic feedback loop leads to an all-or-nothing toggle switch in the underlying core components, while the output signal of the SAC still behaves like a rheostat switch. The results of this study support the hypothesis that the SAC signal varies with increasing number of attached kinetochores, even though it might still contain toggle switches in some of its components.
Ji, Z., Gao, H., Jia, L., Li, B. and Yu, H. (2017). A sequential multi-target Mps1 phosphorylation cascade promotes spindle checkpoint signaling. Elife 6. PubMed ID: 28072388
The master spindle checkpoint kinase Mps1 senses kinetochore-microtubule attachment and promotes checkpoint signaling to ensure accurate chromosome segregation. The kinetochore scaffold Knl1, when phosphorylated by Mps1, recruits checkpoint complexes Bub1-Bub3 and BubR1-Bub3 to unattached kinetochores. Active checkpoint signaling ultimately enhances the assembly of the mitotic checkpoint complex (MCC) consisting of BubR1-Bub3, Mad2, and Cdc20, which inhibits the anaphase-promoting complex or cyclosome bound to Cdc20 (APC/C(Cdc20)) to delay anaphase onset. Using in vitro reconstitution in a multi-organism study, this study shows that Mps1 promotes APC/C inhibition by MCC components through phosphorylating Bub1 and Mad1. Phosphorylated Bub1 binds to Mad1-Mad2. Phosphorylated Mad1 directly interacts with Cdc20. Mutations of Mps1 phosphorylation sites in Bub1 or Mad1 abrogate the spindle checkpoint in human cells. Therefore, Mps1 promotes checkpoint activation through sequentially phosphorylating Knl1, Bub1, and Mad1. This sequential multi-target phosphorylation cascade makes the checkpoint highly responsive to Mps1 and to kinetochore-microtubule attachment.
Golub, O., Wee, B., Newman, R. A., Paterson, N. M. and Prehoda, K. E. (2017). Activation of Discs large by aPKC aligns the mitotic spindle to the polarity axis during asymmetric cell division. Elife 6. PubMed ID: 29185419
Asymmetric division generates cellular diversity by producing daughter cells with different fates. In animals, the mitotic spindle aligns with Par complex polarized fate determinants, ensuring that fate determinant cortical domains are bisected by the cleavage furrow. This study investigated the mechanisms that couple spindle orientation to polarity during asymmetric cell division of Drosophila neuroblasts. The tumor suppressor Discs large (Dlg) was found to link the Par complex component atypical Protein Kinase C (aPKC) to the essential spindle orientation factor GukHolder (GukH). Dlg is autoinhibited by an intramolecular interaction between its SH3 and GK domains, preventing Dlg interaction with GukH at cortical sites lacking aPKC. When co-localized with aPKC, Dlg is phosphorylated in its SH3 domain which disrupts autoinhibition and allows GukH recruitment by the GK domain. This work establishes a molecular connection between the polarity and spindle orientation machineries during asymmetric cell division.
Kachaner, D., Garrido, D., Mehsen, H., Normandin, K., Lavoie, H. and Archambault, V. (2017). Coupling of Polo kinase activation to nuclear localization by a bifunctional NLS is required during mitotic entry. Nat Commun 8(1): 1701. PubMed ID: 29167465
The Polo kinase is a master regulator of mitosis and cytokinesis conserved from yeasts to humans. Polo is composed of an N-term kinase domain (KD) and a C-term polo-box domain (PBD), which regulates its subcellular localizations. The PBD and KD can interact and inhibit each other, and this reciprocal inhibition is relieved when Polo is phosphorylated at its activation loop. How Polo activation and localization are coupled during mitotic entry is unknown. This study reports that PBD binding to the KD masks a nuclear localization signal (NLS). Activating phosphorylation of the KD leads to exposure of the NLS and entry of Polo into the nucleus before nuclear envelope breakdown. Failures of this mechanism result in misregulation of the Cdk1-activating Cdc25 phosphatase and lead to mitotic and developmental defects in Drosophila. These results uncover spatiotemporal mechanisms linking master regulatory enzymes during mitotic entry.

Wednesday, December 27

Li, Q., Tjong, H., Li, X., Gong, K., Zhou, X. J., Chiolo, I. and Alber, F. (2017). The three-dimensional genome organization of Drosophila melanogaster through data integration. Genome Biol 18(1): 145. PubMed ID: 28760140
Genome structures are dynamic and non-randomly organized in the nucleus of higher eukaryotes. To maximize the accuracy and coverage of three-dimensional genome structural models, it is important to integrate all available sources of experimental information about a genome's organization. It remains a major challenge to integrate such data from various complementary experimental methods. This study presents an approach for data integration to determine a population of complete three-dimensional genome structures that are statistically consistent with data from both genome-wide chromosome conformation capture (Hi-C) and lamina-DamID experiments. These structures resolve the genome at the resolution of topological domains, and reproduce simultaneously both sets of experimental data. Importantly, this data deconvolution framework allows for structural heterogeneity between cells, and hence accounts for the expected plasticity of genome structures. As a case study Drosophila melanogaster embryonic cells, for which both data types are available, were chosen. The three-dimensional genome structures have strong predictive power for structural features not directly visible in the initial data sets, and reproduce experimental hallmarks of the D. melanogaster genome organization from imaging experiments. Also they reveal a number of new insights about genome organization and its functional relevance, including the preferred locations of heterochromatic satellites of different chromosomes, and observations about homologous pairing that cannot be directly observed in the original Hi-C or lamina-DamID data. This approach allows systematic integration of Hi-C and lamina-DamID data for complete three-dimensional genome structure calculation, while also explicitly considering genome structural variability.
Ma, Y. and Buttitta, L. (2017). Chromatin organization changes during the establishment and maintenance of the postmitotic state. Epigenetics Chromatin 10(1): 53. PubMed ID: 29126440
Genome organization changes during development as cells differentiate. Chromatin motion becomes increasingly constrained and heterochromatin clusters as cells become restricted in their developmental potential. These changes coincide with slowing of the cell cycle, which can also influence chromatin organization and dynamics. Terminal differentiation is often coupled with permanent exit from the cell cycle, and existing data suggest a close relationship between a repressive chromatin structure and silencing of the cell cycle in postmitotic cells. This study examined the relationship between chromatin organization, terminal differentiation and cell cycle exit. These studies focused on the Drosophila wing, where epithelial cells transition from active proliferation to a postmitotic state in a temporally controlled manner. Two stages of G0 were found in this tissue, a flexible G0 period where cells can be induced to reenter the cell cycle under specific genetic manipulations and a state that is called "robust," where cells become strongly refractory to cell cycle reentry. Compromising the flexible G0 by driving ectopic expression of cell cycle activators causes a global disruption of the clustering of heterochromatin-associated histone modifications such as H3K27 trimethylation and H3K9 trimethylation, as well as their associated repressors, Polycomb and Heterochromatin protein 1 (HP1). However, this disruption is reversible. When cells enter a robust G0 state, even in the presence of ectopic cell cycle activity, clustering of heterochromatin-associated modifications is restored. If cell cycle exit is bypassed, cells in the wing continue to terminally differentiate, but heterochromatin clustering is severely disrupted. Heterochromatin-dependent gene silencing does not appear to be required for cell cycle exit, as compromising the H3K27 methyltransferase Enhancer of zeste, and/or HP1 cannot prevent the robust cell cycle exit, even in the face of normally oncogenic cell cycle activities. It is concluded that heterochromatin clustering during terminal differentiation is a consequence of cell cycle exit, rather than differentiation. Compromising heterochromatin-dependent gene silencing does not disrupt cell cycle exit.
Li, W., Yi, J., Agbu, P., Zhou, Z., Kelley, R. L., Kallgren, S., Jia, S. and He, X. (2017). Replication stress affects the fidelity of nucleosome-mediated epigenetic inheritance. PLoS Genet 13(7): e1006900. PubMed ID: 28749973
The fidelity of epigenetic inheritance or the precision by which epigenetic information is passed along is an essential parameter for measuring the effectiveness of the process. This study performed quantitative measurement of epigenetic fidelity, using position effect variegation (PEV) in S. pombe as readout, to explore whether replication perturbation affects nucleosome-mediated epigenetic inheritance. Replication stresses, due to either hydroxyurea treatment or various forms of genetic lesions of the replication machinery, reduce the inheritance accuracy of CENP-A/Cnp1 nucleosome positioning within centromere. Mechanistically, this study demonstrates that excessive formation of single-stranded DNA, a common molecular abnormality under these conditions, might have correlation with the reduction in fidelity of centromeric chromatin duplication. Furthermore, it was shown that replication stress broadly changes chromatin structure at various loci in the genome, such as telomere heterochromatin expanding and mating type locus heterochromatin spreading out of the boundaries. Interestingly, the levels of inheritable expanding at sub-telomeric heterochromatin regions are highly variable among independent cell populations. Finally, this study shows that HU treatment of the multi-cellular organisms C. elegans and D. melanogaster affects epigenetically programmed development and PEV, illustrating the evolutionary conservation of the phenomenon. Replication stress, in addition to its demonstrated role in genetic instability, promotes variable epigenetic instability throughout the epigenome.
Dasari, V., Srivastava, S., Khan, S. and Mishra, R. K. (2017). Epigenetic factors Polycomb (Pc) and Suppressor of zeste (Su(z)2) negatively regulate longevity in Drosophila melanogaster. Biogerontology [Epub ahead of print]. PubMed ID: 29177687
The process of aging is a hallmark of the natural life span of all organisms and individuals within a population show variability in the measures of age related performance. Longevity and the rate of aging are influenced by several factors such as genetics, nutrition, stress, and environment. Many studies have focused on the genes that impact aging and there is increasing evidence that epigenetic factors regulate these genes to control life span. Polycomb (PcG) and Trithorax (trxG) protein complexes maintain the expression profiles of developmentally important genes and regulate many cellular processes. This study reports that mutations of PcG and trxG members affect the process of aging in Drosophila melanogaster, with perturbations mostly associated with retardation in aging. Mutations in polycomb repressive complex (PRC1) components Pc and Su(z)2 increase fly survival. Using an inducible UAS-GAL4 system, it was shown that this effect is tissue-specific; knockdown in fat body, but not in muscle or brain tissues, enhances life span. It is hypothesized that these two proteins influence life span via pathways independent of their PRC1 functions, with distinct effects on response to oxidative stress. These observations highlight the role of global epigenetic regulators in determining life span.
Huang, C., Yang, F., Zhang, Z., Zhang, J., Cai, G., Li, L., Zheng, Y., Chen, S., Xi, R. and Zhu, B. (2017). Mrg15 stimulates Ash1 H3K36 methyltransferase activity and facilitates Ash1 Trithorax group protein function in Drosophila. Nat Commun 8(1): 1649. PubMed ID: 29158494
Ash1 is a Trithorax group protein that possesses H3K36-specific histone methyltransferase activity, which antagonizes Polycomb silencing. This study reports the identification of two Ash1 complex subunits, Mrg15 and Nurf55. In vitro, Mrg15 stimulates the enzymatic activity of Ash1. In vivo, Mrg15 is recruited by Ash1 to their common targets, and Mrg15 reinforces Ash1 chromatin association and facilitates the proper deposition of H3K36me2. To dissect the functional role of Mrg15 in the context of the Ash1 complex, this study identified an Ash1 point mutation (Ash1-R1288A) that displays a greatly attenuated interaction with Mrg15. Knock-in flies bearing this mutation display multiple homeotic transformation phenotypes, and these phenotypes are partially rescued by overexpressing the Mrg15-Nurf55 fusion protein, which stabilizes the association of Mrg15 with Ash1. In summary, Mrg15 is a subunit of the Ash1 complex, a stimulator of Ash1 enzymatic activity and a critical regulator of the TrxG protein function of Ash1 in Drosophila.
Cattoni, D. I., Gizzi, A. M. C., Georgieva, M., Di Stefano, M., Valeri, A., Chamousset, D., Houbron, C., Dejardin, S., Fiche, J. B., Gonzalez, I., Chang, J. M., Sexton, T., Marti-Renom, M. A., Bantignies, F., Cavalli, G. and Nollmann, M. (2017). Single-cell absolute contact probability detection reveals chromosomes are organized by multiple low-frequency yet specific interactions. Nat Commun 8(1): 1753. PubMed ID: 29170434
At the kilo- to megabase pair scales, eukaryotic genomes are partitioned into self-interacting modules or topologically associated domains (TADs) that associate to form nuclear compartments. This study combined high-content super-resolution microscopies with state-of-the-art DNA-labeling methods to reveal the variability in the multiscale organization of the Drosophila genome. Association frequencies within TADs and between TAD borders were found to be below ~10%, independently of TAD size, epigenetic state, or cell type. Critically, despite this large heterogeneity, it was possible to visualize nanometer-sized epigenetic domains at the single-cell level. In addition, absolute contact frequencies within and between TADs are to a large extent defined by genomic distance, higher-order chromosome architecture, and epigenetic identity. It is proposed that TADs and compartments are organized by multiple, small-frequency, yet specific interactions that are regulated by epigenetics and transcriptional state.

