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

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


Friday June 30th, 2017

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Mistry, R., Kounatidis, I. and Ligoxygakis, P. (2017). Interaction between familial transmission and a constitutively active immune system shapes gut microbiota in Drosophila melanogaster. Genetics 206(2):889-904. PubMed ID: 28413160
Resident gut bacteria are constantly influencing the immune system. Yet the role of the immune system in shaping microbiota composition during an organism's lifespan has remained unclear. This study used Drosophila as a genetically tractable system with a simple gut bacterial population structure and streamlined genetic backgrounds to address this issue. Depending on their genetic background, young flies had microbiota of different diversities that converged with age to the same Acetobacteraceae-dominated pattern in healthy flies. This pattern was accelerated in immune-compromised flies with higher bacterial load and gut cell death. Nevertheless, immune compromised flies resembled their genetic background, indicating that familial transmission was the main force regulating gut microbiota. In contrast, flies with a constitutively active immune system had microbiota readily distinguishable from their genetic background with the introduction and establishment of previously undetectable bacterial families. This indicated the influence of immunity over familial transmission. Moreover, hyper active immunity and increased enterocyte death resulted in the highest bacterial load observed starting from early adulthood. Cohousing experiments showed that the microenvironment also played an important role in the structure of the microbiota where flies with constitutive immunity defined the gut microbiota of their co-habitants. These data show that in Drosophila, constitutively active immunity shapes the structure and density of gut microbiota.
Kutch, I. C. and Fedorka, K. M. (2017). A test for Y-linked additive and epistatic effects on surviving bacterial infections in Drosophila melanogaster. J Evol Biol. PubMed ID: 28510293
Y and W chromosomes offer a theoretically powerful way for sexual dimorphism to evolve. Consistent with this possibility, Drosophila melanogaster Y-chromosomes can influence gene regulation throughout the genome; particularly immune-related genes. In order for Y-linked regulatory variation (YRV) to contribute to adaptive evolution it must be comprised of additive genetic variance, such that variable Ys induce consistent phenotypic effects within the local gene pool. The potential for Y-chromosomes to adaptively shape gram-negative and gram-positive bacterial defense was tested by introgressing Ys across multiple genetic haplotypes from the same population. No Y-linked additive effects on immune phenotypes were found, suggesting a restricted role for the Y to facilitate dimorphic evolution. A large magnitude Y was found by background interaction that induced rank order reversals of Y-effects across the backgrounds (i.e. sign epistasis). Thus, Y-chromosome effects appeared consistent within backgrounds, but highly variable among backgrounds. This large sign epistatic effect could constrain monomorphic selection in both sexes, considering that autosomal alleles under selection must spend half of their time in a male background where relative fitness values are altered. If the pattern described in this study is consistent for other traits or within other XY (or ZW) systems, then YRV may represent a universal constraint to autosomal trait evolution.
Kounatidis, I., Chtarbanova, S., Cao, Y., Hayne, M., Jayanth, D., Ganetzky, B. and Ligoxygakis, P. (2017). NF-κB immunity in the brain determines fly lifespan in healthy aging and age-related neurodegeneration. Cell Rep 19: 836-848. PubMed ID: 28445733
During aging, innate immunity progresses to a chronically active state. However, what distinguishes those that "age well" from those developing age-related neurological conditions is unclear. This study used Drosophila to explore the cost of immunity in the aging brain. Mutations in intracellular negative regulators of the IMD/NF-κB pathway were shown to predispose flies to toxic levels of antimicrobial peptides, resulting in early locomotor defects, extensive neurodegeneration, and reduced lifespan. These phenotypes are rescued when immunity is suppressed in glia. In healthy flies, suppressing immunity in glial cells results in increased adipokinetic hormonal signaling with high nutrient levels in later life and an extension of active lifespan. Thus, when levels of IMD/NF-κB deviate from normal, two mechanisms are at play: lower levels derepress an immune-endocrine axis, which mobilizes nutrients, leading to lifespan extension, whereas higher levels increase antimicrobial peptides, causing neurodegeneration. Immunity in the fly brain is therefore a key lifespan determinant.

Loch, G., Zinke, I., Mori, T., Carrera, P., Schroer, J., Takeyama, H. and Hoch, M. (2017). Antimicrobial peptides extend lifespan in Drosophila. PLoS One 12(5): e0176689. PubMed ID: 28520752
Antimicrobial peptides (AMPs) are important defense molecules of the innate immune system. High levels of AMPs are induced in response to infections to fight pathogens, whereas moderate levels induced by metabolic stress are thought to shape commensal microbial communities at barrier tissues. Single AMPs were expressed in adult flies either ubiquitously or in the gut by using the inducible GeneSwitch system to tightly regulate AMP expression. Activation of single AMPs, including Drosocin, were found to result in a significant extension of Drosophila lifespan. These animals showed reduced activity of immune pathways over lifetime, less intestinal regenerative processes, reduced stress response and a delayed loss of gut barrier integrity. Furthermore, intestinal Drosocin induction protected the animals against infections with the natural Drosophila pathogen Pseudomonas entomophila, whereas a germ-reduced environment prevented the lifespan extending effect of Drosocin. This study provides new insights into the crosstalk of innate immunity, intestinal homeostasis and ageing.
Paik, D., Monahan, A., Caffrey, D. R., Elling, R., Goldman, W. E. and Silverman, N. (2017). SLC46 family transporters facilitate cytosolic innate immune recognition of monomeric peptidoglycans. J Immunol 199(1):263-270. PubMed ID: 28539433
Tracheal cytotoxin (TCT), a monomer of DAP-type peptidoglycan from Bordetella pertussis, causes cytopathology in the respiratory epithelia of mammals and robustly triggers the Drosophila Imd pathway. PGRP-LE, a cytosolic innate immune sensor in Drosophila, directly recognizes TCT and triggers the Imd pathway, yet the mechanisms by which TCT accesses the cytosol are poorly understood. This study reports that CG8046, a Drosophila SLC46 family transporter, is a novel transporter facilitating cytosolic recognition of TCT, and plays a crucial role in protecting flies against systemic Escherichia coli infection. In addition, mammalian SLC46A2s promote TCT-triggered NOD1 activation in human epithelial cell lines, indicating that SLC46As is a conserved group of peptidoglycan transporter contributing to cytosolic immune recognition.
Tokusumi, Y., Tokusumi, T. and Schulz, R. A. (2017). The nociception genes painless and Piezo are required for the cellular immune response of Drosophila larvae to wasp parasitization. Biochem Biophys Res Commun 486(4):893-897. PubMed ID: 28342875
In vertebrates, interaction between the nervous system and immune system is important to protect a challenged host from stress inputs from external sources. This study demonstrates that sensory neurons are involved in the cellular immune response elicited by wasp infestation of Drosophila larvae. Multidendritic class IV neurons sense contacts from external stimuli and induce avoidance behaviors for host defense. The findings show that inactivation of these sensory neurons impairs the cellular response against wasp parasitization. It was also demonstrated that the nociception genes encoding the mechanosensory receptors Painless and Piezo, both expressed in class IV neurons, are essential for the normal cellular immune response to parasite challenge.

Thursday, June 29th

McClelland, L., Jasper, H. and Biteau, B. (2017). Tis11 mediated mRNA decay promotes the reacquisition of Drosophila intestinal stem cell quiescence. Dev Biol 426(1): 8-16. PubMed ID: 28445691
Adult stem cell proliferation rates are precisely regulated to maintain long-term tissue homeostasis. Defects in the mechanisms controlling stem cell proliferation result in impaired regeneration and hyperproliferative diseases. Many stem cell populations increase proliferation in response to tissue damage and reacquire basal proliferation rates after tissue repair is completed. Although proliferative signals have been extensively studied, much less is known about the molecular mechanisms that restore stem cell quiescence. This study shows that Tis11, an Adenine-uridine Rich Element (ARE) binding protein that promotes mRNA degradation, is required to re-establish basal proliferation rates of adult Drosophila intestinal stem cells (ISC) after a regenerative episode. Tis11 limits ISC proliferation specifically after proliferation has been stimulated in response to heat stress or infection, and Tis11 expression and activity are increased in ISCs during tissue repair. Based on stem cell transcriptome analysis and RNA immunoprecipitation, it is proposed that Tis11 activation represents an integral part of a negative feedback mechanism that limits the expression of key components of several signaling pathways that control ISC function and proliferation. The results identify Tis11 mediated mRNA decay as an evolutionarily conserved mechanism of re-establishing basal proliferation rates of stem cells in regenerating tissues.
Vicidomini, R., Petrizzo, A., di Giovanni, A., Cassese, L., Lombardi, A. A., Pragliola, C. and Furia, M. (2017). Drosophila dyskerin is required for somatic stem cell homeostasis. Sci Rep 7(1): 347. PubMed ID: 28337032
Drosophila represents an excellent model to dissect the roles played by the evolutionary conserved family of eukaryotic dyskerins. These multifunctional proteins are involved in the formation of H/ACA snoRNP and telomerase complexes, both involved in essential cellular tasks. Since fly telomere integrity is guaranteed by a different mechanism, the specific role played by dyskerin in somatic stem cell maintenance was investigated. Focus was placed on Drosophila midgut, a hierarchically organized and well characterized model for stemness analysis. Surprisingly, the ubiquitous loss of the protein uniquely affects the formation of the larval stem cell niches, without altering other midgut cell types. The number of adult midgut precursor stem cells is dramatically reduced, and this effect is not caused by premature differentiation and is cell-autonomous. Moreover, only a few dispersed precursors found in the depleted midguts could maintain stem identity and the ability to divide asymmetrically, and show cell-growth defects or undergo apoptosis. Loss is mainly specifically dependent on defective amplification. These studies establish a strict link between dyskerin and somatic stem cell maintenance in a telomerase-lacking organism, indicating that loss of stemness can be regarded as a conserved, telomerase-independent effect of dyskerin dysfunction.
Jin, Y., Patel, P. H., Kohlmaier, A., Pavlovic, B., Zhang, C. and Edgar, B. A. (2017). Intestinal stem cell pool regulation in Drosophila. Stem Cell Reports [Epub ahead of print]. PubMed ID: 28479306
Intestinal epithelial renewal is mediated by intestinal stem cells (ISCs) that exist in a state of neutral drift, wherein individual ISC lineages are regularly lost and born but ISC numbers remain constant. To test whether an active mechanism maintains stem cell pools in the Drosophila midgut, partial ISC depletion was performed. In contrast to the mouse intestine, Drosophila ISCs failed to repopulate the gut after partial depletion. Even when the midgut was challenged to regenerate by infection, ISCs retained normal proportions of asymmetric division and ISC pools did not increase. The loss of differentiated midgut enterocytes (ECs), however, slows when ISC division is suppressed and accelerates when ISC division increases. This plasticity in rates of EC turnover appears to facilitate epithelial homeostasis even after stem cell pools are compromised. This study identifies unique behaviors of Drosophila midgut cells that maintain epithelial homeostasis.
Ma, X., Zhu, X., Han, Y., Story, B., Do, T., Song, X., Wang, S., Zhang, Y., Blanchette, M., Gogol, M., Hall, K., Peak, A., Anoja, P. and Xie, T. (2017). Aubergine controls germline stem cell self-renewal and progeny differentiation via distinct mechanisms. Dev Cell 41(2): 157-169.e155. PubMed ID: 28441530
Piwi family protein Aubergine (Aub) maintains genome integrity in late germ cells of the Drosophila ovary through Piwi-associated RNA-mediated repression of transposon activities. Although it is highly expressed in germline stem cells (GSCs) and early progeny, it remains unclear whether it plays any roles in early GSC lineage development. This study reports that Aub promotes GSC self-renewal and GSC progeny differentiation. RNA-iCLIP results show that Aub binds the mRNAs encoding self-renewal and differentiation factors in cultured GSCs. Aub controls GSC self-renewal by preventing DNA-damage-induced Chk2 activation and by translationally controlling the expression of self-renewal factors. It promotes GSC progeny differentiation by translationally controlling the expression of differentiation factors, including Bam. Therefore, this study reveals a function of Aub in GSCs and their progeny, which promotes translation of self-renewal and differentiation factors by directly binding to its target mRNAs and interacting with translational initiation factors.

Wednesday, June 28th

Lopez-Arias, B., Turiegano, E., Monedero, I., Canal, I. and Torroja, L. (2017). Presynaptic Abeta40 prevents synapse addition in the adult Drosophila neuromuscular junction. PLoS One 12(5): e0177541. PubMed ID: 28520784
Complexity in the processing of the Amyloid Precursor Protein, which generates a mixture of βamyloid peptides, lies beneath the difficulty in understanding the etiology of Alzheimer's disease. Moreover, whether Aβ peptides have any physiological role in neurons is an unresolved question. By expressing single, defined Aβ peptides in Drosophila, specific effects can be discriminated in vivo. This study shows that in the adult neuromuscular junction (NMJ), presynaptic expression of Aβ40 hinders the synaptic addition that normally occurs in adults, yielding NMJs with an invariable number of active zones at all ages tested. A similar trend is observed for Aβ42 at young ages, but net synaptic loss occurs at older ages in NMJs expressing this amyloid species. In contrast, Aβ42arc produces net synaptic loss at all ages tested, although age-dependent synaptic variations are maintained. Inhibition of the PI3K synaptogenic pathway may mediate some of these effects, because western analyses show that Aβ peptides block activation of this pathway, and Aβ species-specific synaptotoxic effects persists in NMJs overgrown by over-expression of PI3K. Finally, individual Aβ effects are also observed when toxicity is examined by quantifying neurodegeneration and survival. These results suggest a physiological effect of Aβ40 in synaptic plasticity, and imply different toxic mechanisms for each peptide species.
Gehring, K. B., Heufelder, K., Depner, H., Kersting, I., Sigrist, S. J. and Eisenhardt, D. (2017). Age-associated increase of the active zone protein Bruchpilot within the honeybee mushroom body. PLoS One 12(4): e0175894. PubMed ID: 28437454
In honeybees, age-associated structural modifications can be observed in the mushroom bodies. Prominent examples are the synaptic complexes (microglomeruli, MG) in the mushroom body calyces, which were shown to alter their size and density with age. It is not known whether the amount of intracellular synaptic proteins in the MG is altered as well. The presynaptic protein Bruchpilot (BRP) is localized at active zones and is involved in regulating the probability of neurotransmitter release in the fruit fly, Drosophila melanogaster. This study explored the localization of the honeybee BRP (Apis mellifera BRP, AmBRP) in the bee brain and examined age-related changes in the AmBRP abundance in the central bee brain and in microglomeruli of the mushroom body calyces. Predominant AmBRP localization is reported near the membrane of presynaptic boutons within the mushroom body MG. The relative amount of AmBRP was increased in the central brain of two-week old bees whereas the amount of Synapsin, another presynaptic protein involved in the regulation of neurotransmitter release, shows an increase during the first two weeks followed by a decrease. In addition, an age-associated modulation was demonstrated of AmBRP located near the membrane of presynaptic boutons within MG located in mushroom body calyces where sensory input is conveyed to mushroom body intrinsic neurons. The observed age-associated AmBRP modulation might be related to maturation processes or to homeostatic mechanisms that might help to maintain synaptic functionality in old animals.
Inoshita, T., Arano, T., Hosaka, Y., Meng, H., Umezaki, Y., Kosugi, S., Morimoto, T., Koike, M., Chang, H. Y., Imai, Y. and Hattori, N. (2017). Vps35 in cooperation with LRRK2 regulates synaptic vesicle endocytosis through the endosomal pathway in Drosophila. Hum Mol Genet [Epub ahead of print]. PubMed ID: 28482024
Mutations of the retromer component Vps35 and endosomal kinase LRRK2 are linked to autosomal dominant forms of familial Parkinson's disease (PD). However, the physiological and pathological roles of Vps35 and LRRK2 in neuronal functions are poorly understood. This study demonstrated that the loss of Drosophila Vps35 (dVps35) affects synaptic vesicle. recycling, dopaminergic synaptic release and sleep behavior associated with dopaminergic activity, which is rescued by the expression of wild-type dVps35 but not the PD-associated mutant dVps35 D647N. Drosophila LRRK2 dLRRK together with Rab5 and Rab11 is also implicated in synaptic vesicle recycling, and the manipulation of these activities improves the Vps35 synaptic phenotypes. These findings indicate that defects of synaptic vesicle recycling in which two late-onset PD genes, Vps35 and LRRK2, are involved could be key aspects of PD etiology.
Kamalesh, K., Trivedi, D., Toscano, S., Sharma, S., Kolay, S. and Raghu, P. (2017). Phosphatidylinositol 5-phosphate 4-kinase regulates early endosomal dynamics during clathrin-mediated endocytosis. J Cell Sci [Epub ahead of print]. PubMed ID: 28507272
Endocytic turnover is essential to regulate the protein composition and function of the plasma membrane thus regulating the plasma membrane levels of many receptors. In Drosophila photoreceptors, photon absorption by the GPCR rhodopsin 1 (Rh1) triggers its endocytosis by clathrin-mediated endocytosis (CME). CME of Rh1 is regulated by phosphatidylinositol 5 phosphate 4-kinase (PIP4K). Flies lacking PIP4K show mislocalization of Rh1 on expanded endomembranes within the cell body. This mislocalization of Rh1 was dependent on the formation of an expanded Rab5 compartment. The Rh1 trafficking defect in dPIP4K-depleted cells could be suppressed by downregulating Rab5 function or by selectively reconstituting dPIP4K in the PI3P enriched early endosomal (EE) compartment of photoreceptors. It was also found that loss of PIP4K was associated with increased CME and an enlarged Rab5 compartment in cultured Drosophila cells. Collectively these findings define PIP4K as a novel regulator of early endosomal homeostasis during CME.

