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September 22nd, 2023 - Adult Physiology and Metabolism

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Collins, D. H., Prince, D. C., Donelan, J. L., Chapman, T. and Bourke, A. F. G. (2023). Developmental diet alters the fecundity-longevity relationship and age-related gene expression in Drosophila melanogaster. J Gerontol A Biol Sci Med Sci. PubMed ID: 37584665
Summary:
The standard evolutionary theory of aging predicts a negative relationship (trade-off) between fecundity and longevity. However, in principle, the fecundity-longevity relationship can become positive in populations in which individuals have unequal resources. Positive fecundity-longevity relationships also occur in queens of eusocial insects such as ants and bees. Developmental diet is likely to be central to determining trade-offs as it affects key fitness traits, but its exact role remains uncertain. For example, in Drosophila melanogaster, changes in adult diet can affect fecundity, longevity, and gene expression throughout life, but it is unknown how changes in developmental (larval) diet affect fecundity-longevity relationships and gene expression in adults. Using D. melanogaster, this study tested the hypothesis that varying developmental diet alters the directionality of fecundity-longevity relationships in adults, and characterised associated gene expression changes. This study reared larvae on low (20%), medium (100%), and high (120%) yeast diets, and transferred adult females to a common diet. Fecundity and longevity of individual adult females were measure, and gene expression changes with age was profiled. Adult females raised on different larval diets exhibited fecundity-longevity relationships that varied from significantly positive to significantly negative, despite minimal differences in mean life-time fertility or longevity. Treatments also differed in age-related gene expression, including for aging-related genes. Hence the sign of fecundity-longevity relationships in adult insects can be altered and even reversed by changes in larval diet quality. By extension, larval diet differences may represent a key mechanistic factor underpinning positive fecundity-longevity relationships observed in species such as eusocial insects.
Cavigliasso, F., Savary, L., Spangenberg, J. E., Gallart-Ayala, H., Ivanisevic, J. and Kawecki, T. J. (2023). Experimental evolution of metabolism under nutrient restriction: enhanced amino acid catabolism and a key role of branched-chain amino acids. Evol Lett 7(4): 273-284. PubMed ID: 37475747
Summary:
Periodic food shortage is a common ecological stressor for animals, likely to drive physiological and metabolic adaptations to alleviate its consequences, particularly for juveniles that have no option but to continue to grow and develop despite undernutrition. This study examined changes in metabolism associated with adaptation to nutrient shortage, evolved by replicate Drosophila melanogaster populations maintained on a nutrient-poor larval diet for over 240 generations. In a factorial metabolomics experiment it was shown that both phenotypic plasticity and genetically-based adaptation to the poor diet involved wide-ranging changes in metabolite abundance; however, the plastic response did not predict the evolutionary change. Compared to nonadapted larvae exposed to the poor diet for the first time, the adapted larvae showed lower levels of multiple free amino acids in their tissues-and yet they grew faster. By quantifying accumulation of the nitrogen stable isotope (15)N it was shown that adaptation to the poor diet led to an increased use of amino acids for energy generation. This apparent "waste" of scarce amino acids likely results from the trade-off between acquisition of dietary amino acids and carbohydrates observed in these populations. The three branched-chain amino acids (leucine, isoleucine, and valine) showed a unique pattern of depletion in adapted larvae raised on the poor diet. A diet supplementation experiment demonstrated that these amino acids are limiting for growth on the poor diet, suggesting that their low levels resulted from their expeditious use for protein synthesis. These results demonstrate that selection driven by nutrient shortage not only promotes improved acquisition of limiting nutrients, but also has wide-ranging effects on how the nutrients are used. They also show that the abundance of free amino acids in the tissues does not, in general, reflect the nutritional condition and growth potential of an animal.