Tuesday, December 26th

Chen, X. and Dickman, D. (2017). Development of a tissue-specific ribosome profiling approach in Drosophila enables genome-wide evaluation of translational adaptations. PLoS Genet 13(12): e1007117. PubMed ID: 29194454
Recent advances in next-generation sequencing approaches have revolutionized understanding of transcriptional expression in diverse systems. However, measurements of transcription do not necessarily reflect gene translation, the process of ultimate importance in understanding cellular function. To circumvent this limitation, biochemical tagging of ribosome subunits to isolate ribosome-associated mRNA has been developed. However, this approach, called TRAP, lacks quantitative resolution compared to a superior technology, ribosome profiling. This study reports the development of an optimized ribosome profiling approach in Drosophila. First, successful ribosome profiling was demonstrate from a specific tissue, larval muscle, with enhanced resolution compared to conventional TRAP approaches. Next the ability of this technology to define genome-wide translational regulation was validated. This technology was leveraged to test the relative contributions of transcriptional and translational mechanisms in the postsynaptic muscle that orchestrate the retrograde control of presynaptic function at the neuromuscular junction. Surprisingly, no evidence was found that significant changes in the transcription or translation of specific genes are necessary to enable retrograde homeostatic signaling, implying that post-translational mechanisms ultimately gate instructive retrograde communication. Finally, it was shown that a global increase in translation induces adaptive responses in both transcription and translation of protein chaperones and degradation factors to promote cellular proteostasis. Together, this development and validation of tissue-specific ribosome profiling enables sensitive and specific analysis of translation in Drosophila.
Dear, M. L., Shilts, J. and Broadie, K. (2017). Neuronal activity drives FMRP- and HSPG-dependent matrix metalloproteinase function required for rapid synaptogenesis. Sci Signal 10(504). PubMed ID: 29114039
Matrix metalloproteinase (MMP) functions modulate synapse formation and activity-dependent plasticity. Aberrant MMP activity is implicated in fragile X syndrome (FXS), a disease caused by the loss of the RNA-binding protein FMRP and characterized by neurological dysfunction and intellectual disability. Gene expression studies in Drosophila suggest that Mmps cooperate with the heparan sulfate proteoglycan (HSPG) glypican co-receptor Dally-like protein (Dlp) to restrict trans-synaptic Wnt signaling and that synaptogenic defects in the fly model of FXS are alleviated by either inhibition of Mmp or genetic reduction of Dlp. This study used the Drosophila neuromuscular junction (NMJ) glutamatergic synapse to test activity-dependent Dlp and Mmp intersections in the context of FXS. Rapid, activity-dependent synaptic bouton formation depended on secreted Mmp1. Acute neuronal stimulation reduced the abundance of Mmp2 but increased that of both Mmp1 and Dlp, as well as enhanced the colocalization of Dlp and Mmp1 at the synapse. Dlp function promoted Mmp1 abundance, localization, and proteolytic activity around synapses. Dlp glycosaminoglycan (GAG) chains mediated this functional interaction with Mmp1. In the FXS fly model, activity-dependent increases in Mmp1 abundance and activity were lost but were restored by reducing the amount of synaptic Dlp. The data suggest that neuronal activity-induced, HSPG-dependent Mmp regulation drives activity-dependent synaptogenesis and that this is impaired in FXS. Thus, exploring this mechanism further may reveal therapeutic targets that have the potential to restore synaptogenesis in FXS patients.
Das, S., Trona, F., Khallaf, M. A., Schuh, E., Knaden, M., Hansson, B. S. and Sachse, S. (2017). Electrical synapses mediate synergism between pheromone and food odors in Drosophila melanogaster. Proc Natl Acad Sci U S A 114(46): E9962-e9971. PubMed ID: 29087946
In Drosophila melanogaster, the sex pheromone produced by males, cis-vaccenyl acetate (cVA), evokes a stereotypic gender-specific behavior in both males and females. As Drosophila adults feed, mate, and oviposit on food, they perceive the pheromone as a blend against a background of food odors. Previous studies have reported that food odors enhance flies' behavioral response to cVA, specifically in virgin females. However, how and where the different olfactory inputs interact has so far remained unknown. This study has elucidated the neuronal mechanism underlying the response at an anatomical, functional, and behavioral level. The data show that in virgin females cVA and the complex food odor vinegar evoke a synergistic response in the cVA-responsive glomerulus DA1. This synergism, however, does not appear at the input level of the glomerulus, but is restricted to the projection neuron level only. Notably, it is abolished by a mutation in gap junctions in projection neurons and is found to be mediated by electrical synapses between excitatory local interneurons and projection neurons. As a behavioral consequence, this study demonstrated that virgin females in the presence of vinegar become receptive more rapidly to courting males, while male courtship is not affected. Altogether, these results suggest that lateral excitation via gap junctions modulates odor tuning in the antennal lobe and drives synergistic interactions between two ecologically relevant odors, representing food and sex.
Saburova, E. A., Vasiliev, A. N., Kravtsova, V. V., Ryabova, E. V., Zefirov, A. L., Bolshakova, O. I., Sarantseva, S. V. and Krivoi, I. I. (2017). Human APP Gene Expression Alters Active Zone Distribution and Spontaneous Neurotransmitter Release at the Drosophila Larval Neuromuscular Junction. Neural Plast 2017: 9202584. PubMed ID: 28770114
This study provides further insight into the molecular mechanisms that control neurotransmitter release. Experiments were performed on larval neuromuscular junctions of transgenic Drosophila melanogaster lines with different levels of human amyloid precursor protein (APP) production. To express human genes in motor neurons of Drosophila, the UAS-GAL4 system was used. Human APP gene expression increased the number of synaptic boutons per neuromuscular junction. The total number of active zones, detected by Bruchpilot protein puncta distribution, remained unchanged; however, the average number of active zones per bouton decreased. These disturbances were accompanied by a decrease in frequency of miniature excitatory junction potentials without alteration in random nature of spontaneous quantal release. Similar structural and functional changes were observed with co-overexpression of human APP and beta-secretase genes. In Drosophila line with expression of human amyloid-beta42 peptide itself, parameters analyzed did not differ from controls, suggesting the specificity of APP effects. These results confirm the involvement of APP in synaptogenesis and provide evidence to suggest that human APP overexpression specifically disturbs the structural and functional organization of active zone and results in altered Bruchpilot distribution and lowered probability of spontaneous neurotransmitter release.
Goel, P., Li, X. and Dickman, D. (2017). Disparate postsynaptic induction mechanisms ultimately converge to drive the retrograde enhancement of presynaptic efficacy. Cell Rep 21(9): 2339-2347. PubMed ID: 29186673
Retrograde signaling systems are fundamental modes of communication synapses utilize to dynamically and adaptively modulate activity. However, the inductive mechanisms that gate retrograde communication in the postsynaptic compartment remain enigmatic. This study investigated retrograde signaling at the Drosophila neuromuscular junction, where three seemingly disparate perturbations to the postsynaptic cell trigger a similar enhancement in presynaptic neurotransmitter release. This study shows that the same presynaptic genetic machinery and enhancements in active zone structure are utilized by each inductive pathway. However, all three induction mechanisms differ in temporal, translational, and CamKII activity requirements to initiate retrograde signaling in the postsynaptic cell. Intriguingly, pharmacological blockade of postsynaptic glutamate receptors, and not calcium influx through these receptors, is necessary and sufficient to induce rapid retrograde homeostatic signaling through CamKII. Thus, three distinct induction mechanisms converge on the same retrograde signaling system to drive the homeostatic strengthening of presynaptic neurotransmitter release.
Csizmadia, T., Lorincz, P., Hegedus, K., Szeplaki, S., Low, P. and Juhasz, G. (2017). Molecular mechanisms of developmentally programmed crinophagy in Drosophila. J Cell Biol [Epub ahead of print]. PubMed ID: 29066608
At the onset of metamorphosis, Drosophila salivary gland cells undergo a burst of glue granule secretion to attach the forming pupa to a solid surface. This study shows that excess granules evading exocytosis are degraded via direct fusion with lysosomes, a secretory granule-specific autophagic process known as crinophagy. This study found that the tethering complex HOPS (homotypic fusion and protein sorting); the small GTPases Rab2, Rab7, and its effector, PLEKHM1; and a SNAP receptor complex consisting of Syntaxin 13, Snap29, and Vamp7 are all required for the fusion of secretory granules with lysosomes. Proper glue degradation within lysosomes also requires the Uvrag-containing Vps34 lipid kinase complex and the v-ATPase proton pump, whereas Atg genes involved in macroautophagy are dispensable for crinophagy. This work establishes the molecular mechanism of developmentally programmed crinophagy in Drosophila and paves the way for analyzing this process in metazoans.

Friday, December 22nd

Coutinho-Budd, J. C., Sheehan, A. E. and Freeman, M. R. (2017). The secreted neurotrophin Spatzle 3 promotes glial morphogenesis and supports neuronal survival and function. Genes Dev 31(20): 2023-2038. PubMed ID: 29138279
Most glial functions depend on establishing intimate morphological relationships with neurons. Significant progress has been made in understanding neuron-glia signaling at synaptic and axonal contacts, but how glia support neuronal cell bodies is unclear. This study explored the growth and functions of Drosophila cortex glia (which associate almost exclusively with neuronal cell bodies) to understand glia-soma interactions. Cortex glia were shown to tile with one another and with astrocytes to establish unique central nervous system (CNS) spatial domains that actively restrict glial growth, and selective ablation of cortex glia causes animal lethality. In an RNAi-based screen, alphaSNAP (soluble NSF [N-ethylmalemeide-sensitive factor] attachment protein alpha) and several components of vesicle fusion and recycling machinery were identified as essential for the maintenance of cortex glial morphology and continued contact with neurons. Interestingly, loss of the secreted neurotrophin Spatzle 3 (Spz3) phenocopied alphaSNAP phenotypes, which included loss of glial ensheathment of neuron cell bodies, increased neuronal cell death, and defects in animal behavior. Rescue experiments suggest that Spz3 can exert these effects only over very short distances. This work identifies essential roles for glial ensheathment of neuronal cell bodies in CNS homeostasis as well as Spz3 as a novel signaling factor required for maintenance of cortex glial morphology and neuron-glia contact.
Das, R., Bhattacharjee, S., Patel, A. A., Harris, J. M., Bhattacharya, S., Letcher, J. M., Clark, S. G., Nanda, S., Iyer, E. P. R., Ascoli, G. A. and Cox, D. N. (2017). Dendritic cytoskeletal architecture is modulated by combinatorial transcriptional regulation in Drosophila melanogaster. Genetics [Epub ahead of print]. PubMed ID: 29025914
Studies in Drosophila melanogaster have demonstrated that the conserved transcription factors (TFs) Cut and Knot exert combinatorial control over aspects of dendritic cytoskeleton development, promoting actin- and MT-based arbor morphology, respectively. To investigate transcriptional targets of Cut and/or Knot regulation, systematic neurogenomic studies were conducted, coupled with in vivo genetic screens utilizing multi-fluor cytoskeletal and membrane marker reporters. These analyses identified a host of putative Cut and/or Knot effector molecules and a subset of these putative TF targets converge on modulating dendritic cytoskeletal architecture and are grouped into three major phenotypic categories, based upon neuromorphometric analyses:-- complexity enhancer, complexity shifter, and complexity suppressor. Complexity enhancer genes normally function to promote higher order dendritic growth and branching with variable effects on MT stabilization and F-actin organization, whereas complexity shifter and complexity suppressor genes normally function in regulating proximal-distal branching distribution or in restricting higher order branching complexity, respectively, with spatially restricted impacts on the dendritic cytoskeleton. Collectively, this study implicate novel genes and cellular programs by which TFs distinctly and combinatorially govern dendritogenesis via cytoskeletal modulation.
Chen, D., Dale, R. K. and Lei, E. P. (2017). Shep regulates Drosophila neuronal remodeling by controlling transcription of its chromatin targets. Development [Epub ahead of print]. PubMed ID: 29158441
Neuronal remodeling is critical to form the mature nervous system and can lead to neuropsychiatric diseases when disrupted. Global gene expression changes in neurons during remodeling as well as the factors that regulate these changes remain poorly defined. To elucidate this process, RNA-seq was performed on isolated Drosophila larval and pupal neurons and up-regulated synaptic signaling and down-regulated gene expression regulators were found as a result of normal neuronal metamorphosis. The role was tested of alan shepard (shep), which encodes an evolutionarily conserved RNA-binding protein required for proper neuronal remodeling. Depletion of shep in neurons prevents the execution of metamorphic gene expression patterns, and shep-regulated genes correspond to Shep chromatin and/or RNA-binding targets. Reduced expression of a Shep-inhibited target gene brat is sufficient to rescue neuronal remodeling defects of shep knockdown flies. These results reveal direct regulation of transcriptional programs by Shep to regulate neuronal remodeling during metamorphosis.
Chen, Y. C., Mishra, D., Glass, S. and Gerber, B. (2017). Behavioral evidence for enhanced processing of the minor component of binary odor mixtures in larval Drosophila. Front Psychol 8: 1923. PubMed ID: 29163299
A fundamental problem in deciding between mutually exclusive options is that the decision needs to be categorical although the properties of the options often differ but in grade. In this study, larval Drosophila were trained such that in one set of animals odor A was rewarded, but odor B was not (A+/B), whereas a second set of animals was trained reciprocally (A/B+). The preference was tested of the larvae, either for A, or for B, or for "morphed" mixtures of A and B, that is for mixtures differing in the ratio of the two components. As expected, the larvae showed higher preference when only the previously rewarded odor was presented than when only the previously unrewarded odor was presented. For mixtures of A and B that differed in the ratio of the two components, the major component dominated preference behavior-but it dominated less than expected from a linear relationship between mixture ratio and preference behavior. This suggests that a minor component can have an enhanced impact in a mixture, relative to such a linear expectation. The current paradigm may prove useful in understanding how nervous systems generate discrete outputs in the face of inputs that differ only gradually.