Tuesday, June 27th

Liao, P. C., Tandarich, L. C. and Hollenbeck, P. J. (2017). ROS regulation of axonal mitochondrial transport is mediated by Ca2+ and JNK in Drosophila. PLoS One 12(5): e0178105. PubMed ID: 28542430
Mitochondria perform critical functions including aerobic ATP production and calcium (Ca2+) homeostasis, but are also a major source of reactive oxygen species (ROS) production. To maintain cellular function and survival in neurons, mitochondria are transported along axons, and accumulate in regions with high demand for their functions. Oxidative stress and abnormal mitochondrial axonal transport are associated with neurodegenerative disorders. However, little is known about the connection between these two. Using the Drosophila third instar larval nervous system as the in vivo model, this study found that ROS inhibited mitochondrial axonal transport more specifically, primarily due to reduced flux and velocity, but did not affect transport of other organelles. To understand the mechanisms underlying these effects, Ca2+ levels were studied along with the JNK (c-Jun N-terminal Kinase) pathway, which have been shown to regulate mitochondrial transport and general fast axonal transport, respectively. Elevated ROS was found to increase Ca2+ levels, and experimental reduction of Ca2+ to physiological levels rescued ROS-induced defects in mitochondrial transport in primary neuron cell cultures. In addition, in vivo activation of the JNK pathway reduced mitochondrial flux and velocities, while JNK knockdown partially rescued ROS-induced defects in the anterograde direction. It is concluded that ROS have the capacity to regulate mitochondrial traffic, and that Ca2+ and JNK signaling play roles in mediating these effects. In addition to transport defects, ROS produces imbalances in mitochondrial fission-fusion and metabolic state, indicating that mitochondrial transport, fission-fusion steady state, and metabolic state are closely interrelated in the response to ROS.
Kuhn, L., Majzoub, K., Einhorn, E., Chicher, J., Pompon, J., Imler, J. L., Hammann, P. and Meignin, C. (2017). Definition of a RACK1 interaction network in Drosophila melanogaster Using SWATH-MS. G3 (Bethesda)[Epub ahead of print]. PubMed ID: 28522639
Receptor for Activated C kinase 1 (RACK1) is a scaffold protein that has been found in association with several signaling complexes, and with the 40S subunit of the ribosome. Using the model organism Drosophila melanogaster, it has been shown that RACK1 is required at the ribosome for IRES-mediated translation of viruses. This study reports a proteomic characterization of the interactome of RACK1 in Drosophila S2 cells. Label-Free quantitation was carried out using both Data-Dependent and Data-Independent Acquisition and a significant advantage was observed for the Sequential Window Acquisition of all THeoretical fragment-ion spectra (SWATH) method both in terms of identification of interactants and quantification of low abundance proteins. These data represent the first SWATH spectral library available for Drosophila and will be a useful resource for the community. A total of 52 interacting proteins were identified, including several molecules involved in translation such as structural components of the ribosome, factors regulating translation initiation or elongation and RNA binding proteins. Among these 52 proteins, 15 were identified as partners by the SWATH strategy only. Interestingly, these 15 proteins are significantly enriched for the functions translation and nucleic acid binding. This enrichment reflects the engagement of RACK1 at the ribosome and highlights the added value of SWATH analysis. A functional screen did not reveal any protein sharing the interesting properties of RACK1, which is required for IRES-dependent translation and not essential for cell viability. Intriguingly however, 10 of the RACK1 partners identified restrict replication of Cricket paralysis virus, an IRES-containing virus.
Goyal, Y., Levario, T. J., Mattingly, H. H., Holmes, S., Shvartsman, S. Y. and Lu, H. (2017). Parallel imaging of Drosophila embryos for quantitative analysis of genetic perturbations of the Ras pathway. Dis Model Mech [Epub ahead of print]. PubMed ID: 28495673
The Ras pathway patterns the poles of the Drosophila embryo by downregulating the levels and activity of a DNA-binding transcriptional repressor Capicua (Cic). This study demonstrates that the spatiotemporal pattern of Cic during this signaling event can be harnessed for functional studies of the Ras-pathway mutations from human diseases. The approach relies on a new microfluidic device that enables parallel imaging of Cic dynamics in dozens of live embryos. Although the pattern of Cic in early embryos is complex, it can be accurately approximated by a product of one spatial profile and one time-dependent amplitude. Analysis of these functions of space and time alone reveals the differential effects of mutations within the Ras pathway. Given the highly-conserved nature of Ras-dependent control of Cic, this approach opens a new way for functional analysis of multiple sequence variants from developmental abnormalities and cancers.
Jossin, Y., Lee, M., Klezovitch, O., Kon, E., Cossard, A., Lien, W. H., Fernandez, T. E., Cooper, J. A. and Vasioukhin, V. (2017). Llgl1 connects cell polarity with cell-cell adhesion in embryonic neural stem cells. Dev Cell [Epub ahead of print]. PubMed ID: 28552558
Evolutionary Homolog Study
Malformations of the cerebral cortex (MCCs) are devastating developmental disorders. This study reports that mice with embryonic neural stem-cell-specific deletion of Llgl1 Nestin-Cre/Llgl1fl/fl), a mammalian ortholog of the Drosophila cell polarity gene lgl, exhibit MCCs resembling severe periventricular heterotopia (PH). Immunohistochemical analyses and live cortical imaging of PH formation revealed that disruption of apical junctional complexes (AJCs) was responsible for PH in Nestin-Cre/Llgl1fl/fl brains. While it is well known that cell polarity proteins govern the formation of AJCs, the exact mechanisms remain unclear. This study shows that LLGL1 directly binds to and promotes internalization of N-cadherin (see Drosophila Cadherin-N), and N-cadherin/LLGL1 interaction is inhibited by atypical protein kinase C-mediated phosphorylation of LLGL1 (see Drosophila aPKC), restricting the accumulation of AJCs to the basolateral-apical boundary. Disruption of the N-cadherin-LLGL1 interaction during cortical development in vivo is sufficient for PH. These findings reveal a mechanism responsible for the physical and functional connection between cell polarity and cell-cell adhesion machineries in mammalian cells.

Monday, June 26th

Liang, X., Holy, T. E. and Taghert, P. H. (2017). A series of suppressive signals within the Drosophila circadian neural circuit generates sequential daily outputs. Neuron [Epub ahead of print]. PubMed ID: 28552314
The Drosophila circadian neural circuit was studied using whole-brain imaging in vivo. Five major groups of pacemaker neurons display synchronized molecular clocks, yet each exhibits a distinct phase of daily Ca2+ activation. Light and neuropeptide pigment dispersing factor (PDF) from morning cells (s-LNv) together delay the phase of the evening (LNd) group by approximately 12 hr; PDF alone delays the phase of the DN3 group by approximately 17 hr. Neuropeptide sNPF, released from s-LNv and LNd pacemakers, produces Ca2+ activation in the DN1 group late in the night. The circuit also features negative feedback by PDF to truncate the s-LNv Ca2+ wave and terminate PDF release. Both PDF and sNPF suppress basal Ca2+ levels in target pacemakers with long durations by cell-autonomous actions. Thus, light and neuropeptides act dynamically at distinct hubs of the circuit to produce multiple suppressive events that create the proper tempo and sequence of circadian pacemaker neuronal activities.
Lee, S., Kim, Y. J. and Jones, W. D. (2017). Central peptidergic modulation of peripheral olfactory responses. BMC Biol 15(1): 35. PubMed ID: 28476120
Drosophila neuropeptide F (NPF) modulates the responses of a specific population of antennal olfactory sensory neurons (OSNs) to food-derived odors. Knock-down of NPF in NPF neurons specifically reduces the responses of the ab3A neurons to ethyl butyrate, a volatile ester found in apples and other fruits. Knock-down of the NPF receptor (NPFR) in the ab3A neuron reduces their responses and disrupts the ability of the flies to locate food. A sexual dimorphism was identified in ab3A responsiveness: ab3A neurons in females immediately post-eclosion are less responsive to ethyl butyrate than those of both age-matched males and older females. Not only does this change correlate with brain NPF levels, but also NPFR mutants show no such sexual dimorphism. Finally, by way of mechanism, mutation of NPFR seems to cause intracellular clustering of OR22a, the odorant receptor expressed in the ab3A neurons. This modulation of the peripheral odorant responsiveness of the ab3A neurons by NPF is distinct from the modulation of presynaptic gain in the ab3A neurons previously observed with the similarly named but distinct neuropeptide sNPF. Rather than affecting the strength of the output at the level of the first synapse in the antennal lobe, NPF-NPFR signaling may affect the process of odorant detection itself by causing intracellular OR clustering.
Konig, C., Antwi-Adjei, E., Ganesan, M., Kilonzo, K., Viswanathan, V., Durairaja, A., Voigt, A. and Yarali, A. (2017). Aversive olfactory associative memory loses odor specificity over time. J Exp Biol 220(Pt 9): 1548-1553. PubMed ID: 28468811
Avoiding associatively learned predictors of danger is crucial for survival. Aversive memories can, however, become counter-adaptive when they are overly generalized to harmless cues and contexts. In a fruit fly odor-electric shock associative memory paradigm, this study found that learned avoidance lost its specificity for the trained odor and became general to novel odors within a day of training. The possible neural circuit mechanisms of this effect are discussed, and the parallelism is highlighted to over-generalization of learned fear behavior after an incubation period in rodents and humans, with due relevance for post-traumatic stress disorder.
Kim, S. S., Rouault, H., Druckmann, S. and Jayaraman, V. (2017). Ring attractor dynamics in the Drosophila central brain. Science 356(6340): 849-853. PubMed ID: 28473639
Ring attractors are a class of recurrent networks hypothesized to underlie the representation of heading direction. Such network structures, schematized as a ring of neurons whose connectivity depends on their heading preferences, can sustain a bump-like activity pattern whose location can be updated by continuous shifts along either turn direction. A population of fly neurons in the ellipsoid body has been shown to represent the animal's heading via bump-like activity dynamics. This study combined two-photon calcium imaging in head-fixed flying flies with optogenetics to overwrite the existing population representation with an artificial one, which was then maintained by the circuit with naturalistic dynamics. A network with local excitation and global inhibition enforces this unique and persistent heading representation. Ring attractor networks have long been invoked in theoretical work; this study provides physiological evidence of their existence and functional architecture.

Sunday, June 25th

Levin, T. C. and Malik, H. S. (2017). Rapidly evolving Toll-3/4 genes encode male-specific Toll-like receptors in Drosophila. Mol Biol Evol. PubMed ID: 28541576
Animal Toll-like receptors (TLRs) have evolved through a pattern of duplication and divergence. Whereas mammalian TLRs directly recognize microbial ligands, Drosophila Tolls bind endogenous ligands downstream of both developmental and immune signaling cascades. This study found that most Toll genes in Drosophila evolve slowly with little gene turnover (gains/losses), consistent with their important roles in development and indirect roles in microbial recognition. In contrast, the Toll-3/4 genes were found to have experienced an unusually rapid rate of gene gains and losses, resulting in lineage-specific Toll-3/4s and vastly different gene repertoires among Drosophila species, from zero copies (e.g., D. mojavensis) to nineteen copies (e.g., D. willistoni). In D. willistoni, strong evidence was found for positive selection in Toll-3/4 genes, localized specifically to an extracellular region predicted to overlap with the binding site of Spatzle, the only known ligand of insect Tolls. However, because Spatzle genes are not experiencing similar selective pressures, it was hypothesize that Toll-3/4s may be rapidly evolving because they bind to a different ligand, akin to TLRs outside of insects. Unlike other Toll genes in D. melanogaster, Toll-3 and Toll-4 have apparently escaped from essential developmental roles. It is proposed that the Toll-3/4 genes represent an exceptionally rapidly evolving lineage of Drosophila Toll genes, which play an unusual, as-yet-undiscovered role in the male germline.
Duxbury, E. M. L., Rostant, W. G. and Chapman, T. (2017). Manipulation of feeding regime alters sexual dimorphism for lifespan and reduces sexual conflict in Drosophila melanogaster. Proc Biol Sci 284(1854). PubMed ID: 28469030
Sexual dimorphism for lifespan (SDL) is widespread, but poorly understood. A leading hypothesis, which was tested in this study, is that strong SDL can reduce sexual conflict by allowing each sex to maximize its sex-specific fitness. Replicated experimental evolution lines of the fruit fly, Drosophila melanogaster, were used that had been maintained for over 360 generations on either unpredictable 'Random' or predictable 'Regular' feeding regimes. This evolutionary manipulation of feeding regime led to robust, enhanced SDL in Random over control, Regular lines. Enhanced SDL was associated with a significant increase in the fitness of focal males, tested with wild-type (WT) females. This was due to sex-specific changes to male life history, manifested as increased early reproductive output and reduced survival. In contrast, focal female fitness, tested with WT males, did not differ across regimes. Hence increased SDL was associated with a reduction in sexual conflict, which increased male fitness and maintained fitness in females. Differences in SDL were not associated with developmental time or developmental survival. Overall, the results showed that the expression of enhanced SDL, resulting from experimental evolution of feeding regimes, was associated with male-specific changes in life history, leading to increased fitness and reduced sexual conflict.
Kang, L., Garner, H. R., Price, D. K. and Michalak, P. (2017). Kang, L., Garner, H. R., Price, D. K. and Michalak, P. (2017). A test for gene flow among sympatric and allopatric Hawaiian picture-winged Drosophila. J Mol Evol [Epub ahead of print]. PubMed ID: 28492967
The Hawaiian Drosophila are one of the most species-rich endemic groups in Hawaii and a spectacular example of adaptive radiation. Drosophila silvestris and D. heteroneura are two closely related picture-winged Drosophila species that occur sympatrically on Hawaii Island and are known to hybridize in nature, yet exhibit highly divergent behavioral and morphological traits driven largely through sexual selection. Their closest-related allopatric species, D. planitibia from Maui, exhibits hybrid male sterility and reduced behavioral reproductive isolation when crossed experimentally with D. silvestris or D. heteroneura. A modified four-taxon test for gene flow was applied to recently obtained genomes of the three Hawaiian Drosophila species. The analysis indicates recent gene flow in sympatry, but also, although less extensive, between allopatric species. This study underscores the prevalence of gene flow, even in taxonomic groups considered classic examples of allopatric speciation on islands. The potential confounding effects of gene flow in phylogenetic and population genetics inference are discussed, as well as the implications for conservation.
Martinez, J., Tolosana, I., Ok, S., Smith, S., Snoeck, K., Day, J. P. and Jiggins, F. (2017). Symbiont strain is the main determinant of variation in Wolbachia-mediated protection against viruses across Drosophila species. Mol Ecol [Epub ahead of print]. PubMed ID: 28464440
Wolbachia is a common heritable bacterial symbiont in insects. Its evolutionary success lies in the diverse phenotypic effects it has on its hosts coupled to its propensity to move between host species over evolutionary timescales. In a survey of natural host-symbiont associations in a range of Drosophila species, this study found that 10 of 16 Wolbachia strains protected their hosts against viral infection. By moving Wolbachia strains between host species, the symbiont genome was found to have a much greater influence on the level of antiviral protection than the host genome. The reason for this was that the level of protection depended on the density of the symbiont in host tissues, and Wolbachia rather than the host controlled density. The finding that virus resistance and symbiont density are largely under the control of symbiont genes in this system has important implications both for the evolution of these traits and for public health programs using Wolbachia to prevent mosquitoes from transmitting disease.

Saturday, June 24th

Iglesias, P. P. and Hasson, E. (2017). The role of courtship song in female mate choice in South American Cactophilic Drosophila. PLoS One 12(5): e0176119. PubMed ID: 28467464
Courtship songs have undergone a spectacular diversification in the Drosophila buzzatii cluster. Accordingly, it has been suggested that sexual selection has played a significant role in promoting rapid diversification, reproductive isolation and speciation. However, there is no direct evidence (i.e., song playback experiments with wingless males) supporting this tenet. Moreover, several studies have showed that the courtship song in the genus Drosophila is not always used in female mate choice decisions, nor plays the same role when it is taken into account. In this vein, this study used an approach that combines manipulative and playback experiments to explore the importance and the role of courtship song in female mate choice in four species of the D. buzzatii cluster and one species of the closely related D. martensis cluster for outgroup comparison. The importance of courtship song in sexual isolation in sympatry was also investigated between the only semi-cosmopolitan species, D. buzzatii, and the other species of the D. buzzatii cluster. This study revealed that the courtship song is used by females of the D. buzzatii cluster as a criterion for male acceptance or influences the speed with which males are chosen. In contrast, it was shown that this characteristic is not shared by D. venezolana, the representative species of the D. martensis cluster. It was also found that the studied species of the D. buzzatii cluster differ in the role that conspecific and heterospecific songs have in female mate choice and in sexual isolation. These findings support the hypothesis that divergence in female preferences for courtship songs has played a significant role in promoting rapid diversification and reproductive isolation in the D. buzzatii cluster. However, evidence from D. venezolana suggests that the use of the courtship song in female mate choice is not a conserved feature in the D. buzzatii complex.
Humberg, T. H. and Sprecher, S. G. (2017). Age- and wavelength-dependency of Drosophila larval phototaxis and behavioral responses to natural lighting conditions. Front Behav Neurosci 11: 66. PubMed ID: 28473759
Animals use various environmental cues as key determinant for their behavioral decisions. Visual systems are hereby responsible to translate light-dependent stimuli into neuronal encoded information. Even though the larval eyes of the fruit fly Drosophila melanogaster are comparably simple, they comprise two types of photoreceptor neurons (PRs), defined by different Rhodopsin genes expressed. Recent findings support that for light avoidance Rhodopsin5 (Rh5) expressing photoreceptors are crucial, while Rhodopsin6 (Rh6) expressing photoreceptors are dispensable under laboratory conditions. However, it remains debated how animals change light preference during larval life. This study shows that larval negative phototaxis is age-independent as it persists in larvae from foraging to wandering developmental stages. Moreover, whether spectrally different Rhodopsins are employed for the detection of different wavelength of light remains unexplored. This study found that negative phototaxis can be elicit by light with wavelengths ranging from ultraviolet (UV) to green. This behavior is uniquely mediated by Rh5 expressing photoreceptors, and therefore suggest that this photoreceptor-type is able to perceive UV up to green light. In contrast to laboratory tests, field experiments revealed that Drosophila larvae uses both types of photoreceptors under natural lighting conditions. The results demonstrate that Drosophila larval eyes mediate avoidance of light stimuli with a wide, ecological relevant range of quantity (intensities) and quality (wavelengths). Thus, the two photoreceptor-types appear more likely to play a role in different aspects of phototaxis under natural lighting conditions, rather than color discrimination.
Kim, H., Jeong, Y. T., Choi, M. S., Choi, J., Moon, S. J. and Kwon, J. Y. (2017). Involvement of a Gr2a-expressing Drosophila pharyngeal gustatory receptor neuron in regulation of aversion to high-salt foods. Mol Cells [Epub ahead of print]. PubMed ID: 28535667
Regulation of feeding is essential for animal survival. The pharyngeal sense organs can act as a second checkpoint of food quality, due to their position between external taste organs such as the labellum which initially assess food quality, and the digestive tract. Growing evidence provides support that the pharyngeal sensory neurons regulate feeding, but much is still unknown. This study found that a pair of gustatory receptor neurons in the LSO, a Drosophila adult pharyngeal organ which expresses four gustatory receptors, is involved in feeding inhibition in response to high concentrations of sodium ions. RNAi experiments and mutant analysis showed that the gustatory receptor Gr2a is necessary for this process. This feeding preference determined by whether a food source is perceived as appetizing or not is influenced by nutritional conditions, such that when the animal is hungry, the need for energy dominates over how appealing the food source is. These results provide experimental evidence that factors involved in feeding function in a context-dependent manner.
Higuchi, T., Kohatsu, S. and Yamamoto, D. (2017). Quantitative analysis of visually induced courtship elements in Drosophila subobscura. J Neurogenet 31(1-2): 49-57. PubMed ID: 28552034
Fly Motion-detector with an Actuator-Coupled Stimulator (FlyMacs), in which the stimulation of a fly with a moving visual target and recording of induced behaviors are automated under computer control, was employed for the identification of motion features that trigger specific courtship elements in Drosophila subobscura. A female abdomen attached to the actuator, when moved in an appropriate pattern, evokes in the test male tapping-like foreleg motions, midleg swing and proboscis extension, which are considered to be elementary actions in male courtship behavior. Tapping is primarily induced when the target is moving, whereas midleg swing and proboscis extension are most frequently observed after the target stops moving. In contrast to midleg swing, which tends to occur immediately after target cessation (approximately 3000 ms), the incidence of proboscis extension gradually increases with time after target cessation, reaching a plateau at 3000 ms. The results suggest that tapping, midleg swing and proboscis extension are each induced by different movement features of the visual target. These findings do not support the view that a single key stimulus induces the entire courtship ritual. Rather, courtship behaviors in D. subobscura are correlated with movement and position of the target, which suggests that D. subobscura uses sensory information to pattern its courtship.