Chen, W., Yin, Y. and Zhang, Z. (2023). Effects of N-acetylcysteine on CG8005 gene-mediated proliferation and apoptosis of Drosophila S2 embryonic cells. Sci Rep 13(1): 12502. PubMed ID: 37532734
Summary:
To investigate the effect of the antioxidant N-acetylcysteine (NAC) on the proliferation and apoptosis in CG8005 (Deoxyhypusine synthase) gene-interfering Drosophila S2 embryonic cells by scavenging intracellular reactive oxygen species (ROS). The interfering efficiency of CG8005 gene in Drosophila S2 embryonic cells was verified by real-time quantitative PCR (qRT-PCR). Different concentrations of NAC and phosphate buffered saline (PBS) were used to affect the Drosophila S2 embryonic cells. The growth state of Drosophila S2 embryonic cells was observed by light microscope. Two probes dihydroethidium (DHE) and 2,7-dichlorodihydrofluorescein-acetoacetate (DCFH-DA) were used to observe the ROS production in each group after immunofluorescence staining. TUNEL staining and flow cytometry were used to investigate the apoptosis level of Drosophila S2 embryos, and CCK-8 (Cell Counting Kit-8) was used to detect the cell viability of Drosophila S2 embryos. The knockdown efficiency of siCG8005-2 fragment was high and stable, which was verified by interference efficiency (P < 0.05). There was no significant change in the growth of Drosophila S2 embryonic cells after the treatment of NAC as compared to PBS group. Moreover, knockdowning CG8005 gene resulted in an increase in ROS and apoptosis in Drosophila S2 embryonic cells (P &tlt; 0.05) and a decrease in proliferation activity (P < 0.05). In addition, the pretreatment of antioxidant NAC could inhibit ROS production in Drosophila S2 embryonic cells (P < 0.05), reduce cell apoptosis (P < 0.05), and improve cell survival (P < 0.05). The CG8005 gene in Drosophila S2 embryonic cells could regulate the proliferation and apoptosis of S2 embryonic cells by disrupting the redox homeostasis, and antioxidant NAC could inhibit cell apoptosis and promotes cell proliferation by scavenging ROS in Drosophila S2 embryonic cells, which is expected to provide novel insights for the pathogenesis of male infertility and spermatogenesis.
da Silva Soares, N. F., Quagliariello, A., Yigitturk, S. and Martino, M. E. (2023). Gut microbes predominantly act as living beneficial partners rather than raw nutrients. Sci Rep 13(1): 11981. PubMed ID: 37488173
Summary:
Animals and their gut microbes mutually benefit their health. Nutrition plays a central role in this, directly influencing both host and microbial fitness and the nature of their interactions. This makes nutritional symbioses a complex and dynamic tri-system of diet-microbiota-host. Despite recent discoveries on this field, full control over the interplay among these partners is challenging and hinders the resolution of fundamental questions, such as how to parse the gut microbes' effect as raw nutrition or as symbiotic partners? To tackle this, use was made of the well-characterized Drosophila melanogaster/Lactiplantibacillus plantarum experimental model of nutritional symbiosis to generate a quantitative framework of gut microbes' effect on the host. By coupling experimental assays and Random Forest analysis, it was shown that the beneficial effect of L. plantarum strains primarily results from the active relationship as symbionts rather than raw nutrients, regardless of the bacterial strain. Metabolomic analysis of both active and inactive bacterial cells further demonstrated the crucial role of the production of beneficial bacterial metabolites, such as N-acetylated-amino-acids, as result of active bacterial growth and function. Altogether, these results provide a ranking and quantification of the main bacterial features contributing to sustain animal growth.This study has demonstrated that bacterial activity is the predominant and necessary variable involved in bacteria-mediated benefit, followed by strain-specific properties and the nutritional potential of the bacterial cells. This contributes to elucidate the role of beneficial bacteria and probiotics, creating a broad quantitative framework for host-gut microbiome that can be expanded to other model systems.
Dahleh, M. M. M., Araujo, S. M., Bortolotto, V. C., Torres, S. P., Machado, F. R., Meichtry, L. B., Musachio, E. A. S., Guerra, G. P. and Prigol, M. (2023).. The implications of exercise in Drosophila melanogaster: insights into Akt/p38 MAPK/Nrf2 pathway associated with Hsp70 regulation in redox balance maintenance. J Comp Physiol B. PubMed ID: 37500966
Summary:
This study investigated the potential effects of exercise on the responses of energy metabolism, redox balance maintenance, and apoptosis regulation in Drosophila melanogaster to shed more light on the mechanisms underlying the increased performance that this emerging exercise model provides. Three groups were evaluated for seven days: the control (no exercise or locomotor limitations), movement-limited flies (MLF) (no exercise, with locomotor limitations), and EXE (with exercise, no locomotor limitations). The EXE flies demonstrated greater endurance-like tolerance in the swimming test, associated with increased citrate synthase activity, lactate dehydrogenase activity and lactate levels, and metabolic markers in exercise. Notably, the EXE protocol regulated the Akt/p38 MAPK/Nrf2 pathway, which was associated with decreased Hsp70 activation, culminating in glutathione turnover regulation. Moreover, reducing the locomotion environment in the MLF group decreased endurance-like tolerance and did not alter citrate synthase activity, lactate dehydrogenase activity, or lactate levels. The MLF treatment promoted a pro-oxidant effect, altering the Akt/p38 MAPK/Nrf2 pathway and increasing Hsp70 levels, leading to a poorly-regulated glutathione system. Lastly, it was demonstrated that exercise could modulate major metabolic responses in Drosophila melanogaster aerobic and anaerobic metabolism, associated with apoptosis and cellular redox balance maintenance in an emergent exercise model.