Thursday, December 21st

Cai, X., Akber, M., Spirov, A. and Baumgartner, S. (2017). Cortical movement of Bicoid in early Drosophila embryos is actin- and microtubule-dependent and disagrees with the SDD diffusion model. PLoS One 12(10): e0185443. PubMed ID: 28973031
The SDD model (synthesis, diffusion, degradation) proposes that the bicoid (bcd) mRNA is located at the anterior pole of the embryo at all times and serves a source for translation of the Bicoid protein, coupled with diffusion and uniform degradation throughout the embryo. The ARTS model (active RNA transport, synthesis) proposes the mRNA is transported at the cortex along microtubules to form a mRNA gradient which serves as template for the production of Bcd, hence little Bcd movement is involved. Bcd was found to move along the cortex and not in a broad front towards the posterior as the SDD model predicted. Embryos were subjected to hypoxia where the mRNA remained strictly located at the tip at all times, while the protein was allowed to move freely, thus conforming to an ideal experimental setup to test the SDD model. Unexpectedly, Bcd still moved along the cortex. Moreover, cortical Bcd movement was sparse, even under longer hypoxic conditions. Hypoxic embryos treated with drugs compromising microtubule and actin function affected Bcd cortical movement and stability. Vinblastine treatment allowed the simulation of an ideal SDD model whereby the protein moved throughout the embryo in a broad front. In unfertilized embryos, the Bcd protein followed the mRNA which itself was transported into the interior of the embryo utilizing a hitherto undiscovered microtubular network. These data suggest that the Bcd gradient formation is probably more complex than previously anticipated.
Clement, R., Dehapiot, B., Collinet, C., Lecuit, T. and Lenne, P. F. (2017). Viscoelastic dissipation stabilizes cell shape changes during tissue morphogenesis. Curr Biol 27(20): 3132-3142. PubMed ID: 28988857
Tissue morphogenesis relies on the production of active cellular forces. Understanding how such forces are mechanically converted into cell shape changes is essential to understanding of morphogenesis. This study used myosin II pulsatile activity during Drosophila embryogenesis to study how transient forces generate irreversible cell shape changes. Analyzing the dynamics of junction shortening and elongation resulting from myosin II pulses, this study found that long pulses yield less reversible deformations, typically a signature of dissipative mechanics. This is consistent with a simple viscoelastic description, which was used to model individual shortening and elongation events. The model predicts that dissipation typically occurs on the minute timescale, a timescale commensurate with that of force generation by myosin II pulses. This estimate was tested by applying time-controlled forces on junctions with optical tweezers. Finally, it was shown that actin turnover participates in dissipation, as reducing it pharmacologically increases the reversibility of contractile events. These results argue that active junctional deformation is stabilized by actin-dependent dissipation. Hence, tissue morphogenesis requires coordination between force generation and dissipation.
Xie, W., Yang, Y., Gao, S., Song, T., Wu, Y., Li, D., Liu, M. and Zhou, J. (2017). The tumor suppressor CYLD controls epithelial morphogenesis and homeostasis by regulating mitotic spindle behavior and adherens junction assembly. J Genet Genomics 44(7): 343-353. PubMed ID: 28750888
The molecular mechanisms that contribute to the morphogenesis and homeostasis of the epithelium remain elusive. This study reports a novel role for the cylindromatosis (CYLD) tumor suppressor in these events. The results show that CYLD depletion disrupts epithelial organization in both Drosophila egg chambers and mouse skin and intestinal epithelia. Microscopic analysis of proliferating cells in mouse epithelial tissues and cultured organoids reveals that loss of CYLD synergizes with tumor-promoting agents to cause the misorientation of the mitotic spindle. Mechanistic studies show that CYLD accumulates at the cell cortex in epithelial tissues and cultured cells, where it promotes the formation of epithelial adherens junctions through the modulation of microtubule dynamics. These data suggest that CYLD controls epithelial morphogenesis and homeostasis by modulating the assembly of adherens junctions and ensuring proper orientation of the mitotic spindle. These findings thus provide novel insight into the role of CYLD in development, tissue homeostasis, and tumorigenesis.
Curran, S., Strandkvist, C., Bathmann, J., de Gennes, M., Kabla, A., Salbreux, G. and Baum, B. (2017). Myosin II controls junction fluctuations to guide epithelial tissue ordering. Dev Cell 43(4): 480-492. PubMed ID: 29107560
Under conditions of homeostasis, dynamic changes in the length of individual adherens junctions (AJs) provide epithelia with the fluidity required to maintain tissue integrity in the face of intrinsic and extrinsic forces. While the contribution of AJ remodeling to developmental morphogenesis has been intensively studied, less is known about AJ dynamics in other circumstances. AJ dynamics were studied in an epithelium that undergoes a gradual increase in packing order, without concomitant large-scale changes in tissue size or shape. Neighbor exchange events were found to be driven by stochastic fluctuations in junction length, regulated in part by junctional actomyosin. In this context, the developmental increase of isotropic junctional actomyosin reduces the rate of neighbor exchange, contributing to tissue order. A model is proposed in which the local variance in tension between junctions determines whether actomyosin-based forces will inhibit or drive the topological transitions that either refine or deform a tissue.

Wednesday, December 20th

Cavallin, M. et al., (2017). WDR81 mutations cause extreme microcephaly and impair mitotic progression in human fibroblasts and Drosophila neural stem cells. Brain 140(10): 2597-2609. PubMed ID: 28969387
Microlissencephaly is a rare brain malformation characterized by congenital microcephaly and lissencephaly. Microlissencephaly is suspected to result from abnormalities in the proliferation or survival of neural progenitors. Despite the recent identification of six genes involved in microlissencephaly, the pathophysiological basis of this condition remains poorly understood. This study performed trio-based whole exome sequencing in seven subjects from five non-consanguineous families who presented with either microcephaly or microlissencephaly. This led to the identification of compound heterozygous mutations in WDR81, a gene previously associated with cerebellar ataxia, intellectual disability and quadrupedal locomotion. Patient phenotypes ranged from severe microcephaly with extremely reduced gyration with pontocerebellar hypoplasia to moderate microcephaly with cerebellar atrophy. In patient fibroblast cells, WDR81 mutations were associated with increased mitotic index and delayed prometaphase/metaphase transition. Similarly, in vivo, knockdown of the WDR81 orthologue in Drosophila led to increased mitotic index of neural stem cells with delayed mitotic progression. In summary, this study highlights the broad phenotypic spectrum of WDR81-related brain malformations, which include microcephaly with moderate to extremely reduced gyration and cerebellar anomalies. The results suggest that WDR81 might have a role in mitosis that is conserved between Drosophila and humans.
Coyne, A. N., Lorenzini, I., Chou, C. C., Torvund, M., Rogers, R. S., Starr, A., Zaepfel, B. L., Levy, J., Johannesmeyer, J., Schwartz, J. C., Nishimune, H., Zinsmaier, K., Rossoll, W., Sattler, R. and Zarnescu, D. C. (2017). Post-transcriptional inhibition of Hsc70-4/HSPA8 expression leads to synaptic vesicle cycling defects in multiple models of ALS. Cell Rep 21(1): 110-125. PubMed ID: 28978466
Amyotrophic lateral sclerosis (ALS) is a synaptopathy accompanied by the presence of cytoplasmic aggregates containing TDP-43, an RNA-binding protein linked to approximately 97% of ALS cases. Using a Drosophila model of ALS, this study shows that TDP-43 overexpression (OE) in motor neurons results in decreased expression of the Hsc70-4 chaperone at the neuromuscular junction (NMJ). Mechanistically, mutant TDP-43 sequesters hsc70-4 mRNA and impairs its translation. Expression of the Hsc70-4 ortholog, HSPA8, is also reduced in primary motor neurons and NMJs of mice expressing mutant TDP-43. Electrophysiology, imaging, and genetic interaction experiments reveal TDP-43-dependent defects in synaptic vesicle endocytosis. These deficits can be partially restored by OE of Hsc70-4, cysteine-string protein (Csp), or dynamin. This suggests that TDP-43 toxicity results in part from impaired activity of the synaptic CSP/Hsc70 chaperone complex impacting dynamin function. Finally, Hsc70-4/HSPA8 expression is also post-transcriptionally reduced in fly and human induced pluripotent stem cell (iPSC) C9orf72 models, suggesting a common disease pathomechanism.
Berson, A., Sartoris, A., Nativio, R., Van Deerlin, V., Toledo, J. B., Porta, S., Liu, S., Chung, C. Y., Garcia, B. A., Lee, V. M., Trojanowski, J. Q., Johnson, F. B., Berger, S. L. and Bonini, N. M. (2017). TDP-43 promotes neurodegeneration by impairing chromatin remodeling. Curr Biol[Epub ahead of print]. PubMed ID: 29153328
Regulation of chromatin structure is critical for brain development and function. However, the involvement of chromatin dynamics in neurodegeneration is less well understood. This study found, launching from Drosophila models of amyotrophic lateral sclerosis and frontotemporal dementia, that TDP-43 impairs the induction of multiple key stress genes required to protect from disease by reducing the recruitment of the chromatin remodeler Chd1 to chromatin. Chd1 depletion robustly enhances TDP-43-mediated neurodegeneration and promotes the formation of stress granules. Conversely, upregulation of Chd1 restores nucleosomal dynamics, promotes normal induction of protective stress genes, and rescues stress sensitivity of TDP-43-expressing animals. TDP-43-mediated impairments are conserved in mammalian cells, and, importantly, the human ortholog CHD2 physically interacts with TDP-43 and is strikingly reduced in level in temporal cortex of human patient tissue. These findings indicate that TDP-43-mediated neurodegeneration causes impaired chromatin dynamics that prevents appropriate expression of protective genes through compromised function of the chromatin remodeler Chd1/CHD2. Enhancing chromatin dynamics may be a treatment approach to amyotrophic lateral scleorosis (ALS)/frontotemporal dementia (FTD).
Bardai, F. H., Wang, L., Mutreja, Y., Yenjerla, M., Gamblin, T. C. and Feany, M. B. (2017). A conserved cytoskeletal signaling cascade mediates neurotoxicity of FTDP-17 tau mutations in vivo. J Neurosci. PubMed ID: 29138281
The microtubule binding protein tau is strongly implicated in multiple neurodegenerative disorders, including frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17), which is caused by mutations in tau. In vitro, FTDP-17 mutant versions of tau can reduce microtubule binding and increase aggregation of tau, but the mechanism by which these mutations promote disease in vivo is not clear. This study took a combined biochemical and in vivo modeling approach to define functional properties of tau driving neurotoxicity in vivo. Wild type human tau and five FTDP-17 mutant forms of tau were expressed in Drosophila using a site-directed insertion strategy to ensure equivalent levels of expression. Multiple markers of neurodegeneration and neurotoxicity were analyzed in transgenic animals, including analysis of both males and females. FTDP-17 mutations act to enhance phosphorylation of tau and thus promote neurotoxicity in an in vivo setting. Further, it was demonstrated that phosphorylation-dependent excess stabilization of the actin cytoskeleton is a key phosphorylation-dependent mediator of the toxicity of wild type tau, and of all the FTDP-17 mutants tested. Finally, it was shown that important downstream pathways, including autophagy and the unfolded protein response, are co-regulated with neurotoxicity and actin cytoskeletal stabilization in brains of flies expressing wild type human and various FTDP-17 tau mutants, supporting a conserved mechanism of neurotoxicity of wild type tau and FTDP-17 mutant tau in disease pathogenesis.

Tuesday, December 19th

Chan, S. K. K., Cerda-Moya, G., Stojnic, R., Millen, K., Fischer, B., Fexova, S., Skalska, L., Gomez-Lamarca, M., Pillidge, Z., Russell, S. and Bray, S. J. (2017). Role of co-repressor genomic landscapes in shaping the Notch response. PLoS Genet 13(11): e1007096. PubMed ID: 29155828
Repressors are frequently deployed to limit the transcriptional response to signalling pathways. For example, several co-repressors interact directly with the DNA-binding protein CSL (Suppressor of Hairless in Drosophila) and are proposed to keep target genes silenced in the absence of Notch activity. However, the scope of their contributions remains unclear. To investigate co-repressor activity in the context of this well defined signalling pathway, this study analysed the genome-wide binding profile of the best-characterized CSL co-repressor in Drosophila, Hairless, and of a second CSL interacting repressor, SMRTER. As predicted there was significant overlap between Hairless and its CSL DNA-binding partner, both in Kc cells and in wing discs, where they were predominantly found in chromatin with active enhancer marks. However, while the Hairless complex was widely present at some Notch regulated enhancers in the wing disc, no binding was detected at others, indicating that it is not essential for silencing per se. Further analysis of target enhancers confirmed differential requirements for Hairless. SMRTER binding significantly overlapped with Hairless, rather than complementing it, and many enhancers were apparently co-bound by both factors. This analysis indicates that the actions of Hairless and SMRTER gate enhancers to Notch activity and to Ecdysone signalling respectively, to ensure that the appropriate levels and timing of target gene expression are achieved.
Court, H., Ahearn, I. M., Amoyel, M., Bach, E. A. and Philips, M. R. (2017). Regulation of NOTCH signaling by RAB7 and RAB8 requires carboxyl methylation by ICMT. J Cell Biol 216(12): 4165-4182. PubMed ID: 29051265
Isoprenylcysteine carboxyl methyltransferase (ICMT) methylesterifies C-terminal prenylcysteine residues of CaaX proteins and some RAB GTPases. Deficiency of either ICMT or NOTCH1 accelerates pancreatic neoplasia in Pdx1-Cre;LSL-Kras(G12D) mice, suggesting that ICMT is required for NOTCH signaling. This study used Drosophila melanogaster wing vein and scutellar bristle development to screen Rab proteins predicted to be substrates for ICMT (Ste14 in flies). Rab7 and Rab8 were identified as ICMT substrates that when silenced phenocopy ste14 deficiency. ICMT, RAB7, and RAB8 were all required for efficient NOTCH1 signaling in mammalian cells. Overexpression of RAB8 rescued NOTCH activation after ICMT knockdown both in U2OS cells expressing NOTCH1 and in fly wing vein development. ICMT deficiency induced mislocalization of GFP-RAB7 and GFP-RAB8 from endomembrane to cytosol, enhanced binding to RABGDI, and decreased GTP loading of RAB7 and RAB8. Deficiency of ICMT, RAB7, or RAB8 led to mislocalization and diminished processing of NOTCH1-GFP. Thus, NOTCH signaling requires ICMT in part because it requires methylated RAB7 and RAB8.
Casey, A. K., Moehlman, A. T., Zhang, J., Servage, K. A., Kramer, H. and Orth, K. (2017). Fic-mediated deAMPylation is not dependent on homo-dimerization and rescues toxic AMPylation in flies. J Biol Chem [Epub ahead of print]. PubMed ID: 29089387
Protein chaperones play a critical role in proteostasis. The activity of the major endoplasmic reticulum chaperone BiP (GRP78) is regulated by Fic-mediated AMPylation during resting states. By contrast, during times of stress, BiP is deAMPylated. This study shows that constitutive AMPylation by a constitutively-active FicE247G mutant is lethal in Drosophila. This lethality is cell autonomous as directed expression of the constitutively active mutant FicE247G to the fly eye does not kill the fly but rather results in a rough and reduced eye. These lethality and eye phenotypes are rescued by the deAMPylation activity of wild-type Fic. Consistent with Fic acting as a deAMPylation enzyme, its activity was both time and concentration dependent. Furthermore, Fic deAMPylation activity was sufficient to suppress the AMPylation activity mediated by the constitutively active FicE247G mutant in both Drosophila S2 cells and flies. Further, this study shows that the dual enzymatic activity of Fic is regulated by Fic dimerization as it is critical for AMPylation, but not deAMPylation, of BiP.
Chng, W. A., Koch, R., Li, X., Kondo, S., Nagoshi, E. and Lemaitre, B. (2017). Transforming Growth Factor beta/Activin signaling in neurons increases susceptibility to starvation. PLoS One 12(10): e0187054. PubMed ID: 29084242
Animals rely on complex signaling network to mobilize its energy stores during starvation. Previous work has shown that the sugar-responsive TGFbeta/Activin pathway, activated through the TGFbeta ligand Dawdle, plays a central role in shaping the post-prandial digestive competence in the Drosophila midgut. Nevertheless, little is known about the TGFbeta/Activin signaling in sugar metabolism beyond the midgut. This study addresses the importance of Dawdle (Daw) after carbohydrate ingestion. Daw expression was found to be coupled to dietary glucose through the evolutionarily conserved Mio-Mlx transcriptional complex. In addition, Daw activates the TGFbeta/Activin signaling in neuronal populations to regulate triglyceride and glycogen catabolism and energy homeostasis. Loss of those neurons depleted metabolic reserves and rendered flies susceptible to starvation.