Friday, June 23rd

Ghezzi, A., Li, X., Lew, L. K., Wijesekera, T. P. and Atkinson, N. S. (2017). Alcohol-induced neuroadaptation is orchestrated by the histone acetyltransferase CBP. Front Mol Neurosci 10: 103 [Epub ahead of print]. PubMed ID: 28442993
Homeostatic neural adaptations to alcohol underlie the production of alcohol tolerance and the associated symptoms of withdrawal. These adaptations have been shown to persist for relatively long periods of time and are believed to be of central importance in promoting the addictive state. In Drosophila, a single exposure to alcohol results in long-lasting alcohol tolerance and symptoms of withdrawal following alcohol clearance. These persistent adaptations involve mechanisms such as long-lasting changes in gene expression and perhaps epigenetic restructuring of chromosomal regions. Histone modifications have emerged as important modulators of gene expression and are thought to orchestrate and maintain the expression of multi-gene networks. Previously genes that contribute to tolerance were identified as those that show alcohol-induced changes in histone H4 acetylation following a single alcohol exposure. However, the molecular mediator of the acetylation process that orchestrates their expression remains unknown. This study shows that the Drosophila ortholog of mammalian CBP, nejire, is the histone acetyltransferase involved in regulatory changes producing tolerance-alcohol induces nejire expression, nejire mutations suppress tolerance, and transgenic nejire induction mimics tolerance in alcohol-naive animals. Moreover, a loss-of-function mutation in the alcohol tolerance gene slo epistatically suppresses the effects of CBP induction on alcohol resistance, linking nejire to a well-established alcohol tolerance gene network. It is proposed that CBP is a central regulator of the network of genes underlying an alcohol adaptation.
Kellermann, V., van Heerwaarden, B. and Sgro, C. M. (2017). How important is thermal history? Evidence for lasting effects of developmental temperature on upper thermal limits in Drosophila melanogaster. Proc Biol Sci 284(1855) [Epub ahead of print]. PubMed ID: 28539515
A common practice in thermal biology is to take individuals directly from the field and estimate a range of thermal traits. These estimates are then used in studies aiming to understand broad scale distributional patterns, understanding and predicting the evolution of phenotypic plasticity, and generating predictions for climate change risk. However, the use of field-caught individuals in such studies ignores the fact that many traits are phenotypically plastic and will be influenced by the thermal history of the focal individuals. This study aims to determine the extent to which estimates of upper thermal limits (CTmax), a frequently used measure for climate change risk, are sensitive to developmental and adult acclimation temperatures and whether these two forms of plasticity are reversible. Examining a temperate and tropical population of Drosophila melanogaster this study shows that developmental acclimation has a larger and more lasting effect on CTmax than adult acclimation. Evidence was found for an interaction between developmental and adult acclimation, particularly when flies are acclimated for a longer period, and these effects can be population specific. These results suggest that thermal history can have lasting effects on estimates of CTmax. In addition, evidence is provided that developmental and/or adult acclimation are unlikely to contribute to substantial shifts in CTmax and that acclimation capacity may be constrained at higher temperatures.
Colinet, H., Pineau, C. and Com, E. (2017). Large scale phosphoprotein profiling to explore Drosophila cold acclimation regulatory mechanisms. Sci Rep 7(1): 1713. PubMed ID: 28490779
The regulatory mechanisms involved in the acquisition of thermal tolerance are unknown in insects. Reversible phosphorylation is a widespread post-translational modification that can rapidly alter proteins function(s). A large-scale comparative screening was conducted of phosphorylation networks in adult Drosophila flies that were cold-acclimated versus control. Using a modified SIMAC method followed by a multiple MS analysis strategy, a large collection of phosphopeptides (about 1600) and phosphoproteins (about 500) was identified in both groups, with good enrichment efficacy (80%). The saturation curves from the four biological replicates revealed that the phosphoproteome was rather well covered under the experimental conditions. Acclimation evoked a strong phosphoproteomic signal characterized by large sets of unique and differential phosphoproteins. These were involved in several major GO superclusters of which cytoskeleton organization, positive regulation of transport, cell cycle, and RNA processing were particularly enriched. Data suggest that phosphoproteomic changes in response to acclimation were mainly localized within cytoskeletal network, and particularly within microtubule associated complexes. This study opens up novel research avenues for exploring the complex regulatory networks that lead to acquired thermal tolerance.
Kim, G., Huang, J. H., McMullen, J. G., Newell, P. D. and Douglas, A. E. (2017). Physiological responses of insects to microbial fermentation products: insights from the interactions between Drosophila and acetic acid. J Insect Physiol [Epub ahead of print]. PubMed ID: 28522417
Acetic acid is a fermentation product of many microorganisms, including some that inhabit the food and guts of Drosophila. This study investigated the effect of dietary acetic acid on oviposition and larval performance of Drosophila. At all concentrations tested (0.34-3.4%), acetic acid promoted egg deposition by mated females in no-choice assays; and females preferred to oviposit on diet with acetic acid relative to acetic acid-free diet. However, acetic acid depressed larval performance, particularly extending the development time of both larvae colonized with the bacterium Acetobacter pomorum and axenic (microbe-free) larvae. The larvae may incur an energetic cost associated with dissipating the high acid load on acetic acid-supplemented diets. This effect was compounded by suppressed population growth of A. pomorum on the 3.4% acetic acid diet, such that the gnotobiotic Drosophila on this diet displayed traits characteristic of axenic Drosophila, specifically reduced developmental rate and elevated lipid content. It is concluded that acetic acid is deleterious to larval Drosophila, and hypothesized that acetic acid may function as a reliable cue for females to oviposit in substrates bearing microbial communities that promote larval nutrition.

Thursday, June 22nd

Kashima, R., Redmond, P. L., Ghatpande, P., Roy, S., Kornberg, T. B., Hanke, T., Knapp, S., Lagna, G. and Hata, A. (2017). Hyperactive locomotion in a Drosophila model is a functional readout for the synaptic abnormalities underlying fragile X syndrome. Sci Signal 10(477). PubMed ID: 28465421
Fragile X syndrome (FXS) is the most common cause of heritable intellectual disability and autism and affects ~1 in 4000 males and 1 in 8000 females. The discovery of effective treatments for FXS has been hampered by the lack of effective animal models and phenotypic readouts for drug screening. FXS ensues from the epigenetic silencing or loss-of-function mutation of the fragile X mental retardation 1 (FMR1) gene, which encodes an RNA binding protein that associates with and represses the translation of target mRNAs. Previous studies found that the activation of LIM kinase 1 (LIMK1) downstream of augmented synthesis of bone morphogenetic protein (BMP) type 2 receptor (BMPR2) promotes aberrant synaptic development in mouse and Drosophila models of FXS and that these molecular and cellular markers were correlated in patients with FXS. This study reports that larval locomotion is augmented in a Drosophila FXS model. Genetic or pharmacological intervention on the BMPR2-LIMK pathway ameliorated the synaptic abnormality and locomotion phenotypes of FXS larvae, as well as hyperactivity in an FXS mouse model. This study demonstrates that (1) the BMPR2-LIMK pathway is a promising therapeutic target for FXS and (2) the locomotion phenotype of FXS larvae is a quantitative functional readout for the neuromorphological phenotype associated with FXS and is amenable to the screening novel FXS therapeutics.
Kang, J., Shin, S., Perrimon, N. and Shen, J. (2017). An evolutionarily conserved role of presenilin in neuronal protection in the aging Drosophila brain. Genetics [Epub ahead of print]. PubMed ID: 28495961
Mutations in the Presenilin genes are the major genetic cause of Alzheimer's disease. Presenilin and Nicastrin are essential components of γ-secretase, a multi-subunit protease that cleaves Type I transmembrane proteins. The roles of Drosophila Presenilin (Psn) and Nicastrin (Nct) in the adult fly brain are unknown. To knockdown (KD) Psn or Nct selectively in neurons of the adult brain, multiple shRNA lines were generated. Using a ubiquitous driver, these shRNA lines resulted in 80-90% reduction of mRNA and pupal lethality, a phenotype that is shared with Psn and Nct mutants carrying nonsense mutations. Furthermore, expression of these shRNAs in the wing disc caused notching wing phenotypes, which are also shared with Psn and Nct mutants. Similar to Nct, neuron-specific Psn KD using two independent shRNA lines led to early mortality and rough eye phenotypes, which were rescued by a fly Psn transgene. Interestingly, conditional KD (cKD) of Psn or Nct in adult neurons using the elav-Gal4 and tubulin-Gal80ts system caused shortened lifespan, climbing defects, increases in apoptosis and age-dependent neurodegeneration. Together, these findings demonstrate that similar to their mammalian counterparts, Drosophila Psn and Nct are required for neuronal survival during aging and normal lifespan, highlighting an evolutionarily conserved role of Presenilin in neuronal protection in the aging brain.
Chanu, S. I. and Sarkar, S. (2017). Targeted downregulation of dMyc restricts neurofibrillary tangles mediated pathogenesis of human neuronal tauopathies in Drosophila. Biochim Biophys Acta [Epub ahead of print]. PubMed ID: 28529046
Formation of Neurofibrillary Tangles (NFTs) in neuronal tissues has been implicated as the hallmark of disease pathogenesis and tau mediated toxicity in human and mammalian models. However, previous studies had failed to correlate NFT formation with pathogenesis of human neuronal tauopathies in Drosophila disease models. Though, a recent report suggests formation of tau mediated NFTs like structures confined to dopaminergic neurons in Drosophila adult brain, by utilizing various approaches, this study demonstrated distinct and recurrent formation of NFTs in Drosophila neuronal tissues upon expression of wild type or mutant isoforms of human tau, and this appears as the key mediator of the pathogenesis of human neuronal tauopathy in Drosophila. Further, it was shown that tissue specific downregulation of dMyc (Drosophila homolog of human c-myc proto-oncogene) alleviates h-tau mediated cellular and functional deficits by restricting the formation of NFTs in neuronal tissues. Therefore, these findings provide very critical and novel insights about pathogenesis of human neuronal tauopathies in Drosophila disease models.
Frenkel-Pinter, M., Stempler, S., Tal-Mazaki, S., Losev, Y., Singh-Anand, A., Escobar-Alvarez, D., Lezmy, J., Gazit, E., Ruppin, E. and Segal, D. (2017). Altered protein glycosylation predicts Alzheimer's disease and modulates its pathology in disease model Drosophila. Neurobiol Aging. PubMed ID: 28552182
The pathological hallmarks of Alzheimer's disease (AD) are pathogenic oligomers and fibrils of misfolded amyloidogenic proteins (e.g., beta-amyloid and hyper-phosphorylated tau in AD), which cause progressive loss of neurons in the brain and nervous system. In an analysis of available expression data sets this study indicates that many glycosylation-related genes are differentially expressed in brains of AD patients compared with healthy controls. The robust differences found enabled prediction of the occurrence of AD with remarkable accuracy in a test cohort and identification of a set of key genes whose expression determines this classification. Then the effect of reducing expression of homologs of 6 of these genes in was studied in transgenic Drosophila overexpressing human tau, a well-established invertebrate AD model. These experiments have led to the identification of glycosylation genes that may augment or ameliorate tauopathy phenotypes. These results indicate that OstDelta, l(2)not and beta4GalT7 are tauopathy suppressors, whereas pgnat5 and CG33303 are enhancers, of tauopathy. These results suggest that specific alterations in protein glycosylation may play a causal role in AD etiology.

Wednesday, June 21st

Piskadlo, E., Tavares, A. and Oliveira, R. A. (2017). Metaphase chromosome structure is dynamically maintained by Condensin I-directed DNA (de)catenation. Elife 6. PubMed ID: 28477406
Mitotic chromosome assembly remains a big mystery in biology. Condensin complexes are pivotal for chromosome architecture yet how they shape mitotic chromatin remains unknown. Using acute inactivation approaches and live-cell imaging in Drosophila embryos, this study dissects the role of condensin I in the maintenance of mitotic chromosome structure with unprecedented temporal resolution. Removal of condensin I from pre-established chromosomes results in rapid disassembly of centromeric regions while most chromatin mass undergoes hyper-compaction. This is accompanied by drastic changes in the degree of sister chromatid intertwines. While wild-type metaphase chromosomes display residual levels of catenations, upon timely removal of condensin I, chromosomes present high levels of de novo Topoisomerase II (TopoII)-dependent re-entanglements, and complete failure in chromosome segregation. TopoII is thus capable of re-intertwining previously separated DNA molecules and condensin I continuously required to counteract this erroneous activity. It is proposed that maintenance of chromosome resolution is a highly dynamic bidirectional process.
Farrell, D. L., Weitz, O., Magnasco, M. O. and Zallen, J. A. (2017). SEGGA: a toolset for rapid automated analysis of epithelial cell polarity and dynamics. Development 144(9): 1725-1734. PubMed ID: 28465336
Epithelial remodeling determines the structure of many organs in the body through changes in cell shape, polarity and behavior and is a major area of study in developmental biology. Accurate and high-throughput methods are necessary to systematically analyze epithelial organization and dynamics at single-cell resolution. This study developed SEGGA, an easy-to-use software for automated image segmentation, cell tracking and quantitative analysis of cell shape, polarity and behavior in epithelial tissues. SEGGA is free, open source, and provides a full suite of tools that allow users with no prior computational expertise to independently perform all steps of automated image segmentation, semi-automated user-guided error correction, and data analysis. This study uses SEGGA to analyze changes in cell shape, cell interactions and planar polarity during convergent extension in the Drosophila embryo. These studies demonstrate that planar polarity is rapidly established in a spatiotemporally regulated pattern that is dynamically remodeled in response to changes in cell orientation. These findings reveal an unexpected plasticity that maintains coordinated planar polarity in actively moving populations through the continual realignment of cell polarity with the tissue axes.
Karg, T., Elting, M. W., Vicars, H., Dumont, S. and Sullivan, W. (2017). The chromokinesin Klp3a and microtubules facilitate acentric chromosome segregation. J Cell Biol. PubMed ID: 28500183
Although poleward segregation of acentric chromosomes is well documented, the underlying mechanisms remain poorly understood. This study demonstrates that microtubules play a key role in poleward movement of acentric chromosome fragments generated in Drosophila melanogaster neuroblasts. Acentrics segregate with either telomeres leading or lagging in equal frequency and are preferentially associated with peripheral bundled microtubules. In addition, laser ablation studies demonstrate that segregating acentrics are mechanically associated with microtubules. Finally, this study shows that successful acentric segregation requires the chromokinesin Klp3a. Reduced Klp3a function results in disorganized interpolar microtubules and shortened spindles. Normally, acentric poleward segregation occurs at the periphery of the spindle in association with interpolar microtubules. In klp3a mutants, acentrics fail to localize and segregate along the peripheral interpolar microtubules and are abnormally positioned in the spindle interior. These studies demonstrate an unsuspected role for interpolar microtubules in driving acentric segregation.
Kyogoku, H. and Kitajima, T.S. (2017). Large cytoplasm is linked to the error-prone nature of oocytes. Dev Cell 41: 287-298. PubMed ID: 28486131
Evolutionary Homolog Study:
Chromosome segregation during meiosis in oocytes is error prone. The uniquely large cytoplasmic size of oocytes, which provides support for embryogenesis after fertilization, might be a predisposing factor for meiotic errors. However, this hypothesis remains unproven. This study shows that cytoplasmic size affects the functionality of the acentrosomal spindle. Artificially decreasing the cytoplasmic size in mouse oocytes allows the acentrosomal spindle poles to have a better-focused distribution of microtubule-organizing centers and to biorient chromosomes more efficiently, whereas enlargement of the cytoplasmic size has the opposite effects. Moreover, the cytoplasmic size-dependent dilution of nuclear factors, including anaphase inhibitors that are preformed at the nuclear membrane, limits the spindle's capacity to prevent anaphase entry with misaligned chromosomes. The present study defines a large cytoplasmic volume as a cell-intrinsic feature linked to the error-prone nature of oocytes. This may represent a trade-off between meiotic fidelity and post-fertilization developmental competence.

Tuesday, June 20th

Green, J., Adachi, A., Shah, K. K., Hirokawa, J. D., Magani, P. S. and Maimon, G. (2017). A neural circuit architecture for angular integration in Drosophila. Nature [Epub ahead of print]. PubMed ID: 28538731
Many animals keep track of their angular heading over time while navigating through their environment. However, a neural-circuit architecture for computing heading has not been experimentally defined in any species. This study describes a set of clockwise- and anticlockwise-shifting neurons in the Drosophila central complex whose wiring and physiology provide a means to rotate an angular heading estimate based on the fly's angular velocity. Each class of shifting neurons exists in two subtypes, with spatiotemporal activity profiles that suggest different roles for each subtype at the start and end of tethered-walking turns. Shifting neurons are required for the heading system to properly track the fly's heading in the dark, and stimulation of these neurons induces predictable shifts in the heading signal. The central features of this biological circuit are analogous to those of computational models proposed for head-direction cells in rodents and may shed light on how neural systems, in general, perform integration.
Julienne, H., Buhl, E., Leslie, D. S. and Hodge, J. J. L. (2017). Drosophila PINK1 and parkin loss-of-function mutants display a range of non-motor Parkinson's disease phenotypes. Neurobiol Dis 104: 15-23. PubMed ID: 28435104
Parkinson's disease (PD) is more commonly associated with its motor symptoms and the related degeneration of dopamine (DA) neurons. However, PD patients also display a wide range of non-motor symptoms, including memory deficits and disruptions of their sleep-wake cycles. These have a large impact on their quality of life, but their etiology is poorly understood. The fruit fly Drosophila has already been successfully used to model PD, and has been used extensively to study relevant non-motor behaviours in other contexts, but little attention has yet been paid to modelling non-motor symptoms of PD in this genetically tractable organism. This study examined memory performance and circadian rhythms in flies with loss-of-function mutations in two PD genes: PINK1 and parkin. Learning and memory abnormalities were found in both mutant genotypes, as well as a weakening of circadian rhythms that is underpinned by electrophysiological changes in clock neurons. This study paves the way for further work that may help us understand the mechanisms underlying these neglected aspects of PD, thus identifying new targets for treatments to address these non-motor problems specifically and perhaps even to halt disease progression in its prodromal phase.
Hattori, D., Aso, Y., Swartz, K. J., Rubin, G. M., Abbott, L. F. and Axel, R. (2017). Representations of novelty and familiarity in a mushroom body compartment. Cell 169(5): 956-969. PubMed ID: 28502772
Animals exhibit a behavioral response to novel sensory stimuli about which they have no prior knowledge. This study has examined the neural and behavioral correlates of novelty and familiarity in the olfactory system of Drosophila. Novel odors elicit strong activity in output neurons (MBONs) of the α'3 compartment of the mushroom body that is rapidly suppressed upon repeated exposure to the same odor. This transition in neural activity upon familiarization requires odor-evoked activity in the dopaminergic neuron innervating this compartment. Moreover, exposure of a fly to novel odors evokes an alerting response that can also be elicited by optogenetic activation of α'3 MBONs. Silencing these MBONs eliminates the alerting behavior. These data suggest that the α'3 compartment plays a causal role in the behavioral response to novel and familiar stimuli as a consequence of dopamine-mediated plasticity at the Kenyon cell-MBONα'3 synapse.
Grebler, R., Kistenpfennig, C., Rieger, D., Bentrop, J., Schneuwly, S., Senthilan, P. R. and Helfrich-Forster, C. (2017). Drosophila Rhodopsin 7 can partially replace the structural role of Rhodopsin 1, but not its physiological function. J Comp Physiol A Neuroethol Sens Neural Behav Physiol. PubMed ID: 28500442
Rhodopsin 7 (Rh7), a new invertebrate Rhodopsin gene, was discovered in the genome of Drosophila melanogaster in 2000 and thought to encode for a functional Rhodopsin protein. Indeed, Rh7 exhibits most hallmarks of the known Rhodopsins, except for the G-protein-activating QAKK motif in the third cytoplasmic loop that is absent in Rh7. This study shows that Rh7 can partially substitute Rh1 in the outer receptor cells (R1-6) for rhabdomere maintenance, but that it cannot activate the phototransduction cascade in these cells. This speaks against a role of Rh7 as photopigment in R1-6, but does not exclude that it works in the inner photoreceptor cells.

Monday, June 19th

Cook, M. S., Cazin, C., Amoyel, M., Yamamoto, S., Bach, E. and Nystul, T. (2017). Neutral competition for Drosophila follicle and cyst stem cell niches requires vesicle trafficking genes. Genetics [Epub ahead of print]. PubMed ID: 28512187
The process of selecting for cellular fitness through competition plays a critical role in both development and disease. The germarium, a structure at the tip of the ovariole of a Drosophila ovary, contains two follicle stem cells (FSCs) that undergo neutral competition for the stem cell niche. Using the FSCs as a model, a genetic screen through a collection of 126 mutants in essential genes on the X chromosome was performed to identify candidates that increase or decrease competition for the FSC niche. Approximately 55% and 6% of the mutations screened were obtained as putative FSC hypo- or hypercompetitors, respectively. A large majority of mutations were found in vesicle trafficking genes (11 out of the 13 in the collection of mutants) are candidate hypocompetition alleles, and the hypocompetition phenotype was confirmed for four of these alleles. Sec16 and another COP II vesicle trafficking component, Sar1, are required for follicle cell differentiation. Lastly, it was demonstrated that although some components of vesicle trafficking are also required for neutral competition in the cyst stem cells (CySCs) of the testis, there are important tissue-specific differences. These results demonstrate a critical role for vesicle trafficking in stem cell niche competition and differentiation, and a number of putative candidates for further exploration were identified.
Cheng, M. H., Andrejka, L., Vorster, P. J., Hinman, A. and Lipsick, J. S. (2017). The Drosophila LIN54 homolog Mip120 controls two aspects of oogenesis. Biol Open [Epub ahead of print]. PubMed ID: 28522430
The conserved multi-protein MuvB core associates with the Myb oncoproteins and with the RB-E2F-DP tumor suppressor proteins in complexes that regulate cell proliferation, differentiation, and apoptosis. Drosophila Mip120, a homolog of LIN54, is a sequence-specific DNA-binding protein within the MuvB core. A mutant of Drosophila mip120 was previously shown to cause female and male sterility. This study shows that Mip120 regulates two different aspects of oogenesis. First, in the absence of the Mip120 protein, egg chambers arrest during the transition from stage 7 to 8 with a failure of the normal program of chromosomal dynamics in the ovarian nurse cells. Specifically, the decondensation, disassembly and dispersion of the endoreplicated polytene chromosomes fail to occur without Mip120. The conserved carboxy-terminal DNA-binding and protein-protein interaction domains of Mip120 are necessary but are not sufficient for this process. Second, a lack of Mip120 was shown to cause a dramatic increase in the expression of benign gonial cell neoplasm (bgcn), a gene that is normally expressed in only a small number of cells within the ovary including the germline stem cells.
Ji, S., Li, C., Hu, L., Liu, K., Mei, J., Luo, Y., Tao, Y., Xia, Z., Sun, Q. and Chen, D. (2017). Bam-dependent deubiquitinase complex can disrupt germ-line stem cell maintenance by targeting cyclin A. Proc Natl Acad Sci U S A. PubMed ID: 28484036
Drosophila germ-line stem cells (GSCs) provide an excellent model to study the regulatory mechanisms of stem cells in vivo. Bag of marbles (bam) has been demonstrated to be necessary and sufficient to promote GSC and cystoblast differentiation. Despite extensive investigation of its regulation and genetic functions, the biochemical nature of the Bam protein has been unknown. This study reports that Bam is an ubiquitin-associated protein and controls the turnover of cyclin A (CycA). Mechanistically, it was found that Bam associated with Otu to form a deubiquitinase complex that stabilized CycA by deubiquitination, thus providing a mechanism to explain how ectopic expression of Bam in GSCs promotes differentiation. Collectively, these findings not only identify a biochemical function of Bam, which contributes to GSC fate determination, but also emphasizes the critical role of proper expression of cyclin proteins mediated by both ubiquitination and deubiquitination pathways in balancing stem cell self-renewal and differentiation.
Kim, J., Lu, C., Srinivasan, S., Awe, S., Brehm, A. and Fuller, M. T. (2017). Blocking promiscuous activation at cryptic promoters directs cell type-specific gene expression. Science 356(6339): 717-721. PubMed ID: 28522526
To selectively express cell type-specific transcripts during development, it is critical to maintain genes required for other lineages in a silent state. This study shows in the Drosophila male germline stem cell lineage that a spermatocyte-specific zinc finger protein, Kumgang (Kmg; CG5204), working with the chromatin remodeler dMi-2 prevents transcription of genes normally expressed only in somatic lineages. By blocking transcription from normally cryptic promoters, Kmg restricts activation by Aly, a component of the testis-meiotic arrest complex, to transcripts for male germ cell differentiation. These results suggest that as new regions of the genome become open for transcription during terminal differentiation, blocking the action of a promiscuous activator on cryptic promoters is a critical mechanism for specifying precise gene activation (Kim, 2017).