Dey, M., Ganguly, A. and Dahanukar, A. (2023). An inhibitory mechanism for suppressing high salt intake in Drosophila. Chem Senses 48. PubMed ID: 37201555
Summary:
High concentrations of dietary salt are harmful to health. Like most animals, Drosophila melanogaster are attracted to foods that have low concentrations of salt, but show strong taste avoidance of high salt foods. Salt in known on multiple classes of taste neurons, activating Gr64f sweet-sensing neurons that drive food acceptance and 2 others (Gr66a bitter and Ppk23 high salt) that drive food rejection. This study finds that NaCl elicits a bimodal dose-dependent response in Gr64f taste neurons, which show high activity with low salt and depressed activity with high salt. High salt also inhibits the sugar response of Gr64f neurons, and this action is independent of the neuron's taste response to salt. Consistent with the electrophysiological analysis, feeding suppression in the presence of salt correlates with inhibition of Gr64f neuron activity, and remains if high salt taste neurons are genetically silenced. Other salts such as Na2SO4, KCl, MgSO4, CaCl2, and FeCl3 act on sugar response and feeding behavior in the same way. A comparison of the effects of various salts suggests that inhibition is dictated by the cationic moiety rather than the anionic component of the salt. Notably, high salt-dependent inhibition is not observed in Gr66a neurons-response to a canonical bitter tastant, denatonium, is not altered by high salt. Overall, this study characterizes a mechanism in appetitive Gr64f neurons that can deter ingestion of potentially harmful salts.

Thursday, September 21st - Disease Models

Del Rocio Perez Baca, M., Jacobs, E. Z., Vantomme, L., ..., Vergult, S. and Callewaert, B. (2023). A novel neurodevelopmental syndrome caused by loss-of-function of the Zinc Finger Homeobox 3 (ZFHX3) gene. medRxiv. PubMed ID: 37292950
Summary:
This study identified loss-of-function variation in ZFHX3 as a novel cause for syndromic intellectual disability (ID). ZFHX3 is a zinc-finger homeodomain transcription factor involved in multiple biological processes including cell differentiation and tumorigenesis. Clinical and morphometric data were collected of 41 individuals with protein truncating variants (PTVs) or (partial) deletions of ZFHX3. Immunoprecipitation followed by mass spectrometry indicated potential binding partners of endogenous ZFHX3 in neural stem cells. A reversed genetic approach characterized the ZFHX3 orthologue, Zfh2 in Drosophila melanogaster. Loss-of-function variation of ZFHX3 consistently associates with (mild) ID and/or behavioural problems, postnatal growth retardation, feeding difficulties, and recognizable facial characteristics, including the rare occurrence of cleft palate. Nuclear abundance of ZFHX3 increases during human brain development and neuronal differentiation in neural stem cells and SH-SY5Y cells, ZFHX3 interacts with the chromatin remodelling BRG1/Brm-associated factor complex and the cleavage and polyadenylation complex. In line with a role for chromatin remodelling, ZFHX3 haploinsufficiency associates with a specific DNA methylation profile in leukocyte-derived DNA. The target genes of ZFHX3 are implicated in neuron and axon development. In Drosophila melanogaster, zfh2, considered to be the ZFHX3 orthologue, is expressed in the third instar larval brain. Ubiquitous and neuron-specific knockdown of zfh2 results in adult lethality underscoring a key role for zfh2 in development and neurodevelopment. Interestingly, ectopic expression of zfh2 as well as ZFHX3 in the developing wing disc results in a thoracic cleft phenotype. Collectively, these data shows that loss-of-function variants in ZFHX3 are a cause of syndromic ID, that associates with a specific DNA methylation profile. Furthermore, this study shows that ZFHX3 participates in chromatin remodelling and mRNA processing.