Monday, December 18th

Fedina, T. Y., Arbuthnott, D., Rundle, H. D., Promislow, D. E. L. and Pletcher, S. D. (2017). Tissue-specific insulin signaling mediates female sexual attractiveness. PLoS Genet 13(8): e1006935. PubMed ID: 28817572
Global manipulation of insulin signaling, a nutrient-sensing pathway governing investment in survival versus reproduction, affects female sexual attractiveness in Drosophila. This study demonstrates that these effects on attractiveness derive from insulin signaling in the fat body and ovarian follicle cells, whose signals are integrated by pheromone-producing cells called oenocytes. Functional ovaries were required for global insulin signaling effects on attractiveness, and manipulations of insulin signaling specifically in late follicle cells recapitulated effects of global manipulations. Interestingly, modulation of insulin signaling in the fat body produced opposite effects on attractiveness, suggesting a competitive relationship with the ovary. Furthermore, all investigated tissue-specific insulin signaling manipulations that changed attractiveness also changed fecundity in the corresponding direction, pointing to insulin pathway activity as a reliable link between fecundity and attractiveness cues. The cues themselves, cuticular hydrocarbons, responded distinctly to fat body and follicle cell manipulations, indicating independent readouts of the pathway activity from these two tissues. Thus, this study describes a system in which female attractiveness results from an apparent connection between attractiveness cues and an organismal state of high fecundity, both of which are created by lowered insulin signaling in the fat body and increased insulin signaling in late follicle cells.
Chen, D., Tao, X., Zhou, L., Sun, F., Sun, M. and Fang, X. (2017). Spaghetti, a homolog of human RPAP3 (RNA polymerase II-associated protein 3), determines the fate of germline stem cells in Drosophila ovary. Cell Biol Int. PubMed ID: 29110400
The Drosophila ovary provides an attractive model for studying the extrinsic or intrinsic factors that regulate the fate of germline stem cells (GSCs). Using this model, this study identified a new role for Drosophila spaghetti (spag), encoding a homolog of human RNA polymerase II-associated protein 3 (RPAP3), in regulating the fate of ovarian GSCs. Results from spag knockdown and genetic mosaic studies suggest that spag functions as an intrinsic factor for GSCs maintenance. Loss of Spag by, either spag RNAi or null mutation failed to trigger apoptosis in ovarian GSCs. Overexpression of spag led to negligible increases in the number of GSC/Cystoblast (CB) cells, suggesting that an excess of Spag is not sufficient to accelerate the proliferation of GSCs or delay CBs' differentiation. This study provides evidence supporting that spag is involved in adult stem cells maintenance. In addition, the RNAi screen results showed that knockdown of Hsp90, one of known Spag interacting partners, led to loss of ovarian GSCs in Drosophila. Heterozygous mutations in hsp90 (hsp90/+) dramatically accelerated the GSC loss in spag RNAi ovaries, suggesting that the Spag-contained complex possibly plays an essential role in controlling the GSCs fate.
Bailey, A. S., Batista, P. J., Gold, R. S., Chen, Y. G., de Rooij, D. G., Chang, H. Y. and Fuller, M. T. (2017). The conserved RNA helicase YTHDC2 regulates the transition from proliferation to differentiation in the germline. Elife 6. PubMed ID: 29087293
The switch from mitosis to meiosis is the key event marking onset of differentiation in the germline stem cell lineage. In Drosophila, the translational repressor Bgcn is required for spermatogonia to stop mitosis and transition to meiotic prophase and the spermatocyte state. This study shows that the mammalian Bgcn homolog YTHDC2 facilitates a clean switch from mitosis to meiosis in mouse germ cells, revealing a conserved role for YTHDC2 in this critical cell fate transition. YTHDC2-deficient male germ cells enter meiosis but have a mixed identity, maintaining expression of Cyclin A2 and failing to properly express many meiotic markers. Instead of continuing through meiotic prophase, the cells attempt an abnormal mitotic-like division and die. YTHDC2 binds multiple transcripts including Ccna2 and other mitotic transcripts, binds specific piRNA precursors, and interacts with RNA granule components, suggesting that proper progression of germ cells through meiosis is licensed by YTHDC2 through post-transcriptional regulation.
Chen, D. Y., Crest, J. and Bilder, D. (2017). A cell migration tracking tool supports coupling of tissue rotation to elongation. Cell Rep 21(3): 559-569. PubMed ID: 29045826
Cell migration is indispensable to morphogenesis and homeostasis. Live imaging allows mechanistic insights, but long-term observation can alter normal biology, and tools to track movements in vivo without perturbation are lacking. This study developed a tool called M-TRAIL (matrix-labeling technique for real-time and inferred location), which reveals migration histories in fixed tissues. Using clones that overexpress GFP-tagged extracellular matrix (ECM) components, motility trajectories are mapped based on durable traces deposited onto basement membrane. M-TRAIL was applied to Drosophila follicle rotation, comparing in vivo and ex vivo migratory dynamics. The rate, trajectory, and cessation of rotation in wild-type (WT) follicles measured in vivo and ex vivo were identical, as was rotation failure in fat2 mutants. However, follicles carrying intracellularly truncated Fat2, previously reported to lack rotation ex vivo, in fact rotate in vivo at a reduced speed, thus revalidating the hypothesis that rotation is required for tissue elongation. The M-TRAIL approach could be applied to track and quantitate in vivo cell motility in other tissues and organisms.

Friday, December 15th

Brown, E. B., Patterson, C., Pancoast, R. and Rollmann, S. M. (2017). Artificial selection for odor-guided behavior in Drosophila reveals changes in food consumption. BMC Genomics 18(1): 867. PubMed ID: 29132294
Appropriate behavioral responses to the chemical cues of predators are important for organismal survival and can influence traits such as organismal life span and food consumption. However, understanding the genetic mechanisms underlying odor-guided behavior, correlated responses in other traits, and how these constrain or promote their evolution, remain an important challenge. This study performed artificial selection for attractive and aversive behavioral responses to four chemical compounds, two aromatics (4-ethylguaiacol and 4-methylphenol) and two esters (methyl hexanoate and ethyl acetate), for thirty generations. Artificial selection for odor-guided behavior revealed symmetrical responses to selection for each of the four chemical compounds. Next, whether selection for odor-guided behavior resulted in correlated responses in life history traits and/or food consumption. Changes were found in food consumption upon selection for behavioral responses to aromatics was tested. In many cases, lines selected for increased attraction to aromatics showed an increase in food consumption. RNA sequencing of lines selected for responses to 4-ethylguaiacol was performed to identify candidate genes associated with odor-guided behavior and its impact on food consumption. The study detected 91 genes that were differentially expressed among lines, many of which were associated with metabolic processes. RNAi-mediated knockdown of select candidate genes further supports their role in odor-guided behavior and/or food consumption. This study identifies novel genes underlying variation in odor-guided behavior and further elucidates the genetic mechanisms underlying the interrelationship between olfaction and feeding.
Chen, D., Kolomenskiy, D., Nakata, T. and Liu, H. (2017). Forewings match the formation of leading-edge vortices and dominate aerodynamic force production in revolving insect wings. Bioinspir Biomim [Epub ahead of print]. PubMed ID: 29052556
In many flying insects, forewings and hindwings are coupled mechanically to achieve flapping flight synchronously while being driven by action of the forewings. How the forewings and hindwings as well as their morphologies contribute to aerodynamic force production and flight control remains unclear. This study demonstrates that the forewings can produce most of the aerodynamic forces even with the hindwings removed through a computational fluid dynamic study of three revolving insect wing models, which are identical to the wing morphologies and Reynolds numbers of hawkmoth (Manduca sexta), bumblebee (Bombus ignitus) and fruitfly (Drosophila melanogaster). The forewing morphologies match the formation of leading-edge vortices (LEV) and are responsible for generating sufficient lift forces at the mean angles of attack and the Reynolds numbers where the three representative insects fly. The LEV formation and pressure loading keep almost unchanged with the hindwing removed, and even lead to some improvement in power factor and aerodynamic efficiency. Moreover, the results indicate that the size and strength of the LEVs can be well quantified with introduction of a conical LEV angle, which varies remarkably with angles of attack and Reynolds numbers but within the forewing region while showing less sensitivity to the wing morphologies. This implies that the forewing morphology very likely plays a dominant role in achieving low-Reynolds number aerodynamic performance in natural flyers as well as in revolving and/or flapping micro air vehicles.
Anreiter, I., Kramer, J. M. and Sokolowski, M. B. (2017). Epigenetic mechanisms modulate differences in Drosophila foraging behavior. Proc Natl Acad Sci U S A 114(47): 12518-12523. PubMed ID: 29078350
Little is known about how genetic variation and epigenetic marks interact to shape differences in behavior. The foraging (for) gene regulates behavioral differences between the rover and sitter Drosophila melanogaster strains, but the molecular mechanisms through which it does so have remained elusive. This study shows that the epigenetic regulator G9a interacts with for to regulate strain-specific adult foraging behavior through allele-specific histone methylation of a for promoter (pr4). Rovers have higher pr4 H3K9me dimethylation, lower pr4 RNA expression, and higher foraging scores than sitters. The rover-sitter differences disappear in the presence of G9a null mutant alleles, showing that G9a is necessary for these differences. Furthermore, rover foraging scores can be phenocopied by transgenically reducing pr4 expression in sitters. This compelling evidence shows that genetic variation can interact with an epigenetic modifier to produce differences in gene expression, establishing a behavioral polymorphism in Drosophila.
Bretman, A., Rouse, J., Westmancoat, J. D. and Chapman, T. (2017). The role of species-specific sensory cues in male responses to mating rivals in Drosophila melanogaster fruitflies. Ecol Evol 7(22): 9247-9256. PubMed ID: 29187965
Complex sets of cues can be important in recognizing and responding to conspecific mating competitors and avoiding potentially costly heterospecific competitive interactions. Drosophila males can detect sensory inputs from conspecifics to assess the level of competition. They respond to rivals by significantly extending mating duration and gain significant fitness benefits from doing so. This study tested the idea that the multiple sensory cues used by D. melanogaster males to detect conspecifics also function to minimize "off-target" responses to heterospecific males that they might encounter (Drosophila simulans, Drosophila yakuba, Drosophila pseudoobscura, or Drosophila virilis). Focal D. melanogaster males exposed to D. simulans or D. pseudoobscura subsequently increased mating duration, but to a lesser extent than following exposure to conspecific rivals. The magnitude of rivals' responses expressed by D. melanogaster males did not align with genetic distance between species, and none of the sensory manipulations caused D. melanogaster to respond to males of all other species tested. However, when "false" sensory cues were removed or provided, D. melanogaster males became more likely to show increased mating duration responses to heterospecific males. It is suggested that benefits of avoiding inaccurate assessment of the competitive environment may shape the evolution of recognition cues.