Sunday, June 18th

Gill, J., Park, Y., McGinnis, J. P., Perez-Sanchez, C., Blanchette, M. and Si, K. (2017). Regulated intron removal integrates motivational state and experience. Cell 169(5): 836-848. PubMed ID: 28525754
Myriad experiences produce transient memory, yet, contingent on the internal state of the organism and the saliency of the experience, only some memories persist over time. How experience and internal state influence the duration of memory at the molecular level remains unknown. A self-assembled aggregated state of Drosophila Orb2A protein is required specifically for long-lasting memory. In the adult fly brain the mRNA encoding Orb2A protein exists in an unspliced non-protein-coding form. The convergence of experience and internal drive transiently increases the spliced protein-coding Orb2A mRNA. A screen identified pasilla, the fly ortholog of mammalian Nova-1/2, as a mediator of Orb2A mRNA processing. A single-nucleotide substitution in the intronic region that reduces Pasilla binding and intron removal selectively impairs long-term memory. It is posited that pasilla-mediated processing of unspliced Orb2A mRNA integrates experience and internal state to control Orb2A protein abundance and long-term memory formation.
Akkouche, A., Mugat, B., Barckmann, B., Varela-Chavez, C., Li, B., Raffel, R., Pelisson, A. and Chambeyron, S. (2017). Piwi is required during Drosophila embryogenesis to license dual-strand piRNA clusters for transposon repression in adult ovaries. Mol Cell 66(3): 411-419. PubMed ID: 28457744
Most piRNAs in the Drosophila female germline are transcribed from heterochromatic regions called dual-strand piRNA clusters. Histone 3 lysine 9 trimethylation (H3K9me3) is required for licensing piRNA production by these clusters. However, it is unclear when and how they acquire this permissive heterochromatic state. This study shows that transient Piwi depletion in Drosophila embryos results in H3K9me3 decrease at piRNA clusters in ovaries. This is accompanied by impaired biogenesis of ovarian piRNAs, accumulation of transposable element transcripts, and female sterility. Conversely, Piwi depletion at later developmental stages does not disturb piRNA cluster licensing. These results indicate that the identity of piRNA clusters is epigenetically acquired in a Piwi-dependent manner during embryonic development, which is reminiscent of the widespread genome reprogramming occurring during early mammalian zygotic development.
Bartoletti, R., Capozzoli, B., Moore, J., Moran, J., Shrawder, B. and Vivekanand, P. (2017). Short hairpin RNA is more effective than long hairpin RNA in eliciting pointed loss-of-function phenotypes in Drosophila. Genesis [Epub ahead of print]. PubMed ID: 28464429
Pointed (Pnt) is a transcriptional activator that functions downstream of the highly conserved Receptor Tyrosine Kinase (RTK) signaling pathway. Pnt is an ETS family transcription factor and encodes for two proteins, PntP1 and PntP2. However, while PntP1 is constitutively active, PntP2 is only active after being phosphorylated by MAPK in the RTK pathway. As mutations in pnt perturb the development of several tissues, the effect and efficacy of using RNAi to target Pnt was examined. pnt RNAi was expressed in the eyes, oocyte, and heart cells using three different RNAi lines: Valium20, Valium10, and VDRC. Valium20 is distinct since it generates a short hairpin RNA (shRNA), while Valium10 and VDRC produce long hairpin dsRNA. It was found that for each tissue examined Valium20 exhibited the strongest phenotype while the Valium10 and VDRC lines produced varying levels of severity; the long hairpin RNA produced by the Valium10 and VDRC lines are unable to effectively knockdown pnt in embryonic tissues.
Tassetto, M., Kunitomi, M. and Andino, R. (2017). Circulating immune cells mediate a systemic RNAi-based adaptive antiviral response in Drosophila. Cell 169(2): 314-325. PubMed ID: 28388413
Effective antiviral protection in multicellular organisms relies on both cell-autonomous and systemic immunity. Systemic immunity mediates the spread of antiviral signals from infection sites to distant uninfected tissues. In arthropods, RNA interference (RNAi) is responsible for antiviral defense. This study shows that flies have a sophisticated systemic RNAi-based immunity mediated by macrophage-like haemocytes. Haemocytes take up dsRNA from infected cells and, through endogenous transposon reverse transcriptases, produce virus-derived complementary DNAs (vDNA). These vDNAs template de novo synthesis of secondary viral siRNAs (vsRNA), which are secreted in exosome-like vesicles. Strikingly, exosomes containing vsRNAs, purified from haemolymph of infected flies, confer passive protection against virus challenge in naive animals. Thus, similar to vertebrates, insects use immune cells to generate immunological memory in the form of stable vDNAs that generate systemic immunity, which is mediated by the vsRNA-containing exosomes.

Saturday, June 17th

Calles-Garcia, D., Yang, M., Soya, N., Melero, R., Menade, M., Ito, Y., Vargas, J., Lukacs, G. L., Kollman, J. M., Kozlov, G. and Gehring, K. (2017). Single-particle electron microscopy structure of UDP-glucose:glycoprotein glucosyltransferase suggests a selectivity mechanism for misfolded proteins. J Biol Chem. PubMed ID: 28490633
The enzyme UDP-glucose:glycoprotein glucosyltransferase (UGGT) mediates quality control of glycoproteins in the endoplasmic reticulum by attaching glucose to N-linked glycan of misfolded proteins. As a sensor, UGGT ensures that misfolded proteins are recognized by the lectin chaperones and do not leave the secretory pathway. The structure of UGGT and the mechanism of its selectivity for misfolded proteins have been unknown for 25 years. This study used negative-stain electron microscopy and small-angle X-ray scattering to determine the structure of UGGT from Drosophila melanogaster at 18 Å resolution. Three-dimensional reconstructions revealed a cage-like structure with a large central cavity. Particle classification revealed flexibility that precluded determination of a high-resolution structure. Introduction of biotinylation sites into a fungal UGGT expressed in E. coli allowed identification of the catalytic and first thioredoxin-like domain. Hydrogen-deuterium exchange mass spectrometry was used to map the binding site of an accessory protein, Sep15, to the first thioredoxin-like domain. The UGGT structural features identified suggest that the central cavity contains the catalytic site and is lined with hydrophobic surfaces. This enhances the binding of misfolded substrates with exposed hydrophobic residues and excludes folded proteins with hydrophilic surfaces. In conclusion, this study has determined the UGGT structure, which enabled development of a plausible functional model of the mechanism for UGGT's selectivity for misfolded glycoproteins.
Fish, J. E., Gutierrez, M. C., Dang, L. T., Khyzha, N., Chen, Z., Veitch, S., Cheng, H. S., Khor, M., Antounians, L., Njock, M. S., Boudreau, E., Herman, A. M., Rhyner, A. M., Ruiz, O. E., Eisenhoffer, G. T., Medina-Rivera, A., Wilson, M. D. and Wythe, J. D. (2017). Dynamic regulation of VEGF-inducible genes by an ERK-ERG-p300 transcriptional network. Development. PubMed ID: 28536097
Evolutionary Homolog Study

The transcriptional pathways activated downstream of Vascular Endothelial Growth Factor (VEGF; see Drosophila Pvf1) signaling during angiogenesis remain incompletely characterized. By assessing the signals responsible for induction of the Notch ligand, Delta-Like 4 (DLL4; see Drosophila Delta) in endothelial cells this study found that activation of the MAPK/ERK pathway mirrors the rapid and dynamic induction of DLL4 transcription and that this pathway is required for DLL4 expression. Furthermore, VEGF/ERK signaling induces phosphorylation and activation of the ETS transcription factor ERG (see Drosophila Pointed), a prerequisite for DLL4 induction. Transcription of DLL4 coincides with dynamic ERG-dependent recruitment of the transcriptional co-activator p300 (see Drosophila Nejire). Genome-wide gene expression profiling identified a network of VEGF-responsive and ERG-dependent genes, and ERG ChIP-seq revealed the presence of conserved ERG-bound putative enhancer elements near these target genes. Functional experiments performed in vitro and in vivo confirm that this network of genes requires ERK, ERG, and p300 activity. Finally, genome-editing and transgenic approaches demonstrate that a highly conserved ERG-bound enhancer located upstream of HLX (a transcription factor implicated in sprouting angiogenesis; see Drosophila Homeodomain protein 2.0) is required for its VEGF-mediated induction. Collectively, these findings elucidate a novel transcriptional pathway contributing to VEGF-dependent angiogenesis.

Nakamura, T., et al. (2017). Novel role of Rac-Mid1 signaling in medial cerebellar development. Development 144(10): 1863-1875. PubMed ID: 28512198
Evolutionary Homolog Study
Rac (see Drosophila Rac1) signaling impacts a relatively large number of downstream targets; however, few studies have established an association between Rac pathways and pathological conditions. This study generated mice with double knockout of Rac1 and Rac3 (Atoh1-Cre;Rac1flox/flox;Rac3-/- ) in cerebellar granule neurons (CGNs). Impaired tangential migration at E16.5 was observed, as well as numerous apoptotic CGNs at the deepest layer of the external granule layer (EGL) in the medial cerebellum of Atoh1-Cre;Rac1flox/flox;Rac3-/- mice at P8. Atoh1-Cre;Rac1flox/flox;Rac3-/- CGNs differentiated normally until expression of p27kip1 (see Drosophila Dacapo) and NeuN in the deep EGL at P5. Primary CGNs and cerebellar microexplants from Atoh1-Cre;Rac1flox/flox;Rac3-/- mice exhibited impaired neuritogenesis, which was more apparent in Map2-positive dendrites. Such findings suggest that impaired tangential migration and final differentiation of CGNs have resulted in decreased cerebellum size and agenesis of the medial internal granule layer, respectively. Furthermore, Rac depleted/deleted cells exhibited decreased levels of Mid1 and impaired mTORC1 signaling. Mid1 depletion in CGNs produced mild impairments in neuritogenesis and reductions in mTORC1 signaling. Thus, a novel Rac-signaling pathway (Rac1-Mid1-mTORC1) may be involved in medial cerebellar development.
Gibbs, E. B., Lu, F., Portz, B., Fisher, M. J., Medellin, B. P., Laremore, T. N., Zhang, Y. J., Gilmour, D. S. and Showalter, S. A. (2017). Phosphorylation induces sequence-specific conformational switches in the RNA polymerase II C-terminal domain. Nat Commun 8: 15233. PubMed ID: 28497798
The carboxy-terminal domain (CTD) of the RNA polymerase II (Pol II) large subunit cycles through phosphorylation states that correlate with progression through the transcription cycle and regulate nascent mRNA processing. Structural analyses of yeast and mammalian CTD are hampered by their repetitive sequences. This study identified a region of the Drosophila melanogaster CTD that is essential for Pol II function in vivo and capitalized on natural sequence variations within it to facilitate structural analysis. Mass spectrometry and NMR spectroscopy reveal that hyper-Ser5 phosphorylation transforms the local structure of this region via proline isomerization. The sequence context of this switch tunes the activity of the phosphatase Ssu72, leading to the preferential de-phosphorylation of specific heptads. Together, context-dependent conformational switches and biased dephosphorylation suggest a mechanism for the selective recruitment of cis-proline-specific regulatory factors and region-specific modulation of the CTD code that may augment gene regulation in developmentally complex organisms.

Friday, June 16th

Sharp, B., Paquet, E., Naef, F., Bafna, A. and Wijnen, H. (2017). A new promoter element associated with daily time keeping in Drosophila. Nucleic Acids Res. PubMed ID: 28407113
Circadian clocks are autonomous daily timekeeping mechanisms that allow organisms to adapt to environmental rhythms as well as temporally organize biological functions. Clock-controlled timekeeping involves extensive regulation of rhythmic gene expression. To date, relatively few clock-associated promoter elements have been identified and characterized. In an unbiased search of core clock gene promoters from 12 species of Drosophila, a 29-bp consensus sequence was discovered that has been designated as the Clock-Associated Transcriptional Activation Cassette or 'CATAC'. To experimentally address the spatiotemporal expression information associated with this element, constructs were generated with four separate native CATAC elements upstream of a basal promoter driving expression of either the yeast Gal4 or firefly luciferase reporter genes. Reporter assays showed that presence of wild-type, but not mutated CATAC elements, imparted increased expression levels as well as rhythmic regulation. Part of the CATAC consensus sequence resembles the E-box binding site for the core circadian transcription factor CLOCK/CYCLE (CLK/CYC), and CATAC-mediated expression rhythms are lost in the presence of null mutations in either cyc or the gene encoding the CLK/CYC inhibitor, period (per). Nevertheless, the results indicate that CATAC's enhancer function persists in the absence of CLK/CYC. Thus, CATAC represents a novel cis-regulatory element encoding clock-controlled regulation.
Adhikary, R., Tan, Y. X., Liu, J., Zimmermann, J., Holcomb, M., Yvellez, C., Dawson, P. E. and Romesberg, F. E. (2017). Conformational heterogeneity and DNA recognition by the morphogen Bicoid. Biochemistry [Epub ahead of print]. PubMed ID: 28547993
The morphogenic activity of the Drosophila transcription factor Bicoid (Bcd) is controlled by its DNA binding homeodomain. The Bcd homeodomain appears to be unique as it can bind multiple DNA sequences and even RNA. All homeodomain proteins adopt a three-helix fold, with residues of the third helix mediating recognition of the nucleic acid target via interactions with the major groove. To begin to directly characterize the conformational heterogeneity in the homeodomain, C-D bonds were introduced within each structural element and their absorptions were characterized in the free and bound states, as well as during thermal denaturation. The data reveal that while residues within the first two helices experience unique environments, each environment is well-defined and similar in the presence and absence of bound DNA. In contrast, the data are consistent with residues within the recognition helix adopting multiple conformations, and while the binding of DNA does alter the environments, the conformational heterogeneity is similar in the bound and unbound states. Finally, thermal denaturation studies reveal that the conformational heterogeneity observed in this and previous studies results not from local instability and unfolding, as has been suggested for other transcription factors, but rather from the population of multiple stable conformations within the folded state of the protein. The results have important implications for how Bcd recognizes its different targets to mediate its critical developmental functions.
Fukaya, T., Lim, B. and Levine, M. (2017). Rapid rates of Pol II elongation in the Drosophila embryo. Curr Biol 27(9): 1387-1391. PubMed ID: 28457866
Elongation of RNA polymerase II (Pol II) is thought to be an important mechanism for regulating gene expression. This study measured the first wave of de novo transcription in living Drosophila embryos using dual-fluorescence detection of nascent transcripts containing 5' MS2 and 3' PP7 RNA stem loops. Pol II elongation rates of 2.4-3.0 kb/min were observed, approximately twice as fast as earlier estimates. The revised rates permit substantial levels of zygotic gene activity prior to the mid-blastula transition. Evidence is provided that variable rates of elongation are not a significant source of differential gene activity, suggesting that transcription initiation and Pol II release are the key determinants of gene control in development.
Rogers, W. A., Goyal, Y., Yamaya, K., Shvartsman, S. Y. and Levine, M. S. (2017). Uncoupling neurogenic gene networks in the Drosophila embryo. Genes Dev 31(7): 634-638. PubMed ID: 28428262
The EGF signaling pathway specifies neuronal identities in the Drosophila embryo by regulating developmental patterning genes such as intermediate neuroblasts defective (ind). EGFR is activated in the ventral midline and neurogenic ectoderm by the Spitz ligand, which is processed by the Rhomboid protease. CRISPR/Cas9 was used to delete defined rhomboid enhancers mediating expression at each site of Spitz processing. Surprisingly, the neurogenic ectoderm, not the ventral midline, was found to be the dominant source of EGF patterning activity. It is suggested that Drosophila is undergoing an evolutionary transition in central nervous system (CNS)-organizing activity from the ventral midline to the neurogenic ectoderm.