Deshpande, P., Chimata, A. V., Snider, E., Singh, A., Kango-Singh, M. and Singh, A. (2023). N-Acetyltransferase 9 ameliorates Aβ42-mediated neurodegeneration in the Drosophila eye. Cell Death Dis 14(7): 478. PubMed ID: 37507384
Summary:
Alzheimer's disease (AD), a progressive neurodegenerative disorder, manifests as accumulation of amyloid-beta-42 (Aβ42) plaques and intracellular accumulation of neurofibrillary tangles (NFTs) that results in microtubule destabilization. Targeted expression of human Aβ42 (GMR > Aβ42) in developing Drosophila eye retinal neurons results in Aβ42 plaque(s) and mimics AD-like extensive neurodegeneration. However, there remains a gap in understanding of the underlying mechanism(s) for Aβ42-mediated neurodegeneration. To address this gap in information, a forward genetic screen was conducted, and N-acetyltransferase 9 (Mnat9) was identified as a genetic modifier of GMR > Aβ42 neurodegenerative phenotype. Mnat9 is known to stabilize microtubules by inhibiting c-Jun-N- terminal kinase (JNK) signaling. This study found that gain-of-function of Mnat9 rescues GMR > Aβ42 mediated neurodegenerative phenotype whereas loss-of-function of Mnat9 exhibits the converse phenotype of enhanced neurodegeneration. A new neuroprotective function of Mnat9 is proposed in downregulating the JNK signaling pathway to ameliorate Aβ42-mediated neurodegeneration, which is independent of its acetylation activity. Transgenic flies expressing human NAT9 (hNAT9), also suppresses Aβ42-mediated neurodegeneration thereby suggesting functional conservation in the interaction of fly Mnat9 or hNAT9 with JNK-mediated neurodegeneration. These studies add to the repertoire of molecular mechanisms that mediate cell death response following accumulation of Aβ42 and may provide new avenues for targeting neurodegeneration.
Chen, H., Li, Y., Gao, J., Cheng, Q., Liu, L. and Cai, R. (2023). Activation of Pgk1 Results in Reduced Protein Aggregation in Diverse Neurodegenerative Conditions. Mol Neurobiol 60(9): 5090-5101. PubMed ID: 37249790
Summary:
The prevention of protein condensates has emerged as a new drug target to treat diverse neurodegenerative disorders. Previous work has shown that terazosin (TZ), a prescribed antagonist of the α1 adrenergic receptor, is an activator of phosphoglycerate kinase 1 (Pgk1; see Drosophila Foraging) and Hsp90. This study aimed to determine whether TZ prevents the formation of diverse pathological condensates in cell cultures and animal disease models. In primary neuron culture, TZ treatment reduced both the protein density and abundance of fused in sarcoma (FUS)-P525L-GFP, a disease-associated mutant form of FUS. Regarding the mechanism, this study found that increased intracellular ATP levels were critical for the reduction in protein aggregate density. In addition, Hsp90 activation by TZ enhanced Hsp90 interaction with ULK1, a master regulator of autophagy. Through in vivo studies, neuron-specific overexpression of tau in Drosophila, mouse models of APP/PS1 Alzheimer's disease (AD), and a rat model of multiple system atrophy (MSA) were examined via the viral expression of α-synuclein in the striatum. TZ prevented and reversed the formation of pathological protein condensates. Together, these results suggest that activation of Pgk1 in cytosol may dissolve pathological protein aggregates via increased ATP levels and degrade these proteins via autophagy; the FUS-P525L degradation pathway in nucleus is unclear.
De Donno, M. D., Puricella, A., D'Attis, S., Specchia, V. and Bozzetti, M. P. (2023). Expression of Transposable Elements in the Brain of the Drosophila melanogaster Model for Fragile X Syndrome. Genes (Basel) 14(5). PubMed ID: 37239420
Summary:
Fragile X syndrome is a neuro-developmental disease affecting intellectual abilities and social interactions. Drosophila melanogaster represents a consolidated model to study neuronal pathways underlying this syndrome, especially because the model recapitulates complex behavioural phenotypes. Drosophila Fragile X protein, or FMRP, is required for a normal neuronal structure and for correct synaptic differentiation in both the peripheral and central nervous systems, as well as for synaptic connectivity during development of the neuronal circuits. At the molecular level, FMRP has a crucial role in RNA homeostasis, including a role in transposon RNA regulation in the gonads of D. m. Transposons are repetitive sequences regulated at both the transcriptional and post-transcriptional levels to avoid genomic instability. De-regulation of transposons in the brain in response to chromatin relaxation has previously been related to neurodegenerative events in Drosophila models. This study demonstrates for the first time that FMRP is required for transposon silencing in larval and adult brains of Drosophila "loss of function" dFmr1 mutants. This study highlights that flies kept in isolation, defined as asocial conditions, experience activation of transposable elements. In all, these results suggest a role for transposons in the pathogenesis of certain neurological alterations in Fragile X as well as in abnormal social behaviors.