Thursday, December 14th

Kohzaki, H., Asano, M. and Murakami, Y. (2018). DNA replication machinery is required for development in Drosophila. Front Biosci (Landmark Ed) 23: 493-505. PubMed ID: 28930557
In Drosophila, some factors involved in chromosome replication seem to be involved in gene amplification and endoreplication, which are actively utilized in particular tissue development, but direct evidence has not been shown. Therefore, this study examined the effect of depletion of replication factors on these processes. First, it was confirmed that RNAi knockdown can be used for the depletion of replication factors by comparing the phenotypes of RNAi knockdown and deletion or point mutants of the components of DNA licensing factor, MCM2, MCM4 and Cdt1. Next, it was found that tissue-specific RNAi knockdown of replication factors caused tissue-specific defects, probably due to defects in DNA replication. In particular, depletion inhibited gene amplification of the chorion gene in follicle cells and endoreplication in salivary glands, showing that chromosomal DNA replication factors are required for these processes. Finally, using RNAi, the genes for chromosomal DNA replication that affected tissue development. Interestingly, wing specific knockdown of Mcm10 induced wing formation defects. These results suggest that some components of chromosomal replication machinery are directly involved in tissue development.
Hinnant, T. D., Alvarez, A. A. and Ables, E. T. (2017). Temporal remodeling of the cell cycle accompanies differentiation in the Drosophila germline. Dev Biol 429(1): 118-131. PubMed ID: 28711427
To describe how the cell cycle is remodeled in germ cells as they differentiate in situ, the Drosophila Fluorescence Ubiquitin-based Cell Cycle Indicator (Fly-FUCCI) system, in which degradable versions of GFP::E2f1 and RFP::CycB fluorescently label cells in each phase of the cell cycle was used. The lengths of the G1, S, and G2 phases of the cell cycle during oogenesis were found to change dramatically over the course of differentiation, and the 4/8-cell cyst was identified as a key developmental transition state in which cells prepare for specialized cell cycles. The data suggest that the transcriptional activator E2f1, which controls the transition from G1 to S phase, is a key regulator of mitotic divisions in the early germline. The data support the model that E2f1 is necessary for proper GSC proliferation, self-renewal, and daughter cell development. In contrast, while E2f1 degradation by the Cullin 4 (Cul4)-containing ubiquitin E3 ligase (CRL4) is essential for developmental transitions in the early germline, the data do not support a role for E2f1 degradation as a mechanism to limit GSC proliferation or self-renewal. Taken together, these findings provide further insight into the regulation of cell proliferation and the acquisition of differentiated cell fate, with broad implications across developing tissues.
Kim, T., Lara-Gonzalez, P., Prevo, B., Meitinger, F., Cheerambathur, D. K., Oegema, K. and Desai, A. (2017). Kinetochores accelerate or delay APC/C activation by directing Cdc20 to opposing fates. Genes Dev 31(11): 1089-1094. PubMed ID: 28698300
Evolutionary Homolog Study
Mitotic duration is determined by activation of the anaphase-promoting complex/cyclosome (APC/C) bound to its coactivator, Cdc20 (see Drosophila Fizzy. Kinetochores, the microtubule-interacting machines on chromosomes, restrain mitotic exit when not attached to spindle microtubules by generating a Cdc20-containing complex that inhibits the APC/C. This study, shows that flux of Cdc20 through kinetochores also accelerates mitotic exit by promoting its dephosphorylation by kinetochore-localized protein phosphatase 1 (see Drosophila Flapwing), which allows Cdc20 to activate the APC/C. Both APC/C activation and inhibition depend on Cdc20 fluxing through the same binding site at kinetochores. The microtubule attachment status of kinetochores therefore optimizes mitotic duration by controlling the balance between opposing Cdc20 fates.
Cahoon, C. K., Yu, Z., Wang, Y., Guo, F., Unruh, J. R., Slaughter, B. D. and Hawley, R. S. (2017). Superresolution expansion microscopy reveals the three-dimensional organization of the Drosophila synaptonemal complex. Proc Natl Acad Sci U S A 114(33): E6857-e6866. PubMed ID: 28760978
The synaptonemal complex (SC) assembles between homologous chromosomes and is essential for accurate chromosome segregation at the first meiotic division. In Drosophila, many SC components within the complex have been dissected through a combination of genetic analyses and superresolution and electron microscopy. The inability to optically resolve the minute distances between proteins in the complex has precluded its 3D characterization. A recently described technology termed expansion microscopy (ExM) uniformly increases the size of a biological sample, thereby circumventing the limits of optical resolution. By adapting the ExM protocol to render it compatible with structured illumination microscopy, it is possible to examine the 3D organization of several known Drosophila SC components. These data provide evidence that two layers of SC are assembled. It is further speculated that each SC layer may connect two nonsister chromatids, and a 3D model of the Drosophila SC is presented based on these findings.
Lane, S. I. R., Morgan, S. L., Wu, T., Collins, J. K., Merriman, J. A., ElInati, E., Turner, J. M. and Jones, K. T. (2017). DNA damage induces a kinetochore-based ATM/ATR-independent SAC arrest unique to the first meiotic division in mouse oocytes. Development 144(19): 3475-3486. PubMed ID: 28851706
Evolutionary Homolog Study
Mouse oocytes carrying DNA damage arrest in meiosis I, thereby preventing creation of embryos with deleterious mutations. The arrest is dependent on activation of the spindle assembly checkpoint, which results in anaphase-promoting complex (APC) inhibition. However, little is understood about how this checkpoint is engaged following DNA damage. This study found that within minutes of DNA damage checkpoint proteins are assembled at the kinetochore, not at damage sites along chromosome arms, such that the APC is fully inhibited within 30 min. Despite this robust response, there is no measurable loss in k-fibres, or tension across the bivalent. Through pharmacological inhibition this study observed that the response is dependent on Mps1 kinase (see Drosophila Mps1), aurora kinase (see Drosophila Aurora B) and Haspin (see Drosophila Haspin). Using oocyte-specific knockouts this study found the response does not require the DNA damage response kinases ATM or ATR. Furthermore, checkpoint activation does not occur in response to DNA damage in fully mature eggs during meiosis II, despite the divisions being separated by just a few hours. Therefore, mouse oocytes have a unique ability to sense DNA damage rapidly by activating the checkpoint at their kinetochores.
Overlack, K., Bange, T., Weissmann, F., Faesen, A. C., Maffini, S., Primorac, I., Muller, F., Peters, J. M. and Musacchio, A. (2017). BubR1 promotes Bub3-dependent APC/C inhibition during spindle assembly checkpoint signaling. Curr Biol 27(19): 2915-2927 e2917. PubMed ID: 28943088
Evolutionary Homolog Study
The spindle assembly checkpoint (SAC) prevents premature sister chromatid separation during mitosis. Phosphorylation of unattached kinetochores by the Mps1 kinase (see Drosophila Mps1) promotes recruitment of SAC machinery that catalyzes assembly of the SAC effector mitotic checkpoint complex (MCC). The SAC protein Bub3 (see Drosophila Bub3) is a phospho-amino acid adaptor that forms structurally related stable complexes with functionally distinct paralogs named Bub1 (see Drosophila Bub1) and BubR1 (see Drosophila Bub1R). A short motif ("loop") of Bub1, but not the equivalent loop of BubR1, enhances binding of Bub3 to kinetochore phospho-targets. This study asked whether the BubR1 loop directs Bub3 to different phospho-targets. The BubR1 loop is essential for SAC function and cannot be removed or replaced with the Bub1 loop. BubR1 loop mutants bind Bub3 and are normally incorporated in MCC in vitro but have reduced ability to inhibit the MCC target anaphase-promoting complex (APC/C), suggesting that BubR1:Bub3 recognition and inhibition of APC/C requires phosphorylation. Thus, small sequence differences in Bub1 and BubR1 direct Bub3 to different phosphorylated targets in the SAC signaling cascade.

Wednesday, December 13th

Baker, R., Nakamura, N., Chandel, I., Howell, B., Lyalin, D. and Panin, V. M. (2017). Protein O-mannosyltransferases affect sensory axon wiring and dynamic chirality of body posture in the Drosophila embryo. J Neurosci. PubMed ID: 29167399
Genetic defects in protein O-mannosyltransferases, POMT1 and POMT2, underlie severe muscular dystrophies. POMT genes are evolutionarily conserved in metazoan organisms. In Drosophila, both male and female POMT mutants show a clockwise rotation of adult abdominal segments, suggesting a chirality of underlying pathogenic mechanisms. This study describes and analyzes a similar phenotype in POMT mutant embryos that show left-handed body torsion. The experiments demonstrated that coordinated muscle contraction waves are associated with asymmetric embryo rolling, unveiling a new chirality marker in Drosophila development. Using genetic and live imaging approaches, it was revealed that the torsion phenotype results from differential rolling and aberrant patterning of peristaltic waves of muscle contractions. The results demonstrated that peripheral sensory neurons are required for normal contractions that prevent accumulation of torsion. POMT mutants show abnormal axonal connections of sensory neurons. POMT transgenic expression limited to sensory neurons significantly rescued the torsion phenotype, axonal connectivity defects and abnormal contractions in POMT mutant embryos. Taken together, these data suggested that protein O-mannosylation is required for normal sensory feedback to control coordinated muscle contractions and body posture. This mechanism may shed light on analogous functions of POMT genes in mammals and help elucidate etiology of neurological defects in muscular dystrophies.
Bozler, J., Kacsoh, B. Z., Chen, H., Theurkauf, W. E., Weng, Z. and Bosco, G. (2017). A systems level approach to temporal expression dynamics in Drosophila reveals clusters of long term memory genes. PLoS Genet 13(10): e1007054. PubMed ID: 29084214
The ability to integrate experiential information and recall it in the form of memory is observed in a wide range of taxa, and is a hallmark of highly derived nervous systems. Storage of past experiences is critical for adaptive behaviors that anticipate both adverse and positive environmental factors. The process of memory formation and consolidation involve many synchronized biological events including gene transcription, protein modification, and intracellular trafficking: However, many of these molecular mechanisms remain illusive. With Drosophila as a model system this study used a nonassociative memory paradigm and a systems level approach to uncover novel transcriptional patterns. RNA sequencing of Drosophila heads during and after memory formation identified a number of novel memory genes. Tracking the dynamic expression of these genes over time revealed complex gene networks involved in long term memory. In particular, this study focuses on two functional gene clusters of signal peptides and proteases. Bioinformatics network analysis and prediction in combination with high-throughput RNA sequencing identified previously unknown memory genes, which when genetically knocked down resulted in behaviorally validated memory defects.
Bahrampour, S., Gunnar, E., Jonsson, C., Ekman, H. and Thor, S. (2017). Neural lineage progression controlled by a temporal proliferation program. Dev Cell 43(3): 332-348.e334. PubMed ID: 29112852
Great progress has been made in identifying transcriptional programs that establish stem cell identity. In contrast, only limited insight has been gained into how these programs are down-graded in a timely manner to halt proliferation and allow for cellular differentiation. Drosophila embryonic neuroblasts undergo such a temporal progression, initially dividing to bud off daughters that divide once (type I), then switching to generating non-dividing daughters (type 0), and finally exiting the cell cycle. This study identifies six early transcription factors that drive neuroblast and type I daughter proliferation. Early factors are gradually replaced by three late factors, acting to trigger the type I-->0 daughter proliferation switch and eventually to stop neuroblasts. Early and late factors regulate each other and four key cell-cycle genes, providing a logical genetic pathway for these transitions. The identification of this extensive driver-stopper temporal program controlling neuroblast lineage progression may have implications for studies in many other systems.
Cao, L. H., Yang, D., Wu, W., Zeng, X., Jing, B. Y., Li, M. T., Qin, S., Tang, C., Tu, Y. and Luo, D. G. (2017). Odor-evoked inhibition of olfactory sensory neurons drives olfactory perception in Drosophila. Nat Commun 8(1): 1357. PubMed ID: 29116083
Inhibitory response occurs throughout the nervous system, including the peripheral olfactory system. While odor-evoked excitation in peripheral olfactory cells is known to encode odor information, the molecular mechanism and functional roles of odor-evoked inhibition remain largely unknown. This study examined Drosophila olfactory sensory neurons and found that inhibitory odors triggered outward receptor currents by reducing the constitutive activities of odorant receptors, inhibiting the basal spike firing in olfactory sensory neurons. Remarkably, this odor-evoked inhibition of olfactory sensory neurons elicited by itself a full range of olfactory behaviors from attraction to avoidance, as did odor-evoked olfactory sensory neuron excitation. These results indicated that peripheral inhibition is comparable to excitation in encoding sensory signals rather than merely regulating excitation. Furthermore, it was demonstrated that a bidirectional code with both odor-evoked inhibition and excitation in single olfactory sensory neurons increases the odor-coding capacity, providing a means of efficient sensory encoding.