Thursday, June 15th

Du, L., Zhou, A., Patel, A., Rao, M., Anderson, K. and Roy, S. (2017). Unique patterns of organization and migration of FGF-expressing cells during Drosophila morphogenesis. Dev Biol [Epub ahead of print]. PubMed ID: 28502613
In Drosophila, the FGF homolog, branchless (bnl) is expressed in a dynamic and spatiotemporally restricted pattern to induce branching morphogenesis of the trachea, which expresses the Bnl-receptor, breathless (btl). A new strategy has been developed to determine bnl-expressing cells and study their interactions with the btl-expressing cells. To enable targeted gene expression specifically in the bnl expressing cells, a new LexA based bnl enhancer trap line was generated using CRISPR/Cas9 based genome editing. With this tool, new bnl-expressing cells, their unique organization and functional interactions with the btl-expressing cells were uncovered in a larval tracheoblast niche in the leg imaginal discs, in larval photoreceptors of the developing retina, and in the embryonic central nervous system. The targeted expression system also facilitated live imaging of simultaneously labeled Bnl sources and tracheal cells, which revealed a unique morphogenetic movement of the embryonic bnl- source. Migration of bnl- expressing cells may create a dynamic spatiotemporal pattern of the signal source necessary for the directional growth of the tracheal branch. The genetic tool and the comprehensive profile of expression, organization, and activity of various types of bnl-expressing cells described in this study provided an important foundation for future research investigating the mechanisms underlying Bnl signaling in tissue morphogenesis.
Zhang, C. U. and Cadigan, K. M. (2017). The matrix protein Tiggrin regulates plasmatocyte maturation in Drosophila larva. Development [Epub ahead of print]. PubMed ID: 28526755
The lymph gland (LG) is a major source of hematopoiesis during Drosophila development. In this tissue, prohemocytes differentiate into multiple lineages including macrophage-like plasmatocytes, which comprise the vast majority of mature hemocytes. Previous studies have uncovered genetic pathways that regulate prohemocyte maintenance and some cell fate choices between hemocyte lineages. However, less is known about how the plasmatocyte pool of the LG is established and matures. This study reports that Tiggrin, a matrix protein expressed in the LG, is a specific regulator of plasmatocyte maturation. Tiggrin mutants exhibit precocious maturation of plasmatocytes, while Tiggrin overexpression blocks this process, resulting in a buildup of intermediate progenitors (IPs) expressing prohemocyte and hemocyte markers. These IPs likely represent a transitory state in prohemocyte to plasmatocyte differentiation. It was also found that overexpression of Wee1 kinase, which slows G2/M progression, results in a phenotype similar to Tiggrin overexpression while String/Cdc25 expression phenocopies Tiggrin mutants. Further analysis revealed that Wee1 inhibits plasmatocyte maturation through up-regulation of Tiggrin transcription. These results elucidate connections between the extracellular matrix and cell cycle regulators in the regulation of hematopoiesis.
Firulli, B. A., Milliar, H., Toolan, K. P., Harkin, J., Fuchs, R. K., Robling, A. G. and Firulli, A. B. (2017). Defective Hand1 phosphoregulation uncovers essential roles for Hand1 in limb morphogenesis. Development [Epub ahead of print]. PubMed ID: 28576769
Evolutionary Homolog Study
The morphogenesis of the vertebrate limbs is a complex process where cell signaling and transcriptional regulation coordinate diverse structural adaptations across species. This study examined the consequences of altering Hand1 (see Drosophila Hand) dimer choice regulation within the developing vertebrate limbs. Although Hand1 deletion via the limb-specific Prx1-Cre reveals a non-essential role for Hand1 in limb morphogenesis, altering Hand1 phosphoregulation, and consequently Hand1 dimerization affinities, results in a severe truncation of anterior-proximal limb elements. Molecular analysis reveals a non-cell autonomous mechanism that causes widespread cell death within embryonic limb bud. In addition, changes were observed in proximal anterior gene regulation including a reduction in the expression of Irx3&5 (see Drosophila Araucan), Gli3 (see Drosophila Ci), and Alx4 (see Drosophila Aristaless), all of which are upregulated in Hand2 limb conditional knockouts. A reduction of Hand2 and Shh (see Drosophila Hedgehog) gene dosage improves the integrity of anterior limb structures validating this proposed mechanism.
Bahm, I., Barriga, E. H., Frolov, A., Theveneau, E., Frankel, P. and Mayor, R. (2017). PDGF controls contact inhibition of locomotion by regulating N-cadherin during neural crest migration. Development [Epub ahead of print]. PubMed ID: 28526750
Evolutionary Homolog Study
A fundamental property of neural crest (NC) migration is Contact inhibition of locomotion (CIL), a process by which cells change their direction of migration upon cell contact. CIL has been proven to be essential for NC migration in amphibian and zebrafish by controlling cell polarity in a cell contact dependent manner. Cell contact during CIL requires the participation of the cell adhesion molecule N-cadherin (see Drosophila CadN), which starts to be expressed by NC cells as a consequence of the switch between E- and N-cadherins during epithelial to mesenchymal transition (EMT). However, the mechanism that controls the upregulation of N-cadherin remains unknown. This study shows that PDGFRα (see Drosophila Pvr) and its ligand PDGF-A (see Drosophila Pvf1) are co-expressed in migrating cranial NC. Inhibition of PDGF-A/PDGFRα blocks NC migration by inhibiting N-cadherin and, consequently impairing CIL. Moreover, PI3K/AKT (see Drosophila Akt) was found to be a downstream effector of the PDGFRα cellular response during CIL. These results lead to a proposal that PDGF-A/PDGFRα signalling is a tissue-autonomous regulator of CIL by controlling N-cadherin upregulation during EMT. Finally, it was shown that once NC have undergone EMT, the same PDGF-A/PDGFRα works as NC chemoattractant guiding their directional migration.

Wednesday, June 14th

Byrnes, A. E. and Slep, K. C. (2017). TOG-tubulin binding specificity promotes microtubule dynamics and mitotic spindle formation. J Cell Biol [Epub ahead of print]. PubMed ID: 28512144
XMAP215, CLASP, and Crescerin use arrayed tubulin-binding tumor overexpressed gene (TOG) domains to modulate microtubule dynamics. It was hypothesized that TOGs have distinct architectures and tubulin-binding properties that underlie each family's ability to promote microtubule polymerization or pause. As a model, this study investigated the pentameric TOG array of a Drosophila melanogaster XMAP215 member, Msps. Msps TOGs were found to have distinct architectures that bind either free or polymerized tubulin, and that a polarized array drives microtubule polymerization. An engineered TOG1-2-5 array fully supported Msps-dependent microtubule polymerase activity. Requisite for this activity was a TOG5-specific N-terminal HEAT repeat that engaged microtubule lattice-incorporated tubulin. TOG5-microtubule binding maintained mitotic spindle formation as deleting or mutating TOG5 compromised spindle architecture and increased the mitotic index. Mad2 knockdown released the spindle assembly checkpoint triggered when TOG5-microtubule binding was compromised, indicating that TOG5 is essential for spindle function. These results reveal a TOG5-specific role in mitotic fidelity and support the hypothesis that architecturally distinct TOGs arranged in a sequence-specific order underlie TOG array microtubule regulator activity.
An, Y., Xue, G., Shaobo, Y., Mingxi, D., Zhou, X., Yu, W., Ishibashi, T., Zhang, L. and Yan, Y. (2017). Apical constriction is driven by a pulsatile apical myosin network in delaminating Drosophila neuroblasts. Development [Epub ahead of print]. PubMed ID: 28506995
Cell delamination is a conserved morphogenetic process important for generation of cell diversity and maintenance of tissue homeostasis. This study used Drosophila embryonic neuroblasts as a model to study the apical constriction process during cell delamination. Dynamic myosin signals both around the cell adherens junctions and underneath the cell apical surface in the neuroectoderm. On the cell apical cortex the non-junctional myosin forms flows and pulses, which are termed as medial myosin pulses. Quantitative differences in medial myosin pulse intensity and frequency are critical to distinguish delaminating neuroblasts from their neighbors. Inhibition of medial myosin pulses blocks delamination. The fate of neuroblasts is set apart from their neighbors by Notch signaling-mediated lateral inhibition. When Notch signaling activity was inhibited in the embryo, it was observed that small clusters of cells undergo apical constriction and display an abnormal apical myosin pattern. Together, this study demonstrates that a contractile actomyosin network across the apical cell surface is organized to drive apical constriction in delaminating neuroblasts.
Chanet, S., Miller, C. J., Vaishnav, E. D., Ermentrout, B., Davidson, L. A. and Martin, A. C. (2017). Actomyosin meshwork mechanosensing enables tissue shape to orient cell force. Nat Commun 8: 15014. PubMed ID: 28504247
Sculpting organism shape requires that cells produce forces with proper directionality. Thus, it is critical to understand how cells orient the cytoskeleton to produce forces that deform tissues. During Drosophila gastrulation, actomyosin contraction in ventral cells generates a long, narrow epithelial furrow, termed the ventral furrow, in which actomyosin fibres and tension are directed along the length of the furrow. Using a combination of genetic and mechanical perturbations that alter tissue shape, this study demonstrated that geometrical and mechanical constraints act as cues to orient the cytoskeleton and tension during ventral furrow formation. An in silico model of two-dimensional actomyosin meshwork contraction was developed, demonstrating that actomyosin meshworks exhibit an inherent force orienting mechanism in response to mechanical constraints. Together, these in vivo and in silico data provide a framework for understanding how cells orient force generation, establishing a role for geometrical and mechanical patterning of force production in tissues.
Chan, E. H., Chavadimane Shivakumar, P., Clement, R., Laugier, E. and Lenne, P. F. (2017). Patterned cortical tension mediated by N-cadherin controls cell geometric order in the Drosophila eye. Elife 6. PubMed ID: 28537220
Adhesion molecules hold cells together but also couple cell membranes to a contractile actomyosin network, which limits the expansion of cell contacts. Despite their fundamental role in tissue morphogenesis and tissue homeostasis, how adhesion molecules control cell shapes and cell patterns in tissues remains unclear. This study address this question in vivo using the Drosophila eye. Cone cell shapes were shown to depend little on adhesion bonds and mostly on contractile forces. However, N-cadherin has an indirect control on cell shape. At homotypic contacts, junctional N-cadherin bonds downregulate Myosin-II contractility. At heterotypic contacts with E-cadherin, unbound N-cadherin induces an asymmetric accumulation of Myosin-II, which leads to a highly contractile cell interface. Such differential regulation of contractility is essential for morphogenesis as loss of N-cadherin disrupts cell rearrangements. These results establish a quantitative link between adhesion and contractility and reveal an unprecedented role of N-cadherin on cell shapes and cell arrangements.

Tuesday, June 13th

Baumann, A. A., Texada, M. J., Chen, H. M., Etheredge, J. N., Miller, D. L., Picard, S., Warner, R., Truman, J. W. and Riddiford, L. M. (2017). Genetic tools to study juvenile hormone action in Drosophila. Sci Rep 7(1): 2132. PubMed ID: 28522854
The insect juvenile hormone receptor is a basic helix-loop-helix (bHLH), Per-Arnt-Sim (PAS) domain protein, a novel type of hormone receptor. In higher flies like Drosophila, the ancestral receptor germ cell-expressed (gce) gene has duplicated to yield the paralog Methoprene-tolerant (Met). These paralogous receptors share redundant function during development but play unique roles in adults. Some aspects of JH function apparently require one receptor or the other. To provide a foundation for studying JH receptor function, endogenous JH receptor expression was recapitulated with single cell resolution. Using Bacteria Artificial Chromosome (BAC) recombineering and a transgenic knock-in, a spatiotemporal expressional atlas was generated of Met and gce throughout development. JH receptor expression was recapitulated in known JH target tissues, in which temporal expression corresponds with periods of hormone sensitivity. Larval expression largely supports the notion of functional redundancy. Furthermore, this study provides the neuroanatomical distribution of JH receptors in both the larval and adult central nervous system, which will serve as a platform for future studies regarding JH action on insect behavior.
Zimmerman, S. G., Merrihew, G. E., MacCoss, M. J. and Berg, C. A. (2017). Proteomics analysis identifies orthologs of human chitinase-like proteins as inducers of tube-morphogenesis defects in Drosophila. Genetics [Epub ahead of print]. PubMed ID: 28404605
Elevated levels of human chitinase-like proteins (CLPs) are associated with numerous chronic inflammatory diseases and several cancers and can promote disease progression by remodeling tissue, activating signaling cascades, stimulating proliferation and migration, and by regulating adhesion. This study has identified Imaginal disc growth factors (Idgfs), orthologs of human CLPs CHI3L1, CHI3L2, and OVGP1, in a proteomics analysis designed to discover factors that regulate tube morphogenesis. The approach used magnetic beads to isolate a small population of specialized ovarian cells, cells that non-autonomously regulate morphogenesis of epithelial tubes that form and secrete eggshell structures called dorsal appendages. Elevated levels were detected of four of the six Idgf family members (Idgf1, Idgf2, Idgf4, and Idgf6) in flies mutant for Bullwinkle, which encodes a transcription factor and is a known regulator of dorsal-appendage tube morphogenesis. During oogenesis, dysregulation of Idgfs (either gain or loss of function) disrupts the formation of the dorsal-appendage tubes. Previous studies demonstrate roles for Drosophila Idgfs in innate immunity, wound healing, and cell proliferation and motility in cell culture. This study identified a novel role for Idgfs in both normal and aberrant tubulogenesis processes.
Brock, A. R., Seto, M. and Smith-Bolton, R. K. (2017). Cap-n-collar promotes tissue regeneration by regulating ROS and JNK signaling in the Drosophila wing imaginal disc. Genetics [Epub ahead of print]. PubMed ID: 28512185
Regeneration is a complex process that requires an organism to recognize and repair tissue damage, as well as grow and pattern new tissue. This study describes a genetic screen to identify novel regulators of regeneration. The Drosophila melanogaster larval wing primordium was ablated by inducing apoptosis in a spatially and temporally controlled manner, and the tissue was allowed to regenerate and repattern. To identify genes that regulate regeneration, a dominant modifier screen was carried out by assessing the amount and quality of regeneration in adult wings heterozygous for isogenic deficiencies. Thirty-one regions on the right arm of the third chromosome were identified that modify the regenerative response. Interestingly, several distinct phenotypes were observed: mutants that regenerated poorly, mutants that regenerated faster or better than wild type, and mutants that regenerated imperfectly and had patterning defects. One deficiency region was mapped to cap-n-collar (cnc), the Drosophila Nrf2 ortholog, which is required for regeneration. Cnc regulates reactive oxygen species levels in the regenerating epithelium, and affects JNK signaling, growth, debris localization, and pupariation timing. This study presents the results of the screen and proposes a model wherein Cnc regulates regeneration by maintaining an optimal level of reactive oxygen species to promote JNK signaling.
Sinagoga, K. L., Stone, W. J., Schiesser, J. V., Schweitzer, J. I., Sampson, L., Zheng, Y. and Wells, J. M. (2017). Distinct roles for the mTOR pathway in postnatal morphogenesis, maturation and function of pancreatic islets. Development [Epub ahead of print]. PubMed ID: 28576773
Evolutionary Homolog Study
While much is known about the molecular pathways that regulate embryonic development and adult homeostasis of the endocrine pancreas, little is known about what regulates early postnatal development and maturation of islets. Given that birth marks the first exposure to enteral nutrition, this study investigated how nutrient-regulated signaling pathways influence postnatal islet development. To do this loss-of-function studies were performed of mechanistic target of rapamycin (mTOR; see Drosophila Tor), a highly conserved kinase within a nutrient-sensing pathway known to regulate cellular growth, morphogenesis and metabolism. Deletion of mTOR in pancreatic endocrine cells had no significant effect on their embryonic development. However, within the first 2 weeks after birth, mTOR-deficient islets became dysmorphic, beta-cell maturation and function was impaired, and animals lost islet mass. Moreover, it was discovered that these distinct functions of mTOR are mediated by separate downstream branches of the pathway, in that mTORC1 (Raptor; see Drosophila Raptor) is the main complex mediating maturation and function of islets, whereas mTORC2 (Rictor; see Drosophila Rictor) impacts islet mass and architecture. Taken together, these findings suggest that nutrient-sensing may be a trigger for postnatal beta cell maturation and islet development.

Monday, June 12th

Yeung, K., Boija, A., Karlsson, E., Holmqvist, P. H., Tsatskis, Y., Nisoli, I., Yap, D. B., Lorzadeh, A., Moksa, M., Hirst, M., Aparicio, S., Fanto, M., Stenberg, P., Mannervik, M. and McNeill, H. (2017). Atrophin controls developmental signaling pathways via interactions with Trithorax-like. Elife 6. PubMed ID: 28327288
Mutations in human Atrophin1, a transcriptional corepressor, cause dentatorubral-pallidoluysian atrophy, a neurodegenerative disease. Drosophila Atrophin (Atro) mutants display many phenotypes, including neurodegeneration, segmentation, patterning and planar polarity defects. Despite Atro's critical role in development and disease, relatively little is known about Atro's binding partners and downstream targets. This study present the first genomic analysis of Atro using ChIP-seq against endogenous Atro. ChIP-seq identified 1300 potential direct targets of Atro including engrailed, and components of the Dpp and Notch signaling pathways. Atro regulates Dpp and Notch signaling in larval imaginal discs, at least partially via regulation of thickveins and fringe. In addition, bioinformatics analyses, sequential ChIP and coimmunoprecipitation experiments reveal that Atro interacts with the Drosophila GAGA Factor, Trithorax-like (Trl), and they bind to the same loci simultaneously. Phenotypic analyses of Trl and Atro clones suggest that Atro is required to modulate the transcription activation by Trl in larval imaginal discs. Taken together these data indicate that Atro is a major Trl cofactor that functions to moderate developmental gene transcription.
Asif-Laidin, A., Delmarre, V., Laurentie, J., Miller, W. J., Ronsseray, S. and Teysset, L. (2017). Short and long-term evolutionary dynamics of subtelomeric piRNA clusters in Drosophila. DNA Res [Epub ahead of print]. PubMed ID: 28459978
Two Telomeric Associated Sequences, TAS-R and TAS-L, form the principal subtelomeric repeat families identified in Drosophila melanogaster. They are PIWI-interacting RNA (piRNA) clusters involved in repression of Transposable Elements. This study revisited TAS structural and functional dynamics in D. melanogaster and in related species. In silico analysis revealed that TAS-R family members are composed of previously uncharacterized domains. This analysis also showed that TAS-L repeats are composed of arrays of a region termed 'TAS-L like' (TLL) identified specifically in one TAS-R family member, X-TAS. TLL were also present in other species of the melanogaster subgroup. Therefore, it is possible that TLL represents an ancestral subtelomeric piRNA core-cluster. Furthermore, all D. melanogaster genomes tested possessed at least one TAS-R locus, whereas TAS-L can be absent. A screen of 110 D. melanogaster lines showed that X-TAS is always present in flies living in the wild, but often absent in long-term laboratory stocks and that natural populations frequently lost their X-TAS within 2 years upon lab conditioning. Therefore, the unexpected structural and temporal dynamics of subtelomeric piRNA clusters demonstrated in this study suggests that genome organization is subjected to distinct selective pressures in the wild and upon domestication in the laboratory.
Ciabrelli, F., Comoglio, F., Fellous, S., Bonev, B., Ninova, M., Szabo, Q., Xuereb, A., Klopp, C., Aravin, A., Paro, R., Bantignies, F. and Cavalli, G. (2017). Stable Polycomb-dependent transgenerational inheritance of chromatin states in Drosophila. Nat Genet 49(6): 876-886. PubMed ID: 28436983
Transgenerational epigenetic inheritance (TEI) describes the transmission of alternative functional states through multiple generations in the presence of the same genomic DNA sequence. Very little is known about the principles and the molecular mechanisms governing this type of inheritance. In this study, by transiently enhancing 3D chromatin interactions, stable and isogenic Drosophila epilines were established that carry alternative epialleles, as defined by differential levels of Polycomb-dependent trimethylation of histone H3 Lys27 (forming H3K27me3). After being established, epialleles can be dominantly transmitted to naive flies and can induce paramutation. Importantly, epilines can be reset to a naive state by disruption of chromatin interactions. Finally, it was found that environmental changes modulate the expressivity of the epialleles, and this paradigm was extended to naturally occurring phenotypes. This work sheds light on how nuclear organization and Polycomb group (PcG) proteins contribute to epigenetically inheritable phenotypic variability.
Hansen, A. S., Pustova, I., Cattoglio, C., Tjian, R. and Darzacq, X. (2017). CTCF and cohesin regulate chromatin loop stability with distinct dynamics. Elife 6. PubMed ID: 28467304
Evolutionary Homolog Study
Folding of mammalian genomes into spatial domains is critical for gene regulation. The insulator protein CTCF (see Drosophila CTFC) and cohesin (see Drosophila Cohesin) control domain location by folding domains into loop structures, which are widely thought to be stable. Combining genomic and biochemical approaches this study shows that CTCF and cohesin co-occupy the same sites and physically interact as a biochemically stable complex. However, using single-molecule imaging it was found that CTCF binds chromatin much more dynamically than cohesin (~1-2 min vs. ~22 min residence time). Moreover, after unbinding, CTCF quickly rebinds another cognate site unlike cohesin for which the search process is long (~1 min vs. ~33 min). Thus, CTCF and cohesin form a rapidly exchanging 'dynamic complex' rather than a typical stable complex. Since CTCF and cohesin are required for loop domain formation, these results suggest that chromatin loops are dynamic and constantly break and reform throughout the cell cycle.