Ciampelli, C., Galleri, G., Puggioni, S., Fais, M., Iannotta, L., Galioto, M., Becciu, M., Greggio, E., Bernardoni, R., Crosio, C. and Iaccarino, C. (2023). Inhibition of the Exocyst Complex Attenuates the LRRK2 Pathological Effects. Int J Mol Sci 24(16). PubMed ID: 37628835
Summary:
Pathological mutations in leucine-rich repeat kinase 2 (LRRK2) gene are the major genetic cause of Parkinson's disease (PD). Multiple lines of evidence link LRRK2 to the control of vesicle dynamics through phosphorylation of a subset of RAB proteins. However, the molecular mechanisms underlying these processes are not fully elucidated. Previous work has demonstrated that LRRK2 increases the exocyst complex assembly by Sec8 interaction, one of the eight members of the exocyst complex, and that Sec8 over-expression mitigates the LRRK2 pathological effect in PC12 cells. This study extended this analysis using LRRK2 drosophila models and show that the LRRK2-dependent exocyst complex assembly increase is downstream of RAB phosphorylation. Moreover, exocyst complex inhibition rescues mutant LRRK2 pathogenic phenotype in cellular and Drosophila models. Finally, prolonged exocyst inhibition leads to a significant reduction in the LRRK2 protein level, overall supporting the role of the exocyst complex in the LRRK2 pathway. Taken together, this study suggests that modulation of the exocyst complex may represent a novel therapeutic target for PD.
Fujino, Y., Ueyama, M., Ishiguro, T., Ozawa, D., Ito, H., Sugiki, T., Murata, A., Ishiguro, A., Gendron, T., Mori, K., Tokuda, E., Taminato, T., Konno, T., Koyama, A., Kawabe, Y., Takeuchi, T., Furukawa, Y., Fujiwara, T., Ikeda, M., Mizuno, T., Mochizuki, H., Mizusawa, H., Wada, K., Ishikawa, K., Onodera, O., Nakatani, K., Petrucelli, L., Taguchi, H. and Nagai, Y. (2023). FUS regulates RAN translation through modulating the G-quadruplex structure of GGGGCC repeat RNA in C9orf72-linked ALS/FTD. Elife 12. PubMed ID: 37461319
Summary:
Abnormal expansions of GGGGCC repeat sequence in the noncoding region of the C9orf72 gene is the most common cause of familial amyotrophic lateral sclerosis and frontotemporal dementia (C9-ALS/FTD). The expanded repeat sequence is translated into dipeptide repeat proteins (DPRs) by noncanonical repeat-associated non-AUG (RAN) translation. Since DPRs play central roles in the pathogenesis of C9-ALS/FTD, this study investigated the regulatory mechanisms of RAN translation, focusing on the effects of RNA-binding proteins (RBPs) targeting GGGGCC repeat RNAs. Using C9-ALS/FTD model flies, this study demonstrated that the ALS/FTD-linked RBP FUS suppresses RAN translation and neurodegeneration in an RNA-binding activity-dependent manner. Moreover, this study found that FUS directly binds to and modulates the G-quadruplex structure of GGGGCC repeat RNA as an RNA chaperone, resulting in the suppression of RAN translation in vitro. These results reveal a previously unrecognized regulatory mechanism of RAN translation by G-quadruplex-targeting RBPs, providing therapeutic insights for C9-ALS/FTD and other repeat expansion diseases.