Tuesday, December 12th

Bushnell, H. L., Feiler, C. E., Ketosugbo, K. F., Hellerman, M. B., Nazzaro, V. L. and Johnson, R. I. (2017). JNK is antagonized to ensure the correct number of interommatidial cells pattern the Drosophila retina. Dev Biol [Epub ahead of print]. PubMed ID: 29133184
Apoptosis is crucial during the morphogenesis of most organs and tissues, and is utilized for tissues to achieve their proper size, shape and patterning. Many signaling pathways contribute to the precise regulation of apoptosis. This study shows that Jun N-terminal Kinase (JNK) activity contributes to the coordinated removal of interommatidial cells via apoptosis in the Drosophila pupal retina. This is consistent with previous findings that JNK activity promotes apoptosis in other epithelia. However, JNK activity was found to be repressed by Cindr (the CIN85 and CD2AP ortholog) in order to promote cell survival. Reducing the amount of Cindr resulted in ectopic cell death. Increased expression of the Drosophila JNK basket in the setting of reduced cindr expression was found to result in even more severe apoptosis, whilst ectopic death was found to be reduced if retinas were heterozygous for basket. Hence Cindr is required to properly restrict JNK-mediated apoptosis in the pupal eye, resulting in the correct number of interommatidial cells. A lack of precise control over developmental apoptosis can lead to improper tissue morphogenesis.
Byrne, D. J., Harmon, M. J., Simpson, J. C., Blackstone, C. and O'Sullivan, N. C. (2017). Roles for the VCP co-factors Npl4 and Ufd1 in neuronal function in Drosophila melanogaster. J Genet Genomics 44(10): 493-501. PubMed ID: 29037990
The VCP-Ufd1-Npl4 complex regulates proteasomal processing within cells by delivering ubiquitinated proteins to the proteasome for degradation. Mutations in VCP are associated with two neurodegenerative diseases, amyotrophic lateral sclerosis (ALS) and inclusion body myopathy with Paget's disease of the bone and frontotemporal dementia (IBMPFD). Extensive study has revealed crucial functions of VCP within neurons. By contrast, little is known about the functions of Npl4 or Ufd1 in vivo. Using neuronal-specific knockdown of Npl4 or Ufd1 in Drosophila melanogaster, it is inferred that Npl4 contributes to microtubule organization within developing motor neurons. Moreover, Npl4 RNAi flies present with neurodegenerative phenotypes including progressive locomotor deficits, reduced lifespan and increased accumulation of TAR DNA-binding protein-43 homolog (TBPH). Knockdown, but not overexpression, of TBPH also exacerbates Npl4 RNAi-associated adult-onset neurodegenerative phenotypes. In contrast, this study finds that neuronal knockdown of Ufd1 has little effect on neuromuscular junction (NMJ) organization, TBPH accumulation or adult behaviour. These findings suggest the differing neuronal functions of Npl4 and Ufd1 in vivo.
Baumann, D. G., Dai, M. S., Lu, H. and Gilmour, D. S. (2017). GFZF, a glutathione S-transferase protein implicated in cell cycle regulation and hybrid inviability, is a transcriptional co-activator. Mol Cell Biol [Epub ahead of print]. PubMed ID: 29158293
The core promoter of protein-encoding genes plays a central role in regulating transcription. M1BP is a transcriptional activator that associates with a core promoter element known as Motif 1 that resides at thousand of genes in Drosophila. To gain insight into how M1BP functions, this study identified an interacting protein called GFZF. GFZF had been previously identified in genetic screens for factors involved in maintenance of hybrid inviability, the G2-M DNA damage checkpoint, and RAS/MAPK signaling but its contribution to these processes was unknown. This study shows that GFZF resides in the nucleus and functions as a transcriptional co-activator. In addition, GFZF is a glutathione S-transferase(GST). Thus, GFZF is the first transcriptional co-activator with intrinsic GST activity, and its identification as a transcriptional co-activator provides an explanation for its role in numerous biological processes.
Cao, J., Bollepalli, M. K., Hu, Y., Zhang, J., Li, Q., Li, H., Chang, H., Xiao, F., Hardie, R. C., Rong, Y. S. and Hu, W. (2017). A single residue mutation in the Galphaq subunit of the G protein complex causes blindness in Drosophila. G3 (Bethesda). PubMed ID: 29158337
Heterotrimeric G proteins play central roles in many signaling pathways, including the phototransduction cascade in animals. However, the degree of involvement of the G protein subunit Galphaq is not clear since animals with strong loss of function mutations previously reported remain responsive to light stimuli. This study recovered a new allele of Galphaq in Drosophila that abolishes light response in a conventional ERG assay, and reduces sensitivity in whole-cell recordings of dissociated cells by at least 5 orders of magnitude. In addition, mutant eyes demonstrate a rapid rate of degeneration in the presence of light. The new allele is likely the strongest hypomorph described to date. Interestingly, the mutant protein is produced in the eyes but carries a single amino acid change of a conserved hydrophobic residue that has been assigned to the interface of interaction between Galphaq and its downstream effector PLC. This study thus uncovered possibly the first point mutation that specifically affects this interaction in vivo.

Monday, December 11th

Krebs, A. R., Imanci, D., Hoerner, L., Gaidatzis, D., Burger, L. and Schubeler, D. (2017). Genome-wide single-molecule footprinting reveals high RNA Polymerase II turnover at paused promoters. Mol Cell 67(3): 411-422.e414. PubMed ID: 28735898
Transcription initiation entails chromatin opening followed by pre-initiation complex formation and RNA polymerase II recruitment. Subsequent polymerase elongation requires additional signals, resulting in increased residence time downstream of the start site, a phenomenon referred to as pausing. This study harnessed single-molecule footprinting to quantify distinct steps of initiation in vivo throughout the Drosophila genome. This identifies the impact of promoter structure on initiation dynamics in relation to nucleosomal occupancy. Additionally, perturbation of transcriptional initiation reveals an unexpectedly high turnover of polymerases at paused promoters-an observation confirmed at the level of nascent RNAs. These observations argue that absence of elongation is largely caused by premature termination rather than by stable polymerase stalling. In support of this non-processive model, it was observed that induction of the paused heat shock promoter depends on continuous initiation. This study provides a framework to quantify protein binding at single-molecule resolution and refines concepts of transcriptional pausing.
Bali, A. and Shravage, B. V. (2017). Characterization of the Autophagy related gene-8a (Atg8a) promoter in Drosophila melanogaster. Int J Dev Biol 61(8-9): 551-555. PubMed ID: 29139541
Autophagy is an evolutionarily conserved process which is upregulated under various stress conditions, including nutrient stress and oxidative stress. Amongst autophagy related genes (Atgs), Atg8a (LC3 in mammals) is induced several-fold during nutrient limitation in Drosophila. The minimal Atg8a cis-regulatory module (CRM) which mediates transcriptional upregulation under various stress conditions is not known. This study describes the generation and analyses of a series of Atg8a promoter deletions which drive the expression of an mCherry-Atg8a fusion cassette. Expression studies revealed that a 200 bp region of Atg8a is sufficient to drive expression of Atg8a in nutrient rich conditions in fat body and ovaries, as well as under nutrient deficient conditions in the fat body. Furthermore, this 200 bp region can mediate Atg8a upregulation during developmental histolysis of the larval fat body and under oxidative stress conditions induced by H2O2. Finally, the expression levels of Atg8a from this promoter are sufficient to rescue the lethality of the Atg8a mutant. The 200 bp promoter-fusion reporter provides a valuable tool which can be used in genetic screens to identify transcriptional and post-transcriptional regulators of Atg8a (Bali, 2017).
Barr, K. A., Martinez, C., Moran, J. R., Kim, A. R., Ramos, A. F. and Reinitz, J. (2017). Synthetic enhancer design by in silico compensatory evolution reveals flexibility and constraint in cis-regulation. BMC Syst Biol 11(1): 116. PubMed ID: 29187214
Models that incorporate specific chemical mechanisms have been successful in describing the activity of Drosophila developmental enhancers as a function of underlying transcription factor binding motifs. Despite this, the minimum set of mechanisms required to reconstruct an enhancer from its constituent parts is not known. Synthetic biology offers the potential to test the sufficiency of known mechanisms to describe the activity of enhancers, as well as to uncover constraints on the number, order, and spacing of motifs. Using a functional model and in silico compensatory evolution, putative synthetic even-skipped stripe 2 enhancers with varying degrees of similarity to the natural enhancer. These elements represent the evolutionary trajectories of the natural stripe 2 enhancer towards two synthetic enhancers designed ab initio. In the first trajectory, spatially regulated expression was maintained, even after more than a third of binding sites were lost. In the second, sequences with high similarity to the natural element did not drive expression, but a highly diverged sequence about half the length of the minimal stripe 2 enhancer drove ten times greater expression. Additionally, homotypic clusters of Zelda or Stat92E motifs, but not Bicoid, drove expression in developing embryos. The results show that the gene regulation model explains much of the function of the stripe 2 enhancer. Cases where expression deviated from prediction indicates that undescribed factors likely act to modulate expression. Activation driven Bicoid and Hunchback is highly sensitive to spatial organization of binding motifs. In contrast, Zelda and Stat92E drive expression from simple homotypic clusters, suggesting that activation driven by these factors is less constrained. Collectively, the 40 sequences generated in this work provides a powerful training set for building future models of gene regulation.
Baudouin-Gonzalez, L., Santos, M. A., Tempesta, C., Sucena, E., Roch, F. and Tanaka, K. (2017). Diverse cis-regulatory mechanisms contribute to expression evolution of tandem gene duplicates. Mol Biol Evol 34(12): 3132-3147. PubMed ID: 28961967
Pairs of duplicated genes generally display a combination of conserved expression patterns inherited from their unduplicated ancestor and newly acquired domains. However, how the cis-regulatory architecture of duplicated loci evolves to produce these expression patterns is poorly understood. This study directly examined the gene-regulatory evolution of two tandem duplicates, the Drosophila Ly6 genes CG9336 and CG9338, which arose at the base of the drosophilids between 40 and 60 Ma. Comparing the expression patterns of the two paralogs in four Drosophila species with that of the unduplicated ortholog in the tephritid Ceratitis capitata, the study shows that they diverged from each other as well as from the unduplicated ortholog. Moreover, the expression divergence appears to have occurred close to the duplication event and also more recently in a lineage-specific manner. The comparison of the tissue-specific cis-regulatory modules (CRMs) controlling the paralog expression in the four Drosophila species indicates that diverse cis-regulatory mechanisms, including the novel tissue-specific enhancers, differential inactivation, and enhancer sharing, contributed to the expression evolution. This analysis also reveals a surprisingly variable cis-regulatory architecture, in which the CRMs driving conserved expression domains change in number, location, and specificity. Altogether, this study provides a detailed historical account that uncovers a highly dynamic picture of how the paralog expression patterns and their underlying cis-regulatory landscape evolve. It is argued that these findings will encourage studying cis-regulatory evolution at the whole-locus level to understand how interactions between enhancers and other regulatory levels shape the evolution of gene expression.

Friday, December 8th

Adewoye, A. B., Nuzhdin, S. V. and Tauber, E. (2017). Mapping quantitative trait loci underlying circadian light sensitivity in Drosophila. J Biol Rhythms 32(5): 394-405. PubMed ID: 28990443
Despite the significant advance in understanding of the molecular basis of light entrainment of the circadian clock in Drosophila, the underlying genetic architecture is still largely unknown. The aim of this study was to identify loci associated with variation in circadian photosensitivity, which are important for the evolution of this trait. Complementary approaches were used that combined quantitative trait loci (QTL) mapping, complementation testing, and transcriptome profiling to dissect this variation. A major QTL was identified on chromosome 2, which was subsequently fine mapped using deficiency complementation mapping into 2 smaller regions spanning 139 genes, some of which are known to be involved in functions that have been previously implicated in light entrainment. Two genes implicated with the clock and located within that interval, timeless and cycle, failed to complement the QTL, indicating that alleles of these genes contribute to the variation in light response. Specifically, the timeless s/ls polymorphism that has been previously shown to constitute a latitudinal cline in Europe is also segregating in the recombinant inbred lines and is contributing to the phenotypic variation in light sensitivity. This study also profiled gene expression in 2 recombinant inbred strains that differ significantly in their photosensitivity and a total of 368 transcripts were identified that showed differential expression (false discovery rate < 0.1). Of 131 transcripts that showed a significant recombinant inbred line by treatment interaction (i.e., putative expression QTL), 4 are located within QTL2.
Aw, W. C., Garvin, M. R., Melvin, R. G. and Ballard, J. W. O. (2017). Sex-specific influences of mtDNA mitotype and diet on mitochondrial functions and physiological traits in Drosophila melanogaster. PLoS One 12(11): e0187554. PubMed ID: 29166659
This study determined the sex-specific influence of mtDNA type (mitotype) and diet on mitochondrial functions and physiology in two Drosophila melanogaster lines. In many species, males and females differ in aspects of their energy production. These sex-specific influences may be caused by differences in evolutionary history and physiological functions. It was predicted the influence of mtDNA mutations should be stronger in males than females as a result of the organelle's maternal mode of inheritance in the majority of metazoans. In contrast, it was predicted the influence of diet would be greater in females due to higher metabolic flexibility. Four diets were included that differed in their protein: carbohydrate (P:C) ratios as they are the two-major energy-yielding macronutrients in the fly diet. Four mitochondrial function traits (Complex I oxidative phosphorylation, reactive oxygen species production, superoxide dismutase activity, and mtDNA copy number) and four physiological traits (fecundity, longevity, lipid content, and starvation resistance) were examined. Traits were assayed at 11 d and 25 d of age. Consistent with predictions it was observe that the mitotype influenced males more than females supporting the hypothesis of a sex-specific selective sieve in the mitochondrial genome caused by the maternal inheritance of mitochondria. Also, consistent with predictions, it was found that the diet influenced females more than males.
Shilova, V. Y., Zatsepina, O. G., Garbuz, D. G., Funikov, S. Y., Zelentsova, E. S., Schostak, N. G., Kulikov, A. M. and Evgen'ev, M. B. (2017). Heat shock protein 70 from a thermotolerant Diptera species provides higher thermoresistance to Drosophila larvae than correspondent endogenous gene. Insect Mol Biol. PubMed ID: 28796386
Heat shock proteins (Hsp70s) from two Diptera species that drastically differ in their heat shock response and longevity were investigated. Drosophila melanogaster is characterized by the absence of Hsp70 and other hsps under normal conditions and the dramatic induction of hsp synthesis after temperature elevation. The other Diptera species examined belongs to the Stratiomyidae family (Stratiomys singularior) and exhibits high levels of inducible Hsp70 under normal conditions coupled with a thermotolerant phenotype and much longer lifespan. To evaluate the impact of hsp70 genes on thermotolerance and longevity, use was made of a D. melanogaster strain that lacks all hsp70 genes. Single copies of either S. singularior or D. melanogaster hsp70 were introduced into this strain and the transgenic flies wer examined in terms of thermotolerance and longevity. Transgenic strains were developed containing the S. singularior hsp70 gene under control of a D. melanogaster hsp70 promoter. Although these adult flies did synthesize the corresponding mRNA after heat shock, they were not superior to the flies containing a single copy of D. melanogaster hsp70 in thermotolerance and longevity. By contrast, Stratiomyidae Hsp70 provided significantly higher thermotolerance at the larval stage in comparison with endogenous Hsp70.
Abrat, O. B., Storey, J. M., Storey, K. B. and Lushchak, V. I. (2018). High amylose starch consumption induces obesity in Drosophila melanogaster and metformin partially prevents accumulation of storage lipids and shortens lifespan of the insects. Comp Biochem Physiol A Mol Integr Physiol 215: 55-62. PubMed ID: 29054808
There are very few studies that have directly analyzed the effects of dietary intake of slowly digestible starches on metabolic parameters of animals. The present study examined the effects of slowly digestible starch with high amylose content (referred also as amylose starch) either alone, or in combination with metformin on the development, lifespan, and levels of glucose and storage lipids in the fruit fly Drosophila melanogaster. Consumption of amylose starch in concentrations 0.25-10% did not affect D. melanogaster development, whereas 20% starch delayed pupation and reduced the number of larvae that reached the pupal stage. Starch levels in larval food, but not in adult food, determined levels of triacylglycerides in eight-day-old adult flies. Rearing on diet with 20% starch led to shorter lifespan and a higher content of triacylglycerides in the bodies of adult flies as compared with the same parameters in flies fed on 4% starch diet. Food supplementation with 10mM metformin partly attenuated the negative effects of high starch concentrations on larval pupation and decreased triacylglyceride levels in adult flies fed on 20% starch. Long-term consumption of diets supplemented with metformin and starch decreased lifespan of the insects, compared with the diet supplemented with starch only. The data show that in Drosophila high starch consumption may induce a fat fly phenotype and metformin may partially prevent it.