Sunday, June 11th

Chang, C.H. and Larracuente, A.M. (2017). Genomic changes following the reversal of a Y chromosome to an autosome in Drosophila pseudoobscura. Evolution [Epub ahead of print]. PubMed ID: 28322435
Robertsonian translocations resulting in fusions between sex chromosomes and autosomes shape Karyotype evolution by creating new sex chromosomes from autosomes. These translocations can also reverse sex chromosomes back into autosomes, which is especially intriguing given the dramatic differences between autosomes and sex chromosomes. To study the genomic events following a Y chromosome reversal, this study investigated an autosome-Y translocation in Drosophila pseudoobscura. The ancestral Y chromosome fused to a small autosome (the dot chromosome) approximately 10-15 Mya. Single molecule real-time sequencing reads were used to assemble the D. pseudoobscura dot chromosome, including the Y-to-dot translocation. It was found that the intervening sequence between the ancestral Y and the rest of the dot chromosome is only ∼78 Kb and is not repeat-dense, suggesting that the centromere now falls outside, rather than between, the fused chromosomes. The Y-to-dot region is 100 times smaller than the D. melanogaster Y chromosome, owing to changes in repeat landscape. However, a consistent reduction in intron sizes across the Y-to-dot region was not found. Instead, deletions in intergenic regions and possibly a small ancestral Y chromosome size may explain the compact size of the Y-to-dot translocation.
Bargues, N. and Lerat, E. (2017). Evolutionary history of LTR-retrotransposons among 20 Drosophila species. Mob DNA 8: 7. PubMed ID: 28465726
The presence of transposable elements (TEs) in genomes is known to explain in part the variations of genome sizes among eukaryotes. Even among closely related species, the variation of TE amount may be striking, as for example between the two sibling species, Drosophila melanogaster and D. simulans. However, not much is known concerning the TE content and dynamics among other Drosophila species. The sequencing of several Drosophila genomes, covering the two subgenus Sophophora and Drosophila, revealed a large variation of the repeat content among these species but no much information is known concerning their precise TE content. The identification of some consensus sequences of TEs from the various sequenced Drosophila species allowed to get an idea concerning their variety in term of diversity of superfamilies but the used classification remains very elusive and ambiguous. This study focused on LTR-retrotransposons because they represent the most widely represented class of TEs in the Drosophila genomes, describing the phylogenetic relationship of each LTR-retrotransposon family described in 20 Drosophila species, computing their proportion in their respective genomes and identifying several new cases of horizontal transfers. CAll these results give a clearer view on the evolutionary history of LTR retrotransposons among Drosophila that seems to be mainly driven by vertical transmissions although the implications of horizontal transfers, losses and intra-specific diversification are clearly also at play.
Wolff, J. N., Gemmell, N. J., Tompkins, D. M. and Dowling, D. K. (2017). Introduction of a male-harming mitochondrial haplotype via 'Trojan Females' achieves population suppression in fruit flies. Elife 6. PubMed ID: 28467301
Pests are a global threat to biodiversity, ecosystem function, and human health. Pest control approaches are thus numerous, but their implementation costly, damaging to non-target species, and ineffective at low population densities. The Trojan Female Technique (TFT) is a prospective self-perpetuating control technique that is species-specific and predicted to be effective at low densities. The goal of the TFT is to harness naturally-occurring mutations in the mitochondrial genome that impair male fertility while having no effect on females. This study provides proof-of-concept for the TFT, by showing that introduction of a male fertility-impairing mtDNA haplotype into replicated populations of Drosophila melanogaster causes numerical population suppression, with the magnitude of effect positively correlated with its frequency at trial inception. Further development of the TFT could lead to establishing a control strategy that overcomes limitations of conventional approaches, with broad applicability to invertebrate and vertebrate species, to control environmental and economic pests.
Rogers, R. L., Shao, L. and Thornton, K. R. (2017). Tandem duplications lead to novel expression patterns through exon shuffling in Drosophila yakuba. PLoS Genet 13(5): e1006795. PubMed ID: 28531189
One common hypothesis to explain the impacts of tandem duplications is that whole gene duplications commonly produce additive changes in gene expression due to copy number changes. This study used genome wide RNA-seq data from a population sample of Drosophila yakuba to test this 'gene dosage' hypothesis. Little evidence was observed of expression changes in response to whole transcript duplication capturing 5' and 3' UTRs. Among whole gene duplications, evidence was observed that dosage sharing across copies is likely to be common. The lack of expression changes after whole gene duplication suggests that the majority of genes are subject to tight regulatory control and therefore not sensitive to changes in gene copy number. Rather, changes were observed in expression level due to both shuffling of regulatory elements and the creation of chimeric structures via tandem duplication. Additionally, 30 de novo gene structures were observed arising from tandem duplications, 23 of which form with expression in the testes. Thus, the value of tandem duplications is likely to be more intricate than simple changes in gene dosage. The common regulatory effects from chimeric gene formation after tandem duplication may explain their contribution to genome evolution.

Saturday, June 10th

Ton, H. T., Phan, T. X., Abramyan, A. M., Shi, L. and Ahern, G. P. (2017). Identification of a putative binding site critical for general anesthetic activation of TRPA1. Proc Natl Acad Sci U S A 114(14): 3762-3767. PubMed ID: 28320952
General anesthetics suppress CNS activity by modulating the function of membrane ion channels, in particular, by enhancing activity of GABAA receptors (see Drosophila Rdl). In contrast, several volatile (isoflurane, desflurane) and i.v. (propofol) general anesthetics excite peripheral sensory nerves to cause pain and irritation upon administration. These noxious anesthetics activate transient receptor potential ankyrin repeat 1 (TRPA1), a major nociceptive ion channel, but the underlying mechanisms and site of action are unknown. This study exploited the observation that pungent anesthetics activate mammalian but not Drosophila TRPA1. Analysis of chimeric Drosophila and mouse TRPA1 channels reveal a critical role for the fifth transmembrane domain (S5) in sensing anesthetics. Interestingly, this study showed that anesthetics share with the antagonist A-967079 a potential binding pocket lined by residues in the S5, S6, and the first pore helix; isoflurane competitively disrupts A-967079 antagonism, and introducing these mammalian TRPA1 residues into dTRPA1 recapitulates anesthetic agonism. Furthermore, molecular modeling predicts that isoflurane and propofol bind to this pocket by forming H-bond and halogen-bond interactions with Ser-876, Met-915, and Met-956. Mutagenizing Met-915 or Met-956 selectively abolishes activation by isoflurane and propofol without affecting actions of A-967079 or the agonist, menthol. Thus, the combined experimental and computational results reveal the potential binding mode of noxious general anesthetics at TRPA1. These data may provide a structural basis for designing drugs to counter the noxious and vasorelaxant properties of general anesthetics and may prove useful in understanding effects of anesthetics on related ion channels.
Chakravorty, S., Tanner, B. C. W., Foelber, V. L., Vu, H., Rosenthal, M., Ruiz, T. and Vigoreaux, J. O. (2017). Flightin maintains myofilament lattice organization required for optimal flight power and courtship song quality in Drosophila. Proc Biol Sci 284(1854). PubMed ID: 28469022
The indirect flight muscles (IFMs) of Drosophila and other insects with asynchronous flight muscles are characterized by a crystalline myofilament lattice structure. The high-order lattice regularity is considered an adaptation for enhanced power output, but supporting evidence for this claim is lacking. This study shows that IFMs from transgenic flies expressing flightin with a deletion of its poorly conserved N-terminal domain (flnΔN62) have reduced inter-thick filament spacing and a less regular lattice. This resulted in a decrease in flight ability by 33% and in skinned fibre oscillatory power output by 57%, but had no effect on wingbeat frequency or frequency of maximum power output, suggesting that the underlying actomyosin kinetics is not affected and that the flight impairment arises from deficits in force transmission. Moreover, flnΔN62 males were shown to produced an abnormal courtship song characterized by a higher sine song frequency and a pulse song with longer pulses and longer inter-pulse intervals (IPIs), the latter implicated in male reproductive success. When presented with a choice, wild-type females chose control males over mutant males in 92% of the competition events. These results demonstrate that flightin N-terminal domain is required for optimal myofilament lattice regularity and IFM activity, enabling powered flight and courtship song production. As the courtship song is subject to female choice, it is proposed that the low amino acid sequence conservation of the N-terminal domain reflects its role in fine-tuning species-specific courtship songs.
Aleman-Meza, B., Loeza-Cabrera, M., Pena-Ramos, O., Stern, M. and Zhong, W. (2017). High-content behavioral profiling reveals neuronal genetic network modulating Drosophila larval locomotor program. BMC Genet 18(1): 40. PubMed ID: 28499390
Two key questions in understanding the genetic control of behaviors are: what genes are involved and how these genes interact. To answer these questions at a systems level, a high-content profiling of Drosophila larval locomotor behaviors was conducted for over 100 genotypes. 69 genes were studied whose C. elegans orthologs were neuronal signalling genes with significant locomotor phenotypes, and RNAi was conducted with ubiquitous, pan-neuronal, or motor-neuronal Gal4 drivers. Inactivation of 42 genes, including the nicotinic acetylcholine receptors nAChRalpha1 and nAChRalpha3, in the neurons caused significant movement defects. Bioinformatic analysis suggested 81 interactions among these genes based on phenotypic pattern similarities. Comparing the worm and fly data sets, this study found that these genes were highly conserved in having neuronal expressions and locomotor phenotypes. However, the genetic interactions were not conserved for ubiquitous profiles, and may be mildly conserved for the neuronal profiles. Unexpectedly, the data also revealed a possible motor-neuronal control of body size, because inactivation of Rdl and Galphao in the motor neurons reduced the larval body size. Overall, these data established a framework for further exploring the genetic control of Drosophila larval locomotion. In conclusion, high content, quantitative phenotyping of larval locomotor behaviours provides a framework for system-level understanding of the gene networks underlying such behaviours.
Chwen, Y., Lee, G., Yang, Q., Chi, W., Turkson, S. A., Du, W. A., Kemkemer, C., Zeng, Z. B., Long, M. and Zhuang, X. (2017). Genetic architecture of natural variation underlying adult foraging behavior that is essential for survival of Drosophila melanogaster. Genome Biol Evol. PubMed ID: 28472322
Foraging behavior is critical for the fitness of individuals. However, the genetic basis of variation in foraging behavior and the evolutionary forces underlying such natural variation have rarely been investigated. A systematic approach was developed to assay the variation in survival rate in a foraging environment for adult flies derived from a wild Drosophila melanogaster population. Despite being such an essential trait, there is substantial variation of foraging behavior among D. melanogaster strains. Importantly, this study provided the first evaluation of the potential caveats of using inbred Drosophila strains to perform genome-wide association studies on life-history traits, and concluded that inbreeding depression is unlikely a major contributor for the observed large variation in adult foraging behavior. Adult foraging behavior has a strong genetic component and, unlike larval foraging behavior, depends on multiple loci. Identified candidate genes are enriched in those with high expression in adult heads and, demonstrated by expression knock down assay, are involved in maintaining normal functions of the nervous system. This study not only identified candidate genes for foraging behavior that is relevant to individual fitness, but also shed light on the initial stage underlying the evolution of the behavior.

Friday, June 9th

Chen, D., Gu, T., Pham, T. N., Zachary, M. J. and Hewes, R. S. (2017). Regulatory mechanisms of metamorphic neuronal remodeling revealed through a genome-wide modifier screen in Drosophila melanogaster. Genetics [Epub ahead of print]. PubMed ID: 28476867
The RNA binding factor alan shepard (shep) is an important regulator of neuronal remodeling during metamorphosis in Drosophila melanogaster, and loss of shep leads to smaller soma size and fewer neurites in a stage-dependent manner. To shed light on the mechanisms by which shep regulates neuronal remodeling, a genetic modifier screen was conducted for suppressors of shep-dependent wing expansion defects and cellular morphological defects in a set of peptidergic neurons, the bursicon neurons, that promote post-eclosion wing expansion. Out of 702 screened deficiencies that covered 86% of euchromatic genes, 24 deficiencies were isolated as candidate suppressors, and 12 of them at least partially suppressed morphological defects in shep mutant bursicon neurons. With RNAi and mutant alleles of individual genes, Daughters against dpp (Dad) and Olig family (Oli) were identified as shep suppressor genes, and both of them restored the adult cellular morphology of shep-depleted bursicon neurons. Dad encodes an inhibitory Smad protein that inhibits bone morphogenetic protein (BMP) signaling, raising the possibility that shep interacted with BMP signaling through antagonism of Dad. By manipulating expression of the BMP receptor tkv, activated BMP signaling was found to be sufficient to rescue loss-of-shep phenotypes. These findings reveal mechanisms of shep regulation during neuronal development, and they highlight a novel genetic shep interaction with the BMP signaling pathway that controls morphogenesis in mature, terminally differentiated neurons during metamorphosis.
Chow, E. S., Long, D. M. and Giebultowicz, J. M. (2016). Circadian rhythm in mRNA expression of the glutathione synthesis gene Gclc is controlled by peripheral glial clocks in Drosophila melanogaster. C. Physiol Entomol 41(4): 369-377. PubMed ID: 28503020
Circadian coordination of metabolism, physiology, and behaviour is found in all living kingdoms. Clock genes are transcriptional regulators, and their rhythmic activities generate daily rhythms in clock-controlled genes which result in cellular and organismal rhythms. Insects provide numerous examples of rhythms in behaviour and reproduction, but less is known about control of metabolic processes by circadian clocks in insects. Recent data suggest that several pathways involved in protecting cells from oxidative stress may be modulated by the circadian system, including genes involved in glutathione (GSH) biosynthesis. Specifically, rhythmic expression of the gene encoding the catalytic subunit (Gclc) of the rate-limiting GSH biosynthetic enzyme was detected in Drosophila melanogaster heads. The aim of this study was to determine which clocks in the fly multi-oscillatory circadian system are responsible for Gclc rhythms. Genetic disruption of tissue-specific clocks in D. melanogaster revealed that transcriptional rhythms in Gclc mRNA levels occur independently of the central pacemaker neurons, because these rhythms persisted in heads of behaviourally arrhythmic flies with a disabled central clock but intact peripheral clocks. Disrupting the clock specifically in glial cells abolished rhythmic expression of Gclc, suggesting that glia play an important role in Gclc transcriptional regulation, which may contribute to maintaining homeostasis in the fly nervous system.
Chang, P. Y., Su, T. S., Shih, C. T. and Lo, C. C. (2017). The topographical mapping in Drosophila central complex network and its signal routing. Front Neuroinform 11: 26. PubMed ID: 28443014
Neural networks regulate brain functions by routing signals. Therefore, investigating the detailed organization of a neural circuit at the cellular levels is a crucial step toward understanding the neural mechanisms of brain functions. To study how a complicated neural circuit is organized, this study analyzed recently published data on the neural circuit of the Drosophila central complex, a brain structure associated with a variety of functions including sensory integration and coordination of locomotion. Except for a small number of 'atypical' neuron types, it was found that the network structure formed by the identified 194 neuron types can be described by only a few simple mathematical rules. Specifically, the topological mapping formed by these neurons can be reconstructed by applying a generation matrix on a small set of initial neurons. By analyzing how information flows propagate with or without the atypical neurons, it was found that while the general pattern of signal propagation in the central complex follows the simple topological mapping formed by the 'typical' neurons, some atypical neurons can substantially re-route the signal pathways, implying specific roles of these neurons in sensory signal integration. The present study provides insights into the organization principle and signal integration in the central complex.
Chen, Y., Cameron, S., Chang, W. T. and Rao, Y. (2017). Turtle interacts with borderless in regulating glial extension and axon ensheathment. Mol Brain 10(1): 17. PubMed ID: 28535795
Proper recognition between axons and glial processes is required for the establishment of axon ensheathment in the developing nervous system. Recent studies have begun to reveal molecular events underlying developmental control of axon-glia recognition. Previous work has shown that the transmembrane protein Borderless (Bdl) is specifically expressed in wrapping glia (WG), and is required for the extension of glial processes and the ensheathment of photoreceptor axons in the developing Drosophila visual system. The exact mechanism by which Bdl mediates axon-glia recognition, however, remains unknown. This study presents evidence showing that Bdl interacts with the Ig transmembrane protein Turtle (Tutl). Tutl is specifically expressed in photoreceptor axons. Loss of tutl in photoreceptors, like loss of bdl in WG, disrupts glial extension and axon ensheatment. Epistasis analysis shows that Tutl interacts genetically with Bdl. Tutl interacts with Bdl in trans in cultured cells. It is proposed that Tutl interacts with Bdl in mediating axon-glia recognition for WG extension and axon ensheathment.

Thursday, June 8th

Lin, C.M., Xu, J., Yang, W.T., Wang, C., Li, Y.C., Cheng, L.C., Zhang, L. and Hsu, J.C. (2017). Smurf downregulates Echinoid in the amnioserosa to regulate Drosophila dorsal closure. Genetics [Epub ahead of print]. PubMed ID: 28428287
Drosophila dorsal closure is a morphogenetic movement that involves flanking epidermal cells, assembling actomyosin cables and migrating dorsally over the underlying amnioserosa to seal at the dorsal midline. Echinoid (Ed), a cell adhesion molecule of adherens junctions (AJs), participates in several developmental processes. The disappearance of Ed from the amnioserosa is required to define the epidermal leading edge for actomyosin cable assembly and coordinated cell migration. However, the mechanism by which Ed is cleared from amnioserosa is unknown. This study shows that Ed is cleared in amnioserosa by both transcriptional and post-translational mechanisms. First, Ed mRNA transcription is repressed in amnioserosa prior to the onset of dorsal closure. Second, the ubiquitin ligase Smurf downregulates pre-translated Ed by binding to the PPXY motif of Ed. During dorsal closure, Smurf colocalizes with Ed at AJs, and Smurf overexpression prematurely degrades Ed in the amnioserosa. Conversely, Ed persists in the amnioserosa of Smurf mutant embryos which in turn affects actomyosin cable formation. Together, these results demonstrate that transcriptional repression of Ed followed by Smurf-mediated downregulation of pre-translated Ed in amnioserosa regulates the establishment of a taut leading edge during dorsal closure.

Patel, T. and Hobert, O. (2017). Coordinated control of terminal differentiation and restriction of cellular plasticity. Elife 6. PubMed ID: 28422646
Evolutionary Homolog Study:
The acquisition of a specific cellular identity is usually paralleled by a restriction of cellular plasticity. Whether and how these two processes are coordinated is poorly understood. Transcription factors called terminal selectors activate identity-specific effector genes during neuronal differentiation to define the structural and functional properties of a neuron. To study restriction of plasticity, this study ectopically expressed C. elegans CHE-1 (see Drosophila glass), a terminal selector of ASE sensory neuron (see Drosophila sensory neurons) identity. In undifferentiated cells, ectopic expression of CHE-1 results in activation of ASE neuron type-specific effector genes. Once cells differentiate, their plasticity is restricted and ectopic expression of CHE-1 no longer results in activation of ASE effector genes. In striking contrast, removal of the respective terminal selectors of other sensory, inter-, or motor neuron types now enables ectopically expressed CHE-1 to activate its ASE-specific effector genes, indicating that terminal selectors not only activate effector gene batteries but also control the restriction of cellular plasticity. Terminal selectors mediate this restriction at least partially by organizing chromatin. The chromatin structure of a CHE-1 target locus is less compact in neurons that lack their resident terminal selector and genetic epistasis studies with H3K9 methyltransferases suggest that this chromatin modification acts downstream of a terminal selector to restrict plasticity. Taken together, terminal selectors activate identity-specific genes and make non-identity-defining genes less accessible, thereby serving as a checkpoint to coordinate identity specification with restriction of cellular plasticity.

Sun, Z., Amourda, C., Shagirov, M., Hara, Y., Saunders, T. E. and Toyama, Y. (2017). Basolateral protrusion and apical contraction cooperatively drive Drosophila germ-band extension. Nat Cell Biol 19(4): 375-383. PubMed ID: 28346438
During Drosophila germ-band extension (GBE), cell intercalation is the key mechanism for tissue extension, and the associated apical junction remodelling is driven by polarized myosin-II-dependent contraction. However, the contribution of the basolateral cellular mechanics to GBE remains poorly understood. This study characterized how cells coordinate their shape from the apical to the basal side during rosette formation, a hallmark of cell intercalation. Basolateral rosette formation is driven by cells mostly located at the dorsal/ventral part of the rosette (D/V cells). These cells exhibit actin-rich wedge-shaped basolateral protrusions and migrate towards each other. Surprisingly, the formation of basolateral rosettes precedes that of the apical rosettes. Basolateral rosette formation is independent of apical contractility, but requires Rac1-dependent protrusive motility. Furthermore, Src42A was identified as a regulator of basolateral rosette formation. The data show that in addition to apical contraction, active cell migration driven by basolateral protrusions plays a pivotal role in rosette formation and contributes to GBE.
Rizzo, N. P. and Bejsovec, A. (2017). SoxNeuro and shavenbaby act cooperatively to shape denticles in the embryonic epidermis of Drosophila. Development [Epub ahead of print]. PubMed ID: 28506986
During development, extracellular signals are integrated by cells to induce the transcriptional circuitry that controls morphogenesis. In the fly epidermis, Wingless (Wg)/Wnt signaling directs cells to produce either a distinctly-shaped denticle or no denticle, resulting in a segmental pattern of denticle belts separated by smooth, or 'naked', cuticle. Naked cuticle results from Wg repression of shavenbaby (svb), which encodes a transcription factor required for denticle construction. This study has discovered that although the svb promoter responds differentially to altered Wg levels, Svb alone cannot produce the morphological diversity of denticles found in wild-type belts. Instead, a second Wg-responsive transcription factor, SoxNeuro (SoxN), cooperates with Svb to shape the denticles. Co-expressing ectopic SoxN with svb rescued diverse denticle morphologies. Conversely, removing SoxN activity eliminated the residual denticles found in svb mutant embryos. Furthermore, several known Svb target genes are also activated by SoxN, and two novel target genes of SoxN were discovered that are expressed in denticle-producing cells and that are regulated independently of Svb. Thus it is concluded that proper denticle morphogenesis requires transcriptional regulation by both SoxN and Svb.

Wednesday June 7th

Chakraborty, S. and Hasan, G. (2017). Spontaneous Ca2+ influx in Drosophila pupal neurons is modulated by IP3-receptor function and influences maturation of the flight circuit. Front Mol Neurosci 10: 111. PubMed ID: 28473752
Inositol 1,4,5-trisphosphate receptors (IP3R) are Ca2+ channels on the neuronal endoplasmic reticulum (ER) membrane. They are gated by IP3, produced upon external stimulation and activation of G protein-coupled receptors on the plasma membrane (PM). IP3-mediated Ca2+ release, and the resulting depletion of the ER store, triggers entry of extracellular Ca2+ by store-operated Ca2+ entry (SOCE). Mutations in IP3R attenuate SOCE. Compromised IP3R function and SOCE during pupal development of Drosophila leads to flight deficits and mimics suppression of neuronal activity during pupal or adult development. To understand the effect of compromised IP3R function on pupal neuronal calcium signaling, the effects were examined of mutations in the IP3R gene (itpr) on Ca2+ signals in cultured neurons derived from Drosophila pupae. Increased spontaneous Ca2+ influx across was observed the PM of isolated pupal neurons with mutant IP3R and also a loss of SOCE. Both spontaneous Ca2+ influx and reduced SOCE were reversed by over-expression of dOrai and dSTIM, which encode the SOCE Ca2+ channel and the ER Ca2+-sensor that regulates it, respectively. Expression of voltage-gated Ca2+ channels (cac, Ca-alpha1D and Ca-alphaT) was significantly reduced in itpr mutant neurons. However, expression of trp mRNAs and transient receptor potential (TRP) protein were increased, suggesting that TRP channels might contribute to the increased spontaneous Ca2+ influx in neurons with mutant IP3R. Thus, IP3R/SOCE modulates spontaneous Ca2+ influx and expression of PM Ca2+ channels in Drosophila pupal neurons. Spontaneous Ca2+ influx compensates for the loss of SOCE in Drosophila itpr mutant neurons.
Ting, C. Y., McQueen, P. G., Pandya, N., McCreedy, E. S., McAuliffe, M. and Lee, C. H. (2017). Analyzing dendritic morphology in columns and layers. J Vis Exp(121) [Epub ahead of print]. PubMed ID: 28362388
In many regions of the central nervous systems, such as the fly optic lobes and the vertebrate cortex, synaptic circuits are organized in layers and columns to facilitate brain wiring during development and information processing in developed animals. Postsynaptic neurons elaborate dendrites in type-specific patterns in specific layers to synapse with appropriate presynaptic terminals. The fly medulla neuropil is composed of 10 layers and about 750 columns; each column is innervated by dendrites of over 38 types of medulla neurons, which match with the axonal terminals of some 7 types of afferents in a type-specific fashion. This report details the procedures to image and analyze dendrites of medulla neurons. The workflow includes three sections: (1) the dual-view imaging section combines two confocal image stacks collected at orthogonal orientations into a high-resolution 3D image of dendrites; (2) the dendrite tracing and registration section traces dendritic arbors in 3D and registers dendritic traces to the reference column array; (3) the dendritic analysis section analyzes dendritic patterns with respect to columns and layers, including layer-specific termination and planar projection direction of dendritic arbors, and derives estimates of dendritic branching and termination frequencies. The protocols utilize custom plugins built on the open-source MIPAV (Medical Imaging Processing, Analysis, and Visualization) platform and custom toolboxes in the matrix laboratory language. Together, these protocols provide a complete workflow to analyze the dendritic routing of Drosophila medulla neurons in layers and columns, to identify cell types, and to determine defects in mutants.
Solari, P., Rivelli, N., De Rose, F., Picciau, L., Murru, L., Stoffolano, J. G., Jr. and Liscia, A. (2017). Opposite effects of 5-HT/AKH and octopamine on the crop contractions in adult Drosophila melanogaster: Evidence of a double brain-gut serotonergic circuitry. PLoS One 12(3): e0174172. PubMed ID: 28334024
In adult Drosophila melanogaster, the type of sugar - either present within the crop lumen or in the bathing solution of the crop - has no effect on crop muscle contraction. What is important, however, is the volume within the crop lumen. Electrophysiological recordings demonstrated that exogenous applications of serotonin on crop muscles increases both the amplitude and the frequency of crop contraction rate, while adipokinetic hormone mainly enhances the crop contraction frequency. Conversely, octopamine virtually silenced the overall crop activity. The present study reports an analysis of serotonin effects along the gut-brain axis in adult D. melanogaster. Injection of serotonin into the brain between the interocellar area shows that brain applications of serotonin decrease the frequency of crop activity. Based on these results, it is proposed that there are two different, opposite pathways for crop motility control governed by serotonin: excitatory when added in the abdomen (i.e., directly bathing the crop) and inhibitory when supplied within the brain (i.e., by injection). The results point to a double brain-gut serotonergic circuitry suggesting that not only the brain can affect gut functions, but the gut can also affect the central nervous system.
Nitta, Y. and Sugie, A. (2017). DISCO interacting protein 2 determines direction of axon projection under the regulation of c-Jun N-terminal kinase in the Drosophila mushroom body. Biochem Biophys Res Commun. PubMed ID: 28396149
Precisely controlled axon guidance for complex neuronal wiring is essential for appropriate neuronal function. c-Jun N-terminal kinase (JNK) was found to play a role in axon guidance recently as well as in cell proliferation, protection and apoptosis. In spite of many genetic and molecular studies on these biological processes regulated by JNK, how JNK regulates axon guidance accurately has not been fully explained thus far. To address this question, this study used the Drosophila mushroom body (MB) as a model since the α/β axons project in two distinct directions. This study showns that DISCO interacting protein 2 (DIP2) is required for the accurate direction of axonal guidance. DIP2 expression is under the regulation of Basket (Bsk), the Drosophila homologue of JNK. The Bsk/DIP2 pathway is independent from the AP-1 transcriptional factor complex pathway, which is directly activated by Bsk. In conclusion, these findings revealed DIP2 as a novel effector downstream of Bsk modulating the direction of axon projection.

Tuesday June 6th

Yao, C. K., Liu, Y. T., Lee, I. C., Wang, Y. T. and Wu, P. Y. (2017). A Ca2+ channel differentially regulates Clathrin-mediated and activity-dependent bulk endocytosis. PLoS Biol 15(4): e2000931. PubMed ID: 28414717
Clathrin-mediated endocytosis (CME) and activity-dependent bulk endocytosis (ADBE) are two predominant forms of synaptic vesicle (SV) endocytosis, elicited by moderate and strong stimuli, respectively. They are tightly coupled with exocytosis for sustained neurotransmission. However, the underlying mechanisms are ill defined. Previous work has shown that the Flower (Fwe) Ca2+ channel present in SVs is incorporated into the periactive zone upon SV fusion, where it triggers CME, thus coupling exocytosis to CME. This study shows that Fwe also promotes ADBE. Intriguingly, the effects of Fwe on CME and ADBE depend on the strength of the stimulus. Upon mild stimulation, Fwe controls CME independently of Ca2+ channeling. However, upon strong stimulation, Fwe triggers a Ca2+ influx that initiates ADBE. Moreover, knockout of rodent fwe in cultured rat hippocampal neurons impairs but does not completely abolish CME, similar to the loss of Drosophila fwe at the neuromuscular junction, suggesting that Fwe plays a regulatory role in regulating CME across species. In addition, the function of Fwe in ADBE is conserved at mammalian central synapses. Hence, Fwe exerts different effects in response to different stimulus strengths to control two major modes of endocytosis.
Castells-Nobau, A., Nijhof, B., Eidhof, I., Wolf, L., Scheffer-de Gooyert, J. M., Monedero, I., Torroja, L., van der Laak, J. and Schenck, A. (2017). Two algorithms for high-throughput and multi-parametric quantification of Drosophila neuromuscular junction morphology. J Vis Exp(123). PubMed ID: 28518121
The Drosophila larval neuromuscular junction (NMJ), a well-established model for glutamatergic synapses, has been extensively studied for decades. Identification of mutations causing NMJ morphological defects revealed a repertoire of genes that regulate synapse development and function. Many of these were identified in large-scale studies that focused on qualitative approaches to detect morphological abnormalities of the Drosophila NMJ. This protocol describes in detail two image analysis algorithms "Drosophila NMJ Morphometrics" and "Drosophila NMJ Bouton Morphometrics", available as Fiji-compatible macros, for quantitative, accurate and objective morphometric analysis of the Drosophila NMJ. This methodology is developed to analyze NMJ terminals immunolabeled with the commonly used markers Dlg-1 and Brp. Additionally, its wider application to other markers such as Hrp, Csp and Syt is presented in this protocol. The macros are able to assess nine morphological NMJ features: NMJ area, NMJ perimeter, number of boutons, NMJ length, NMJ longest branch length, number of islands, number of branches, number of branching points and number of active zones in the NMJ terminal.
Augustin, H., McGourty, K., Steinert, J. R., Cocheme, H. M., Adcott, J., Cabecinha, M., Vincent, A., Halff, E. F., Kittler, J. T., Boucrot, E. and Partridge, L. (2017). Myostatin-like proteins regulate synaptic function and neuronal morphology. Development [Epub ahead of print]. PubMed ID: 28533206
Growth factors of the TGF-beta superfamily play key roles in regulating neuronal and muscle function. Myostatin (or GDF8) and GDF11 are potent negative regulators of skeletal muscle mass. However, expression of both Myostatin and its cognate receptors in other tissues, including brain and peripheral nerves, suggests a potential wider biological role. This study shows that Myoglianin (MYO), the Drosophila homolog of Myostatin and GDF11, regulates not only body weight and muscle size, but also inhibits neuromuscular synapse strength and composition in a Smad2-dependent manner. Both Myostatin and GDF11 affected synapse formation in isolated rat cortical neuron cultures, suggesting an effect on synaptogenesis beyond neuromuscular junctions. This study also shows that Myoglianin acts in vivo to inhibit synaptic transmission between neurons in the escape response neural circuit of adult flies. Thus, these anti-myogenic proteins act as important inhibitors of synapse function and neuronal growth.
Pym, E., Sasidharan, N., Thompson-Peer, K. L., Simon, D. J., Anselmo, A., Sadreyev, R., Hall, Q., Nurrish, S. and Kaplan, J. M. (2017). Shank is a dose-dependent regulator of Cav1 calcium current and CREB target expression. Elife 6. PubMed ID: 28477407 Evolutionary Homolog Study:
Shank (see Drosophila Shank) is a post-synaptic scaffolding protein that has many binding partners. Shank mutations and copy number variations (CNVs) are linked to several psychiatric disorders, and to synaptic and behavioral defects in mice. It is not known which Shank binding partners are responsible for these defects. This study showed that the C. elegans SHN-1/Shank binds L-type calcium channels and that increased and decreased shn-1 gene dosage alter L-channel current and activity-induced expression of a CRH-1/CREB transcriptional target (gem-4 Copine), which parallels the effects of human Shank copy number variations (CNVs) on Autism spectrum disorders and schizophrenia. These results suggest that an important function of Shank proteins is to regulate L-channel current and activity induced gene expression.

Monday, June 5th

Voolstra, O., Rhodes-Mordov, E., Katz, B., Bartels, J. P., Oberegelsbacher, C., Schotthofer, S. K., Yasin, B., Tzadok, H., Huber, A. and Minke, B. (2017). The phosphorylation state of the Drosophila TRP channel modulates the frequency response to oscillating light in vivo. J Neurosci 37(15): 4213-4224. PubMed ID: 28314815
Drosophila photoreceptors respond to oscillating light of high frequency (approximately 100 Hz), while the detected maximal frequency is modulated by the light rearing conditions, thus enabling high sensitivity to light and high temporal resolution. However, the molecular basis for this adaptive process is unclear. This study reports that dephosphorylation of the light-activated transient receptor potential (TRP) ion channel at S936 is a fast, graded, light-dependent, and Ca2+-dependent process that is partially modulated by the rhodopsin phosphatase retinal degeneration C (RDGC). Electroretinogram measurements of the frequency response to oscillating lights in vivo revealed that dark-reared flies expressing wild-type TRP exhibited a detection limit of oscillating light at relatively low frequencies, which was shifted to higher frequencies upon light adaptation. Strikingly, preventing phosphorylation of the S936-TRP site by alanine substitution in transgenic Drosophila (trpS936A) abolished the difference in frequency response between dark-adapted and light-adapted flies, resulting in high-frequency response also in dark-adapted flies. In contrast, inserting a phosphomimetic mutation by substituting the S936-TRP site to aspartic acid (trpS936D) set the frequency response of light-adapted flies to low frequencies typical of dark-adapted flies. Light-adapted rdgC mutant flies showed relatively high S936-TRP phosphorylation levels and light-dark phosphorylation dynamics. These findings suggest that RDGC is one but not the only phosphatase involved in pS936-TRP dephosphorylation. Together, this study indicates that TRP channel dephosphorylation is a regulatory process that affects the detection limit of oscillating light according to the light rearing condition, thus adjusting dynamic processing of visual information under varying light conditions.
Taylor, E., Alqadri, N., Dodgson, L., Mason, D., Lyulcheva, E., Messina, G. and Bennett, D. (2017). MRL proteins cooperate with activated Ras in glia to drive distinct oncogenic outcomes. Oncogene [Epub ahead of print]. PubMed ID: 28346426
The Mig10/RIAM/Lpd (MRL) adapter protein Lpd (see Drosophila Pico) regulates actin dynamics through interactions with Scar/WAVE and Ena/VASP proteins to promote the formation of cellular protrusions and to stimulate invasive migration. However, the ability of MRL proteins to interact with multiple actin regulators and to promote serum response factor (SRF) signalling has raised the question of whether MRL proteins employ alternative downstream mechanisms to drive oncogenic processes in a context-dependent manner. Using a Drosophila model, overexpression of either human Lpd or its Drosophila orthologue Pico can promote growth and invasion of RasV12-induced cell tumours in the brain. Notably, effects were restricted to two populations of Repo-positive glial cells: an invasive population, characterized by JNK-dependent elevation of Mmp1 expression, and a hyperproliferative population lacking elevated JNK signalling. JNK activation was not triggered by reactive immune cell signalling, implicating the involvement of an intrinsic stress response. The ability to promote dissemination of RasV12-induced tumours was shared by a subset of actin regulators, including, most prominently, Chicadee/Profilin, which directly interacts with Pico, and, Mal, a cofactor for serum response factor that responds to changes in G:F actin dynamics. Suppression of Mal activity partially abrogated the ability of pico to promote invasion of RasV12 tumours. Furthermore, it was found that larval glia are enriched for serum response factor expression, explaining the apparent sensitivity of glial cells to Pico/RasV12 overexpression. Taken together, these findings indicate that MRL proteins cooperate with oncogenic Ras to promote formation of glial tumours, and that, in this context, Mal/serum response factor activation is rate-limiting for tumour dissemination.
Tang, Y., Geng, Q., Chen, D., Zhao, S., Liu, X. and Wang, Z. (2017). Germline proliferation is regulated by somatic endocytic genes via JNK and BMP signaling in Drosophila. Genetics [Epub ahead of print]. PubMed ID: 28315838
Signals derived from the microenvironment contribute greatly to tumorigenesis. This study used Drosophila testis as a model system to address this question, taking the advantage of the ease to distinguish germline and somatic cells and to track the cell numbers. In an EMS mutagenesis screen, Rab5, a key factor in endocytosis, was identified for its non-autonomous role in germline proliferation. The disruption of Rab5 in somatic cyst cells, which escort the development of germline lineage, induced the over-proliferation of under-differentiated but genetically wild-type germ cells. This non-autonomous effect was mediated by the transcriptional activation of Dpp (the fly homolog of BMP) by examining the Dpp-reporter expression and knocking down Dpp to block germline overgrowth. Consistently, the protein levels of Bam, the germline pro-differentiation factor normally accumulated in the absence of BMP/Dpp signaling, decreased in the over-proliferating germ cells. Further, it was discovered that JNK signaling pathway operated between Rab5 and Dpp, because simultaneously inhibiting JNK pathway and Rab5 in cyst cells prevented both dpp transcription and germline tumor growth. Additionally, it was found that multiple endocytic genes, such as avl, TSG101, Vps25, or Cdc42, were required in the somatic cyst cells to restrict germline amplification. These findings indicate that when the endocytic state of the surrounding cells are impaired, genetically wild-type germ cells overgrow. This non-autonomous model of tumorigenesis provides a simple system to dissect the relation between tumor and its niche.
Bulgakova, N. A., Wellmann, J. and Brown, N. H. (2017). Diverse integrin adhesion stoichiometries caused by varied actomyosin activity. Open Biol 7(4). PubMed ID: 28446705
Cells in an organism are subjected to numerous sources of external and internal forces, and are able to sense and respond to these forces. Integrin-mediated adhesion links the extracellular matrix outside cells to the cytoskeleton inside, and participates in sensing, transmitting and responding to forces. While integrin adhesion rapidly adapts to changes in forces in isolated migrating cells, it is not known whether similar or more complex responses occur within intact, developing tissues. Changes in integrin adhesion composition were studied upon different contractility conditions in Drosophila embryonic muscles. All integrin adhesion components tested were still present at muscle attachment sites (MASs) when either cytoplasmic or muscle myosin II was genetically removed, suggesting a primary role of a developmental programme in the initial assembly of integrin adhesions. Contractility does, however, increase the levels of integrin adhesion components, suggesting a mechanism to balance the strength of muscle attachment to the force of muscle contraction. Perturbing contractility in distinct ways, by genetic removal of either cytoplasmic or muscle myosin II or eliminating muscle innervation, each caused unique alterations to the stoichiometry at MASs. This suggests that different integrin-associated proteins are added to counteract different kinds of force increase.