Thursday, December 7th

Molina-Mateo, D., Fuenzalida-Uribe, N., Hidalgo, S., Molina-Fernandez, C., Abarca, J., Zarate, R. V., Escandon, M., Figueroa, R., Tevy, M. F. and Campusano, J. M. (2017). Characterization of a presymptomatic stage in a Drosophila Parkinson's disease model: Unveiling dopaminergic compensatory mechanisms. Biochim Biophys Acta [Epub ahead of print]. PubMed ID: 28716706
Parkinson disease (PD) is a degenerative disorder characterized by several motor symptoms including shaking, rigidity, slow movement and difficult walking, which has been associated to the death of nigro-striatal dopaminergic neurons. >90% of PD patients also present olfactory dysfunction. Although the molecular mechanisms responsible for this disease are not clear, hereditary PD is linked to mutations in specific genes, including the PTEN-induced putative kinase 1 (PINK1). This work provides a thorough temporal description of the behavioral effects induced by a mutation in the PINK1 gene in adult Drosophila. The data suggests that the motor deficits associated to PD are fully revealed only by the third week of age. However, olfactory dysfunction is detected as early as the first week of age. Immunofluorescence and neurochemical data is provided that led to a proposal that compensatory changes occur in this Drosophila model for PD. These compensatory changes are associated to specific components of the dopaminergic system: the biosynthetic enzymes, Tyrosine hydroxylase and Dopa decarboxylase, and the Dopamine transporter, a plasma membrane protein involved in maintaining dopamine extracellular levels at physiologically relevant levels. Thus, these data help define presymptomatic and symptomatic phases in this PD animal model, and that compensatory changes occur in the dopaminergic neurons in the presymptomatic stage.
Lee, B. I., Suh, Y. S., Chung, Y. J., Yu, K. and Park, C. B. (2017). Shedding light on Alzheimer's beta-Amyloidosis: Photosensitized methylene blue inhibits self-assembly of beta-amyloid peptides and disintegrates their aggregates. Sci Rep 7(1): 7523. PubMed ID: 28790398
Abnormal aggregation of beta-amyloid (Abeta) peptides is a major hallmark of Alzheimer's disease (AD). In spite of numerous attempts to prevent the beta-amyloidosis, no effective drugs for treating AD have been developed to date. Among many candidate chemicals, methylene blue (MB) has proved its therapeutic potential for AD in a number of in vitro and in vivo studies; but the result of recent clinical trials performed with MB and its derivative was negative. In this study, with the aid of multiple photochemical analyses, it is reported that photoexcited MB molecules can block Abeta42 aggregation in vitro. Furthermore, an in vivo study using Drosophila AD model demonstrates that photoexcited MB is highly effective in suppressing synaptic toxicity, resulting in a reduced damage to the neuromuscular junction (NMJ), an enhanced locomotion, and decreased vacuole in the brain. The hindrance effect is attributed to Abeta42 oxidation by singlet oxygen (1O2) generated from photoexcited MB. Finally, it was shown that photoexcited MB possess a capability to disaggregate the pre-existing Abeta42 aggregates and reduce Abeta-induced cytotoxicity. This work suggests that light illumination can provide an opportunity to boost the efficacies of MB toward photodynamic therapy of AD in future.
Avery, A. W., Thomas, D. D. and Hays, T. S. (2017). beta-III-spectrin spinocerebellar ataxia type 5 mutation reveals a dominant cytoskeletal mechanism that underlies dendritic arborization. Proc Natl Acad Sci U S A 114(44): E9376-e9385. PubMed ID: 29078305
A spinocerebellar ataxia type 5 (SCA5) L253P mutation in the actin-binding domain (ABD) of beta-III-spectrin causes high-affinity actin binding and decreased thermal stability in vitro. This study shows in mammalian cells, at physiological temperature, that the mutant ABD retains high-affinity actin binding. Significantly, evidence is provided that the mutation alters the mobility and recruitment of beta-III-spectrin in mammalian cells, pointing to a potential disease mechanism. To explore this mechanism, a Drosophila SCA5 model was developed in which an equivalent mutant Drosophila beta-spectrin is expressed in neurons that extend complex dendritic arbors, such as Purkinje cells, targeted in SCA5 pathogenesis. The mutation causes a proximal shift in arborization coincident with decreased beta-spectrin localization in distal dendrites. SCA5 beta-spectrin dominantly mislocalizes alpha-spectrin and ankyrin-2, components of the endogenous spectrin cytoskeleton. These data suggest that high-affinity actin binding by SCA5 beta-spectrin interferes with spectrin-actin cytoskeleton dynamics, leading to a loss of a cytoskeletal mechanism in distal dendrites required for dendrite stabilization and arbor outgrowth.
Arnes, M., Casas-Tinto, S., Malmendal, A. and Ferrus, A. (2017). Amyloid beta42 peptide is toxic to non-neural cells in Drosophila yielding a characteristic metabolite profile and the effect can be suppressed by PI3K. Biol Open 6(11): 1664-1671. PubMed ID: 29141953
The human Abeta42 peptide is associated with Alzheimer's disease through its deleterious effects in neurons. Expressing the human peptide in adult Drosophila in a tissue- and time-controlled manner, this study shows that Abeta42 is also toxic in non-neural cells, neurosecretory and epithelial cell types in particular. This form of toxicity includes the aberrant signaling by Wingless morphogen leading to the eventual activation of Caspase 3. Preventing Caspase 3 activation by means of p53 keeps epithelial cells from elimination but maintains the Abeta42 toxicity yielding more severe deleterious effects to the organism. Metabolic profiling by nuclear magnetic resonance (NMR) of adult flies at selected ages post Abeta42 expression onset reveals characteristic changes in metabolites as early markers of the pathological process. All morphological and most metabolic features of Abeta42 toxicity can be suppressed by the joint overexpression of PI3K.

Wednesday, December 6th

Albert, E. A. and Bokel, C. (2017). A cell based, high throughput assay for quantitative analysis of Hedgehog pathway activation using a Smoothened activation sensor. Sci Rep 7(1): 14341. PubMed ID: 29085027
The Hedgehog (Hh) signalling cascade plays an important role in development and disease. In the absence of Hh ligand, activity of the key signal transducer Smoothened (Smo) is downregulated by the Hh receptor Patched (Ptc). However, the mechanisms underlying this inhibition, and especially its release upon ligand stimulation, are still poorly understood, in part because tools for following Smo activation at the subcellular level were long lacking. To address this deficit this study has developed a high throughput cell culture assay based on a fluorescent sensor for Drosophila Smo activation. A small molecule inhibitor library was screened, and increased Smo sensor fluorescence was observed with compounds aimed at two major target groups, the MAPK signalling cascade and Polo and Aurora kinases. Biochemical validation for selected inhibitors (dobrafenib, tak-733, volasertib) confirmed the screen results and revealed differences in the mode of Smo activation. Furthermore, monitoring Smo activation at the single cell level indicated that individual cells exhibit different threshold responses to Hh stimulation, which may be mechanistically relevant for the formation of graded Hh responses. Together, these results thus provide proof of principle that this assay may become a valuable tool for dissecting the cell biological basis of Hh pathway activation.
Narciso, C. E., Contento, N. M., Storey, T. J., Hoelzle, D. J. and Zartman, J. J. (2017). Release of applied mechanical loading stimulates intercellular calcium waves in Drosophila wing discs. Biophys J 113(2): 491-501. PubMed ID: 28746859
Mechanical forces are critical but poorly understood inputs for organogenesis and wound healing. Calcium ions (Ca2+) are critical second messengers in cells for integrating environmental and mechanical cues, but the regulation of Ca2+ signaling is poorly understood in developing epithelial tissues. This study reports a chip-based regulated environment for microorgans that enables systematic investigations of the crosstalk between an organ's mechanical stress environment and biochemical signaling under genetic and chemical perturbations. This method enabled definition of the essential conditions for generating organ-scale intercellular Ca2+ waves in Drosophila wing discs that are also observed in vivo during organ development. Mechanically induced intercellular Ca2+ waves are shown to require fly extract growth serum as a chemical stimulus. Using the chip-based regulated environment for microorgans, it was demonstrated that not the initial application but instead the release of mechanical loading is sufficient, but not necessary, to initiate intercellular Ca2+ waves. The Ca2+ response depends on the prestress intercellular Ca2+ activity and not on the magnitude or duration of the mechanical stimulation applied. Mechanically induced intercellular Ca2+ waves rely on IP3R-mediated Ca2+-induced Ca2+ release and propagation through gap junctions. Thus, intercellular Ca2+ waves in developing epithelia may be a consequence of stress dissipation during organ growth.
Matsushima, Y., Hirofuji, Y., Aihara, M., Yue, S., Uchiumi, T., Kaguni, L. S. and Kang, D. (2017). Drosophila protease ClpXP specifically degrades DmLRPPRC1 controlling mitochondrial mRNA and translation. Sci Rep 7(1): 8315. PubMed ID: 28814717
ClpXP is the major protease in the mitochondrial matrix in eukaryotes, and is well conserved among species. ClpXP is composed of a proteolytic subunit, ClpP, and a chaperone-like subunit, ClpX. Although it has been proposed that ClpXP is required for the mitochondrial unfolded protein response, additional roles for ClpXP in mitochondrial biogenesis are unclear. This study found that Drosophila leucine-rich pentatricopeptide repeat domain-containing protein 1 (DmLRPPRC1) is a specific substrate of ClpXP. Depletion or introduction of catalytically inactive mutation of ClpP increases DmLRPPRC1 and causes non-uniform increases of mitochondrial mRNAs, accumulation of some unprocessed mitochondrial transcripts, and modest repression of mitochondrial translation in Drosophila Schneider S2 cells. Moreover, DmLRPPRC1 over-expression induces the phenotypes similar to those observed when ClpP is depleted. Taken together, ClpXP regulates mitochondrial gene expression by changing the protein level of DmLRPPRC1 in Drosophila Schneider S2 cells.
Akhmetova, K., Balasov, M., Svitin, A., Chesnokova, E., Renfrow, M. and Chesnokov, I. (2017). Phosphorylation of Pnut at the early steps of Drosophila embryo development affects association of the Septin complex with membrane and is important for viability. G3 (Bethesda). PubMed ID: 29079679
Septin proteins are polymerizing GTPases that are found in most eukaryotic species. Septins are important for cytokinesis and participate in many processes involving spatial modifications of the cell cortex. In Drosophila, septin proteins Pnut, Sep1 and Sep2 form a hexameric septin complex. This study found that septin protein Pnut is phosphorylated during the first two hours of Drosophila embryo development. To study the effect of Pnut phosphorylation in a live organism, a new Drosophila pnut null mutant was created that allows for the analysis of Pnut mutations during embryogenesis. To understand the functional significance of Pnut phosphorylation, Drosophila strains carrying non-phosphorylatable and phospho-mimetic mutant pnut transgenes were established. The expression of the non-phosphorylatable Pnut protein resulted in semi-lethality and abnormal protein localization, whereas, the expression of the phospho-mimetic mutant form of Pnut disrupted the assembly of a functional septin complex and septin filament formation in vitro. Overall, these findings indicate that the controlled phosphorylation of Pnut plays an important role in regulating septin complex functions during organism development.