Sunday, June 4th

Bloch Qazi, M. C., Miller, P. B., Poeschel, P. M., Phan, M. H., Thayer, J. L. and Medrano, C. L. (2017). Transgenerational effects of maternal and grandmaternal age on offspring viability and performance in Drosophila melanogaster. J Insect Physiol 100: 43-52. PubMed ID: 28529156
In non-social insects, fitness is determined by relative lifetime fertility. Fertility generally declines with age as a part of senescence. For females, senescence has profound effects on fitness by decreasing viability and fertility as well as those of her offspring. However, important aspects of these maternal effects, including the cause(s) of reduced offspring performance and carry-over effects of maternal age, are poorly understood. Drosophila melanogaster is a useful system for examining potential transgenerational effects of increasing maternal age, because of their use as a model system for studying the physiology and genetic architecture of both reproduction and senescence. To test the hypothesis that female senescence has transgenerational effects on offspring viability and development, this study measured the effects of maternal age on offspring survival over two generations and under two larval densities in two laboratory strains of flies (Oregon-R and Canton-S). Transgenerational effects of maternal age influence embryonic viability and embryonic to adult viability in both strains. However, the generation causing the effects, and the magnitude and direction of those effects differed by genotype. The effects of maternal age on embryonic-to-adult viability when larvae are stressed was also genotype-specific. Maternal effects involve provisioning: older females produced smaller eggs and larger offspring. These results show that maternal age has profound, complex, and multigenerational consequences on several components of offspring fitness and traits. This study contributes to a body of work demonstrating that female age is an important condition affecting phenotypic variation and viability across multiple generations.
Blenau, W., Daniel, S., Balfanz, S., Thamm, M. and Baumann, A. (2017). Dm5-HT2B: Pharmacological characterization of the fifth serotonin receptor subtype of Drosophila melanogaster. Front Syst Neurosci 11: 28. PubMed ID: 28553207
Serotonin (5-hydroxytryptamine, 5-HT) is an important regulator of physiological and behavioral processes in both protostomes (e.g., insects) and deuterostomes (e.g., mammals). In insects, serotonin has been found to modulate the heart rate and to control secretory processes, development, circadian rhythms, aggressive behavior, as well as to contribute to learning and memory. Serotonin exerts its activity by binding to and activating specific membrane receptors. The clear majority of these receptors belong to the superfamily of G-protein-coupled receptors. In Drosophila melanogaster, a total of five genes have been identified coding for 5-HT receptors. From this family of proteins, four have been pharmacologically examined in greater detail, so far. While Dm5-HT1A, Dm5-HT1B, and Dm5-HT7 couple to cAMP signaling cascades, the Dm5-HT2A receptor leads to Ca2+ signaling in an inositol-1,4,5-trisphosphate-dependent manner. Based on sequence similarity to homologous genes in other insects, a fifth D. melanogaster gene was uncovered coding for a Dm5-HT2B receptor. Knowledge about this receptor's pharmacological properties is very limited. This is quite surprising because Dm5-HT2B has been attributed to distinct physiological functions based on genetic interference with its gene expression. Mutations were described reducing the response of the larval heart to 5-HT, and specific knockdown of Dm5-HT2B mRNA in hemocytes resulted in a higher susceptibility of the flies to bacterial infection. To gain deeper understanding of Dm5-HT2B's pharmacology, this study evaluated the receptor's response to a series of established 5-HT receptor agonists and antagonists in a functional cell-based assay. Metoclopramide and mianserin were identified as two potent antagonists that may allow pharmacological interference with Dm5-HT2B signaling in vitro and in vivo.
Pomatto, L. C., Wong, S., Carney, C., Shen, B., Tower, J. and Davies, K. J. (2017). The age- and sex-specific decline of the 20s proteasome and the Nrf2/CncC signal transduction pathway in adaption and resistance to oxidative stress in Drosophila melanogaster. Aging (Albany NY). PubMed ID: 28373600
Hallmarks of aging include loss of protein homeostasis and dysregulation of stress-adaptive pathways. Loss of adaptive homeostasis, increases accumulation of DNA, protein, and lipid damage. During acute stress, the Cnc-C (Drosophila Nrf2 orthologue) transcriptionally-regulated 20S proteasome degrades damaged proteins in an ATP-independent manner. Exposure to very low, non-toxic, signaling concentrations of the redox-signaling agent hydrogen peroxide (H2O2) cause adaptive increases in the de novo expression and proteolytic activity/capacity of the 20S proteasome in female flies. Female 20S proteasome induction was accompanied by increased tolerance to a subsequent normally toxic but sub-lethal amount of H2O2, and blocking adaptive increases in proteasome expression also prevented full adaptation. This adaptive response is both sex- and age-dependent. Both increased proteasome expression and activity, and increased oxidative-stress resistance, in female flies, were lost with age. In contrast, male flies exhibited no H2O2 adaptation, irrespective of age. Furthermore, aging caused a generalized increase in basal 20S proteasome expression, but proteolytic activity and adaptation were both compromised. Finally, continual knockdown of Keap1 (the cytosolic inhibitor of Cnc-C) in adults resulted in older flies with greater stress resistance than their age-matched controls, but who still exhibited an age-associated loss of adaptive homeostasis.
Brown, M. K., Strus, E. and Naidoo, N. (2017). Reduced sleep during social isolation leads to cellular stress and induction of the Unfolded Protein Response (UPR). Sleep [Epub ahead of print]. PubMed ID: 28541519
Social isolation has a multitude of negative consequences on human health including the ability to endure challenges to the immune system, sleep amount and efficiency, and general morbidity and mortality. These adverse health outcomes are conserved in other social species. In the fruit fly Drosophila melanogaster, social isolation leads to increased aggression, impaired memory and reduced amounts of daytime sleep. There is a correlation between molecules affected by social isolation and those implicated in sleep in Drosophila. Previous work has demonstrated that acute sleep loss in flies and mice induced the unfolded protein response (UPR), an adaptive signaling pathway. One mechanism indicating UPR upregulation is elevated levels of the endoplasmic reticular chaperone BiP/GRP78. BiP overexpression in Drosophila has been shown to led to increased sleep rebound. Increased rebound sleep has also been demonstrated in socially isolated flies. Flies were used to study the effect of social isolation on cellular stress. Socially isolated flies displayed an increase in UPR markers; there were higher BiP levels, increased phosphorylation of the translation initiation factor eIF2alpha and increased splicing of xbp1. These are all indicators of UPR activation. In addition, the effects of isolation on the UPR were reversible; pharmacologically and genetically altering sleep in the flies modulated the UPR. It is concluded that the reduction in sleep observed in socially isolated flies is a cellular stressor that results in UPR induction.

Saturday, June 3rd

Zanon, A., et al. (2017). SLP-2 interacts with Parkin in mitochondria and prevents mitochondrial dysfunction in Parkin-deficient human iPSC-derived neurons and Drosophila. Hum Mol Genet. PubMed ID: 28379402
Mutations in the Parkin gene (PARK2; see Drosophila Parkin) have been linked to a recessive form of Parkinson's disease (PD) characterized by the loss of dopaminergic neurons in the substantia nigra. Deficiencies of mitochondrial respiratory chain complex I activity have been observed in the substantia nigra of PD patients, and loss of Parkin results in the reduction of complex I activity shown in various cell and animal models. Using co-immunoprecipitation and proximity ligation assays on endogenous proteins, this study demonstrates that Parkin interacts with mitochondrial Stomatin-like protein 2 (SLP-2), which also binds the mitochondrial lipid cardiolipin and functions in the assembly of respiratory chain proteins. SH-SY5Y cells with a stable knockdown of Parkin or SLP-2, as well as induced pluripotent stem cell-derived neurons from Parkin mutation carriers, showed decreased complex I activity and altered mitochondrial network morphology. Importantly, induced expression of SLP-2 corrected for these mitochondrial alterations caused by reduced Parkin function in these cells. In-vivo Drosophila studies showed a genetic interaction of Parkin and SLP-2, and further, tissue-specific or global overexpression of SLP-2 transgenes rescued parkin mutant phenotypes, in particular loss of dopaminergic neurons, mitochondrial network structure, reduced ATP production, and flight and motor dysfunction. The physical and genetic interaction between Parkin and SLP-2 and the compensatory potential of SLP-2 suggest a functional epistatic relationship to Parkin and a protective role of SLP-2 in neurons. This finding places further emphasis on the significance of Parkin for the maintenance of mitochondrial function in neurons and provides a novel target for therapeutic strategies.
Arefin, B., Kunc, M., Krautz, R. and Theopold, U. (2017). The immune phenotype of three Drosophila leukemia models. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 28476910
Many leukemia patients suffer from dysregulation of their immune system, making them more susceptible to infections and leading to general weakening (cachexia). Both adaptive and innate immunity are affected. The fruitfly Drosophila melanogaster has an innate immune system including cells of the myeloid lineage (hemocytes). To study Drosophila immunity and physiology during leukemia, three models were established by driving expression of a dominant-active version of the Ras oncogene (RasV12 ) alone or combined with knockdowns of tumor suppressors in Drosophila hemocytes. The results show that phagocytosis, hemocytes migration to wound sites, wound sealing and survival upon bacterial infection of leukemic lines are similar to wild type. In all leukemic models the two major immune pathways (Toll and Imd) are dysregulated. Toll-dependent signaling is activated to comparable extents as after wounding wild type larvae, leading to a proinflammatory status. In contrast, Imd signaling is suppressed. Finally, adult tissue formation was blocked, and degradation was observed of cell masses during metamorphosis of leukemic lines, which is akin to the state of cancer-dependent cachexia. To further analyze the immune competence of leukemic linesa natural infection model was used that involves insect-pathogenic nematodes. Two leukemic lines, which were sensitive to nematode infections, were identified. Further characterization demonstrates that despite the absence of behavioral abnormalities at the larval stage, leukemic larvae show reduced locomotion in the presence of nematodes. Taken together this work establishes new Drosophila models to study the physiological- immune- and behavioral consequences of various forms of leukemia.
Zhang, T., Mishra, P., Hay, B. A., Chan, D. and Guo, M. (2017). Valosin-containing protein (VCP/p97) inhibitors relieve Mitofusin-dependent mitochondrial defects due to VCP disease mutants. Elife 6. PubMed ID: 28322724
Missense mutations of valosin-containing protein (VCP; see Drosophila TER94 ) cause an autosomal dominant disease known as inclusion body myopathy, Paget disease with frontotemporal dementia (IBMPFD) and other neurodegenerative disorders. The pathological mechanism of IBMPFD is not clear and there is no treatment. This study shows that endogenous VCP negatively regulates Mitofusin (see Drosophila Marf), which is required for outer mitochondrial membrane fusion. Because 90% of IBMPFD patients have myopathy, an in vivo IBMPFD model was generated in adult Drosophila muscle, which recapitulates disease pathologies. Common VCP disease mutants were shown to act as hyperactive alleles with respect to regulation of Mitofusin. Importantly, VCP inhibitors suppress mitochondrial defects, muscle tissue damage and cell death associated with IBMPFD models in Drosophila. These inhibitors also suppress mitochondrial fusion and respiratory defects in IBMPFD patient fibroblasts. These results suggest that VCP disease mutants cause IBMPFD through a gain-of-function mechanism, and that VCP inhibitors have therapeutic value.
Zwarts, L., Vulsteke, V., Buhl, E., Hodge, J. J. and Callaerts, P. (2017). SlgA, the homologue of the human schizophrenia associated PRODH gene, acts in clock neurons to regulate Drosophila aggression. Dis Model Mech. PubMed ID: 28331058
Mutations in proline dehydrogenase (PRODH) are linked to behavioral alterations in schizophrenia and as part of DiGeorge and velo-cardio-facial syndromes, but the role of PRODH in their etiology remains unclear. This study established a Drosophila model to study the role of PRODH in behavioral disorders. The distribution was determined of the Drosophila PRODH homolog slgA in the brain, and knock-down and overexpression of human PRODH and slgA in the lateral neurons ventral (LNv) were shown to lead to altered aggressive behavior. SlgA acts in an isoform-specific manner and is regulated by casein kinase II (CkII). The data suggest that these effects are, at least partially, due to effects on mitochondrial function. It is thus shown that precise regulation of proline metabolism is essential to drive normal behavior and Drosophila aggression is a model behavior relevant for the study of mechanisms impaired in neuropsychiatric disorders.

Friday, June 2nd

Mallik, B., Dwivedi, M.K., Mushtaq, Z., Kumari, M., Verma, P.K. and Kumar, V. (2017). Regulation of neuromuscular junction organization by Rab2 and its effector ICA69 in Drosophila. Development [Epub ahead of print]. PubMed ID: 28455372
Mechanisms underlying synaptic differentiation, which involves neuronal membrane and cytoskeletal remodeling, are not completely understood. This study performed a targeted RNAi-mediated screen of Drosophila BAR-domain proteins and identified islet cell autoantigen 69 kDa (dICA69) as one of the key regulators of morphological differentiation of larval neuromuscular junction (NMJ). Drosophila ICA69 colocalizes with α-Spectrin at the NMJ. The conserved N-BAR domain of dICA69 deforms liposomes in vitro. Full length and ICAC but not the N-BAR domain of dICA69 which induces filopodia in cultured cells. Consistent with its cytoskeleton regulatory role, dICA69 mutants show reduced α-Spectrin immunoreactivity at the larval NMJ. Manipulating levels of dICA69 or its interactor dPICK1 alters synaptic level of ionotropic glutamate receptors (iGluRs). Moreover, reducing dPICK1 or dRab2 levels phenocopies dICA69 mutation. Interestingly, dRab2 regulates not only synaptic iGluR but also dICA69 levels. Thus, these data suggest that: a) dICA69 regulates NMJ organization through a pathway that involves dPICK1 and dRab2, and b) dRab2 genetically functions upstream of dICA69 and regulates NMJ organization and targeting/retention of iGluRs by regulating dICA69 levels.

Spencer, A. K., Schaumberg, A. J. and Zallen, J. A. (2017). Scaling of cytoskeletal organization with cell size in Drosophila. Mol Biol Cell 28(11):1519-1529. PubMed ID: 28404752
Spatially organized macromolecular complexes are essential for cell and tissue function, but the mechanisms that organize micron-scale structures within cells are not well understood. Microtubule-based structures such as mitotic spindles scale with cell size, but less is known about the scaling of actin structures within cells. Actin-rich denticle precursors cover the ventral surface of the Drosophila embryo and larva and provide templates for cuticular structures involved in larval locomotion. Using quantitative imaging and statistical modeling, denticle number and spacing were demonstrated to scale with cell size over a wide range of cell lengths in embryos and larvae. Denticle number and spacing are reduced under space-limited conditions, and both features robustly scale over a ten-fold increase in cell length during larval growth. The relationship between cell length and denticle spacing can be recapitulated by specific mathematical equations in embryos and larvae, and accurate denticle spacing requires an intact microtubule network and the microtubule minus-end-binding protein, Patronin. These results identify a novel mechanism of microtubule-dependent actin scaling that maintains precise patterns of actin organization during tissue growth.
Krieg, M., Stühmer, J., Cueva, J.G., Fetter, R., Spilker, K., Cremers, D., Shen, K., Dunn, A.R. and Goodman, M.B. (2017). Genetic defects in β-spectrin and tau sensitize C. elegans axons to movement-induced damage via torque-tension coupling. Elife 6. PubMed ID: 28098556
Evolutionary Homolog Study:
Our bodies are in constant motion and so are the neurons that invade each tissue. Motion-induced neuron deformation and damage are associated with several neurodegenerative conditions. This study investigated the question of how the neuronal cytoskeleton protects axons and dendrites from mechanical stress, exploiting mutations in UNC-70 (see Drosophila karst), PTL-1 (see Drosophila tau) and MEC-7 (see Drosophila β-tubulin) proteins in Caenorhabditis elegans. It was found that mechanical stress induces supercoils and plectonemes in the sensory axons of spectrin and tau double mutants. Biophysical measurements, super-resolution, and electron microscopy, as well as numerical simulations of neurons as discrete, elastic rods provide evidence that a balance of torque, tension, and elasticity stabilizes neurons against mechanical deformation. The study concludes that the spectrin and microtubule cytoskeletons work in combination to protect axons and dendrites from mechanical stress and propose that defects in β-spectrin and tau may sensitize neurons to damage.

An, Y., Xue, G., Shaobo, Y., Mingxi, D., Zhou, X., Yu, W., Ishibashi, T., Zhang, L. and Yan, Y. (2017). Apical constriction is driven by a pulsatile apical myosin network in delaminating Drosophila neuroblasts. Development [Epub ahead of print]. PubMed ID: 28506995
Cell delamination is a conserved morphogenetic process important for generation of cell diversity and maintenance of tissue homeostasis. This study used Drosophila embryonic neuroblasts as a model to study the apical constriction process during cell delamination. Dynamic myosin signals were observed both around the cell adherens junctions and underneath the cell apical surface in the neuroectoderm. On the cell apical cortex the non-junctional myosin forms flows and pulses, which are termed 'medial myosin pulses'. Quantitative differences in medial myosin pulse intensity and frequency are critical to distinguish delaminating neuroblasts from their neighbors. Inhibition of medial myosin pulses blocks delamination. The fate of neuroblasts is set apart from their neighbors by Notch signaling-mediated lateral inhibition. When Notch signaling activity is inhibited in the embryo, small clusters of cells are observed to undergo apical constriction and display an abnormal apical myosin pattern. Together, it was demonstrated that a contractile actomyosin network across the apical cell surface is organized to drive apical constriction in delaminating neuroblasts.

Thursday, June 1st

Umegawachi, T., Yoshida, H., Koshida, H., Yamada, M., Ohkawa, Y., Sato, T., Suyama, M., Krause, H. M. and Yamaguchi, M. (2017). Control of tissue size and development by a regulatory element in the yorkie 3'UTR. Am J Cancer Res 7(3): 673-687. PubMed ID: 28401020
Regulation of the Hippo pathway via phosphorylation of Yorkie (Yki), the Drosophila homolog of human Yes-associated protein 1, is conserved from Drosophila to humans. Overexpression of a non-phosphorylatable form of Yki induces severe overgrowth in adult fly eyes. This study shows that yki mRNA associates with microsomal fractions and forms foci that partially colocalize to processing bodies in the vicinity of endoplasmic reticulum. This localization is dependent on a stem-loop (SL) structure in the 3' untranslated region of yki. Surprisingly, expression of SL deleted yki in eye imaginal discs also results in severe overgrowth phenotypes. When the structure of the SL is disrupted, Yki protein levels increase without a significant effect on RNA levels. When the SL is completely removed, protein levels drastically increase, but in this case, due to increased RNA stability. In the latter case, it was shown that the increased RNA accumulation is due to removal of a putative miR-8 seed sequence in the SL. These data demonstrate the function of two novel regulatory mechanisms, both controlled by the yki SL element, that are essential for proper Hippo pathway mediated growth regulation.
Vallejos Baier, R., Picao-Osorio, J. and Alonso, C. R. (2017). Molecular Regulation of Alternative Polyadenylation (APA) within the Drosophila Nervous System. J Mol Biol [Epub ahead of print]. PubMed ID: 28366829
Alternative polyadenylation (APA) is a widespread gene regulatory mechanism that generates mRNAs with different 3'-ends, allowing them to interact with different sets of RNA regulators such as microRNAs and RNA-binding proteins. This study investigated this problem within the Drosophila nervous system, focusing on the roles played by general cleavage and polyadenylation factors (CPA factors). The expression of the Drosophila orthologues of all mammalian CPA factors (see for example Suppressor of forked, Cpsf160, CstF64 and Hiiragi) was analyzed, and it was noted that their expression decreases during embryogenesis. In contrast to this global developmental decrease in CPA factor expression, cleavage factor I (CFI) expression is actually elevated in the late embryonic central nervous system, suggesting that CFI might play a special role in neural tissues. To test this, the UAS/Gal4 system was used to deplete CFI proteins from neural tissue and it was observed that in this condition, multiple genes switch their APA patterns, demonstrating a role of CFI in APA control during Drosophila neural development. Furthermore, analysis of genes with 3'UTR extensions of different length leads the authors to a suggest a novel relation between 3'UTR length and sensitivity to CPA factor expression. This work work thus contributes to the understanding of the mechanisms of APA control within the developing central nervous system.
Sharma, A., Heinze, S. D., Wu, Y., Kohlbrenner, T., Morilla, I., Brunner, C., Wimmer, E. A., van de Zande, L., Robinson, M. D., Beukeboom, L. W. and Bopp, D. (2017). Male sex in houseflies is determined by Mdmd, a paralog of the generic splice factor gene CWC22. Science 356(6338): 642-645. PubMed ID: 28495751 Evolutionary Homolog Study:
Across species, animals have diverse sex determination pathways, each consisting of a hierarchical cascade of genes and its associated regulatory mechanism. Houseflies have a distinctive polymorphic sex determination system in which a dominant male determiner, the M-factor, can reside on any of the chromosomes. This study identified a gene, Musca domestica male determiner (Mdmd), as the M-factor. Mdmd originated from a duplication of the spliceosomal factor gene CWC22 (nucampholin). Targeted Mdmd disruption results in complete sex reversal to fertile females because of a shift from male to female expression of the downstream genes transformer and doublesex The presence of Mdmd on different chromosomes indicates that Mdmd translocated to different genomic sites. Thus, an instructive signal in sex determination can arise by duplication and neofunctionalization of an essential splicing regulator.
Tamayo, J. V., Teramoto, T., Chatterjee, S., Hall, T. M. and Gavis, E. R. (2017). The Drosophila hnRNP F/H homolog Glorund uses two distinct RNA-binding modes to diversify target recognition. Cell Rep 19(1): 150-161. PubMed ID: 28380354
The Drosophila hnRNP F/H homolog, Glorund (Glo), regulates nanos mRNA translation by interacting with a structured UA-rich motif in the nanos 3' untranslated region. Glo regulates additional RNAs, however, and mammalian homologs bind G-tract sequences to regulate alternative splicing, suggesting that Glo also recognizes G-tract RNA. To gain insight into how Glo recognizes both structured UA-rich and G-tract RNAs, mutational analysis was used guided by crystal structures of Glo's RNA-binding domains, and two discrete RNA-binding surfaces were identified that allow Glo to recognize both RNA motifs. By engineering Glo variants that favor a single RNA-binding mode, it was shown that a subset of Glo's functions in vivo is mediated solely by the G-tract binding mode, whereas regulation of nanos requires both recognition modes. These findings suggest a molecular mechanism for the evolution of dual RNA motif recognition in Glo that may be applied to understanding the functional diversity of other RNA-binding proteins.
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