Tuesday, December 5th

Kaieda, Y., Masuda, R., Nishida, R., Shimell, M., O'Connor, M. B. and Ono, H. (2017). Glue protein production can be triggered by steroid hormone signaling independent of the developmental program in Drosophila melanogaster. Dev Biol 430(1): 166-176. PubMed ID: 28782527
Steroid hormones regulate life stage transitions, allowing animals to appropriately follow a developmental timeline. During insect development, the steroid hormone ecdysone is synthesized and released in a regulated manner by the prothoracic gland (PG) and then hydroxylated to the active molting hormone, 20-hydroxyecdysone (20E), in peripheral tissues. This study manipulated ecdysteroid titers, through temporally controlled over-expression of the ecdysteroid-inactivating enzyme, CYP18A1, in the PG using the GeneSwitch-GAL4 system in the fruit fly Drosophila melanogaster. Expression was monitored of a 20E-inducible glue protein gene, Salivary gland secretion 3 (Sgs3), using a Sgs3:GFP fusion transgene. In wild type larvae, Sgs3-GFP expression is activated at the midpoint of the third larval instar stage in response to the rising endogenous level of 20E. By first knocking down endogenous 20E levels during larval development and then feeding 20E to these larvae at various stages, it was found that Sgs3-GFP expression could be triggered at an inappropriate developmental stage after a certain time lag. This stage-precocious activation of Sgs3 required expression of the Broad-complex, similar to normal Sgs3 developmental regulation, and a small level of nutritional input. It is suggested that these studies provide evidence for a tissue-autonomic regulatory system for a metamorphic event independent from the primary 20E driven developmental progression.
Li, K., Zhang, X., Zuo, Y., Liu, W., Zhang, J. and Moussian, B. (2017). Timed Knickkopf function is essential for wing cuticle formation in Drosophila melanogaster. Insect Biochem Mol Biol 89: 1-10. PubMed ID: 28821399
The insect cuticle is an extracellular matrix that consists of the polysaccharide chitin, proteins, lipids and organic molecules that are arranged in distinct horizontal layers. In Drosophila melanogaster, these layers are not formed sequentially, but, at least partially, at the same time. Timing of the underlying molecular mechanisms is conceivably crucial for cuticle formation. To study this issue, the time period was determined during which the function of Knickkopf (Knk), a key factor of chitin organization, is required for wing cuticle differentiation in D. melanogaster. Although knk is expressed throughout metamorphosis, it was demonstrated that its expression 30 h prior and 48 h after pupariation is essential for correct wing cuticle formation. In other words, expression beyond this period is futile. Importantly, manipulation of Knk expression during this time causes wing bending suggesting an effect of Knk amounts on the physical properties of the wing cuticle. Manipulation of Knk expression also interferes with the structure and function of the cuticle surface. First, it was shown that the shape of surface nano-structures depends on the expression levels of knk. Second, it was found that cuticle impermeability is compromised in wings with reduced knk expression. In summary, despite the extended supply of Knk during metamorphosis, controlled amounts of Knk are important for correct wing cuticle differentiation and function in a concise period of time.
Abbasi, R. and Marcus, J. M. (2017). A new A-P compartment boundary and organizer in holometabolous insect wings. Sci Rep 7(1): 16337. PubMed ID: 29180689
Decades of research on the highly modified wings of Drosophila melanogaster has suggested that insect wings are divided into two Anterior-Posterior (A-P) compartments separated by an axis of symmetry. This axis of symmetry is created by a developmental organizer that establishes symmetrical patterns of gene expression that in turn pattern the A-P axis of the wing. Butterflies possess more typical insect wings and butterfly wing colour patterns provide many landmarks for studies of wing structure and development. Using eyespot colour pattern variation in Vanessa butterflies, this study shows an additional A-P axis of symmetry running between wing sectors 3 and 4. Boundaries of Drosophila mitotic clones suggest the existence of a previously undetected Far-Posterior (F-P) compartment boundary that coincides with this additional A-P axis. A similar compartment boundary is evident in butterfly mosaic gynandromorphs. It is suggested that this additional compartment boundary and its associated developmental organizer create an axis of wing colour pattern symmetry and a gene expression-based combinatorial code, permitting each insect wing compartment to acquire a unique identity and allowing for the individuation of butterfly eyespots.
Ma, M., Cao, X., Dai, J. and Pastor-Pareja, J. C. (2017). Basement membrane manipulation in Drosophila wing discs affects Dpp retention but not growth mechanoregulation. Dev Cell 42(1): 97-106.e104. PubMed ID: 28697337
Basement membranes (BMs) are extracellular matrix polymers basally underlying epithelia, where they regulate cell signaling and tissue mechanics. Constriction by the BM shapes Drosophila wing discs, a well-characterized model of tissue growth. Recently, the hypothesis that mechanical factors govern wing growth has received much attention, but it has not been definitively tested. This study manipulated BM composition to cause dramatic changes in tissue tension. Increased tissue compression when perlecan was knocked down did not affect adult wing size. BM elimination, decreasing compression, reduced wing size but did not visibly affect Hippo signaling, widely postulated to mediate growth mechanoregulation. BM elimination, in contrast, attenuated signaling by bone morphogenetic protein/transforming growth factor beta ligand Dpp, which was not efficiently retained within the tissue and escaped to the body cavity. The results challenge mechanoregulation of wing growth, while uncovering a function of BMs in preserving a growth-promoting tissue environment.

Monday, December 4th

Turissini, D. A. and Matute, D. R. (2017). Fine scale mapping of genomic introgressions within the Drosophila yakuba clade. PLoS Genet 13(9): e1006971. PubMed ID: 28873409
The process of speciation involves populations diverging over time until they are genetically and reproductively isolated. Hybridization between nascent species was long thought to directly oppose speciation. A natural place to look for individuals with admixed ancestry (indicative of introgression) is in regions where species co-occur. In west Africa, D. santomea and D. yakuba hybridize on the island of Sao Tome, while D. yakuba and D. teissieri hybridize on the nearby island of Bioko. This report quantifies the genomic extent of introgression between the three species of the Drosophila yakuba clade (D. yakuba, D. santomea), D. teissieri). The genomes of 86 individuals were sequenced from all three species. A new statistical framework was developed and applied, using a hidden Markov approach, to identify introgression. Introgression was found to have occurred between both species pairs but most introgressed segments are small (on the order of a few kilobases). After ruling out the retention of ancestral polymorphism as an explanation for these similar regions, this study found that the sizes of introgressed haplotypes indicate that genetic exchange is not recent (>1,000 generations ago). It was additionally shown that in both cases, introgression was rarer on X chromosomes than on autosomes which is consistent with sex chromosomes playing a large role in reproductive isolation. Even though the two species pairs have stable contemporary hybrid zones, providing the opportunity for ongoing gene flow, the results indicate that genetic exchange between these species is currently rare.
Siddiq, M. A., Loehlin, D. W., Montooth, K. L. and Thornton, J. W. (2017). Experimental test and refutation of a classic case of molecular adaptation in Drosophila melanogaster. Nat Ecol Evol 1(2): 25. PubMed ID: 28812605
Identifying the genetic basis for adaptive differences between species requires explicit tests of historical hypotheses concerning the effects of past changes in gene sequence on molecular function, organismal phenotype and fitness. This challenge was addressed by combining ancestral protein reconstruction with biochemical experiments and physiological analysis of transgenic animals that carry ancestral genes. A widely held hypothesis of molecular adaptation was tested in this study-that changes in the alcohol dehydrogenase protein (ADH) along the lineage leading to Drosophila melanogaster increased the catalytic activity of the enzyme and thereby contributed to the ethanol tolerance and adaptation of the species to its ethanol-rich ecological niche. These experiments strongly refute the predictions of the adaptive ADH hypothesis and caution against accepting intuitively appealing accounts of historical molecular adaptation that are based on correlative evidence. The experimental strategy employed can be used to decisively test other adaptive hypotheses and the claims they entail about past biological causality.
Schmidt, J. M., Battlay, P., Gledhill-Smith, R. S., Good, R. T., Lumb, C., Fournier-Level, A. and Robin, C. (2017). Insights into DDT Resistance from the Drosophila melanogaster Genetic Reference Panel. Genetics [Epub ahead of print]. PubMed ID: 28935691
Insecticide resistance is considered a classic model of microevolution, where a strong selective agent is applied to a large natural population, resulting in a change in frequency of alleles that confer resistance. While many insecticide resistance variants have been characterized at the gene level, they are typically single genes of large effect identified in highly resistant pest species. In contrast, multiple variants have been implicated in DDT resistance in Drosophila melanogaster, however only the Cyp6g1 locus has previously been shown to be relevant to field populations. This study used genome-wide association studies to identify DDT-associated polygenes and used selective sweep analyses to assess their adaptive significance. Two candidate DDT resistance loci were identified and verified. A largely uncharacterized gene, CG10737, has a function in muscles that ameliorates the effects of DDT, while a putative detoxifying P450, Cyp6w1, shows compelling evidence of positive selection.
Nourmohammad, A., Rambeau, J., Held, T., Kovacova, V., Berg, J. and Lassig, M. (2017). Adaptive evolution of gene expression in Drosophila. Cell Rep 20(6): 1385-1395. PubMed ID: 28793262
Gene expression levels are important quantitative traits that link genotypes to molecular functions and fitness. In Drosophila, population-genetic studies have revealed substantial adaptive evolution at the genomic level, but the evolutionary modes of gene expression remain controversial. This study presents evidence that adaptation dominates the evolution of gene expression levels in flies. 64% of the observed expression divergence across seven Drosophila species are adaptive changes driven by directional selection. The results are derived from time-resolved data of gene expression divergence across a family of related species, using a probabilistic inference method for gene-specific selection. Adaptive gene expression is stronger in specific functional classes, including regulation, sensory perception, sexual behavior, and morphology. Moreover, a large group of genes was identifed with sex-specific adaptation of expression, which predominantly occurs in males. This analysis opens an avenue to map system-wide selection on molecular quantitative traits independently of their genetic basis.

Friday, December 1st

Lynch, Z. R., Schlenke, T. A., Morran, L. T. and de Roode, J. C. (2017). Ethanol confers differential protection against generalist and specialist parasitoids of Drosophila melanogaster. PLoS One 12(7): e0180182. PubMed ID: 28700600
As parasites coevolve with their hosts, they can evolve counter-defenses that render host immune responses ineffective. These counter-defenses are more likely to evolve in specialist parasites than generalist parasites; the latter face variable selection pressures between the different hosts they infect. Natural populations of the fruit fly Drosophila melanogaster are commonly threatened by endoparasitoid wasps in the genus Leptopilina, including the specialist L. boulardi and the generalist L. heterotoma, and both wasp species can incapacitate the cellular immune response of D. melanogaster larvae. Given that ethanol tolerance is high in D. melanogaster and stronger in the specialist wasp than the generalist, this study tested whether fly larvae could use ethanol as an anti-parasite defense and whether its effectiveness would differ against the two wasp species. Fly larvae were found to benefit from eating ethanol-containing food during exposure to L. heterotoma; a two-fold decrease in parasitization intensity and a 24-fold increase in fly survival to adulthood were observed. Although host ethanol consumption did not affect L. boulardi parasitization rates or intensities, it led to a modest increase in fly survival. Thus, ethanol conferred stronger protection against the generalist wasp than the specialist. Test were made to see whether fly larvae can self-medicate by seeking ethanol-containing food after being attacked by wasps, but found no support for this hypothesis was found. Female flies were allowed to choose between control and ethanol-containing oviposition sites in the presence vs. absence of wasps and generally found significant preferences for ethanol regardless of wasp presence. Overall, these results suggest that D. melanogaster larvae obtain protection from certain parasitoid wasp species through their mothers' innate oviposition preferences for ethanol-containing food sources.
Noh, S., Everman, E. R., Berger, C. M. and Morgan, T. J. (2017). Seasonal variation in basal and plastic cold tolerance: Adaptation is influenced by both long- and short-term phenotypic plasticity. Ecol Evol 7(14): 5248-5257. PubMed ID: 28770063
Understanding how thermal selection affects phenotypic distributions across different time scales will allow prediction of the effect of climate change on the fitness of ectotherms. This study tested how seasonal temperature variation affects basal levels of cold tolerance and two types of phenotypic plasticity in Drosophila melanogaster. Developmental acclimation occurs as developmental stages of an organism are exposed to seasonal changes in temperature and its effect is irreversible, while reversible short-term acclimation occurs daily in response to diurnal changes in temperature. Wild flies were selected from a temperate population across seasons and two cold tolerance metrics (chill-coma recovery and cold stress survival) and their responses to developmental and short-term acclimation were measured. Chill-coma recovery responded to seasonal shifts in temperature, and phenotypic plasticity following both short-term and developmental acclimation improved cold tolerance. This improvement indicated that both types of plasticity are adaptive, and that plasticity can compensate for genetic variation in basal cold tolerance during warmer parts of the season when flies tend to be less cold tolerant. A significantly stronger trade-off was observed between basal cold tolerance and short-term acclimation during warmer months. For the longer-term developmental acclimation, a trade-off persisted regardless of season. A relationship between the two types of plasticity may provide additional insight into why some measures of thermal tolerance are more sensitive to seasonal variation than others.
Morimoto, J., Simpson, S. J. and Ponton, F. (2017). Direct and trans-generational effects of male and female gut microbiota in Drosophila melanogaster. Biol Lett 13(7). PubMed ID: 28724687
There is increasing evidence of the far-reaching effects of gut bacteria on physiological and behavioural traits, yet the fitness-related consequences of changes in the gut bacteria composition of sexually interacting individuals remain unknown. To address this question, the gut microbiota of fruit flies, Drosophila melanogaster, were manipulated by monoinfecting flies with either Acetobacter pomorum (AP) or Lactobacillus plantarum (LP). Re-inoculated individuals were paired in all treatment combinations. LP-infected males had longer mating duration and induced higher short-term offspring production in females compared with AP-infected males. Furthermore, females of either re-inoculation state mated with AP-infected males were more likely to have zero offspring after mating, suggesting a negative effect of AP on male fertility. Finally, the effects of male and female gut bacteria interacted to modulate their daughters', but not sons' body mass, revealing a new trans-generational effect of parental gut microbiota. In conclusion, this study shows direct and trans-generational effects of the gut microbiota on mating and reproduction.
Poudel, S., Kim, Y., Gwak, J. S., Jeong, S. and Lee, Y. (2017). Gustatory receptor 22e is essential for sensing chloroquine and strychnine in Drosophila melanogaster. Insect Biochem Mol Biol 88: 30-36. PubMed ID: 28751111
Chloroquine, an amino quinolone derivative commonly used as an anti-malarial drug, is known to impart an unpleasant taste. Little research has been done to study chloroquine taste in insects; therefore, this study examined both the deterrant properties and mechanisms underlying chloroquine perception in fruit flies. The antifeedant effect of chloroquine was identified by screening 21 gustatory receptor (Grs) mutants through behavioral feeding assays and electrophysiology experiments. Two molecular sensors, GR22e and GR33a, were found to act as chloroquine receptors, and chloroquine-mediated activation of GRNs was found to occur through S-type sensilla. At the same time, the chloroquine receptor was successfully recapitulated by expressing GR22e in ectopic gustatory receptor neurons. GR22e was found to form a part of the strychnine receptor. It is suggested that the Drosophila strychnine receptor might have a very complex structure since five different GRs are required for strychnine-induced action potentials.
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