Wednesday, September 20th - Signaling

DeGroot, M. S., Williams, B., Chang, T. Y., Maas Gamboa, M. L., Larus, I. M., Hong, G., Fromme, J. C. and Liu, J. (2023). SMOC-1 interacts with both BMP and glypican to regulate BMP signaling in C. elegans. PLoS Biol 21(8): e3002272. PubMed ID: 37590248
Summary:
Secreted modular calcium-binding proteins (SMOCs) are conserved matricellular proteins found in organisms from Caenorhabditis elegans to humans. SMOC homologs characteristically contain 1 or 2 extracellular calcium (EC)-binding domain(s) and 1 or 2 thyroglobulin type-1 (TY) domain(s). SMOC proteins in Drosophila and Xenopus have been found to interact with cell surface heparan sulfate proteoglycans (HSPGs) to exert both positive and negative influences on the conserved bone morphogenetic protein (BMP) signaling pathway. This study used a combination of biochemical, structural modeling, and molecular genetic approaches to dissect the functions of the sole SMOC protein in C. elegans. CeSMOC-1 binds to the heparin sulfate proteoglycan GPC3 homolog LON-2/glypican, as well as the mature domain of the BMP2/4 homolog DBL-1. Moreover, CeSMOC-1 can simultaneously bind LON-2/glypican and DBL-1/BMP. The interaction between CeSMOC-1 and LON-2/glypican is mediated specifically by the EC domain of CeSMOC-1, while the full interaction between CeSMOC-1 and DBL-1/BMP requires full-length CeSMOC-1. Both in vitro biochemical and in vivo functional evidence id provided demonstrating that CeSMOC-1 functions both negatively in a LON-2/glypican-dependent manner and positively in a DBL-1/BMP-dependent manner to regulate BMP signaling. It was further shown that in silico, Drosophila and vertebrate SMOC proteins can also bind to mature BMP dimers. This work provides a mechanistic basis for how the evolutionarily conserved SMOC proteins regulate BMP signaling.
Eidhof, I., Krebbers, A., van de Warrenburg, B. and Schenck, A. (2023). Ataxia-associated DNA repair genes protect the Drosophila mushroom body and locomotor function against glutamate signaling-associated damage. Front Neural Circuits 17: 1148947. PubMed ID: 37476399
Summary:
The precise control of motor movements is of fundamental importance to all behaviors in the animal kingdom. Efficient motor behavior depends on dedicated neuronal circuits - such as those in the cerebellum - that are controlled by extensive genetic programs. Autosomal recessive cerebellar ataxias (ARCAs) provide a valuable entry point into how interactions between genetic programs maintain cerebellar motor circuits. Previous work identified a striking enrichment of DNA repair genes in ARCAs. How dysfunction of ARCA-associated DNA repair genes leads to preferential cerebellar dysfunction and impaired motor function is however unknown. This study used Drosophila to characterize the function of ARCA-associated DNA repair genes in the mushroom body (MB). The MB was shown to be required for efficient startle-induced and spontaneous motor behaviors. Inhibition of synaptic transmission and loss-of-function of ARCA-associated DNA repair genes in the MB affected motor behavior. These motor deficits correlated with increased levels of MB DNA damage, MB Kenyon cell apoptosis and/or alterations in MB morphology. It was further shown that expression of genes involved in glutamate signaling pathways are highly, specifically, and persistently elevated in the postnatal human cerebellum. Manipulation of glutamate signaling in the MB induced motor defects, Kenyon cell DNA damage and apoptosis. Importantly, pharmacological reduction of glutamate signaling in the ARCA DNA repair models rescued the identified motor deficits, suggesting a role for aberrant glutamate signaling in ARCA-DNA repair disorders. In conclusion, the data highlight the importance of ARCA-associated DNA repair genes and glutamate signaling pathways to the cerebellum, the Drosophila MB and motor behavior. It is proposed that glutamate signaling may confer preferential cerebellar vulnerability in ARCA-associated DNA repair disorders.
Brown, J. and Su, T. T. (2023). E2F1 promotes, JNK and DIAP1 inhibit, and chromosomal position has little effect on radiation-induced Loss of Heterozygosity in Drosophila. bioRxiv. PubMed ID: 37214983
Summary:
Loss of Heterozygosity (LOH) can occur when a heterozygous mutant cell loses the remaining wild type allele to become a homozygous mutant. LOH can have physiological consequences if, for example, the affected gene encodes a tumor suppressor. This study used two fluorescent reporters to study mechanisms of LOH induction by X-rays, a type of ionizing radiation (IR), in Drosophila larval wing discs. IR is used to treat more than half of cancer patients, so understanding its effects is of biomedical relevance. IR-induced LOH does not correlate with the chromosomal position of the LOH locus, unlike previously shown for spontaneously occurring LOH. Like spontaneous LOH, however, IR-induced LOH of X-linked loci shows a sex-dependence, occurring predominately in females. A focused genetic screen identified E2F1 as a key promotor of LOH and further testing suggests a mechanism involving its role in cell cycle regulation rather than apoptosis. The QF/QS LOH reporter was combined with QUAS-transgenes to manipulate gene function after LOH induction. This approach identified JNK signaling and apoptosis as key determinants of LOH maintenance. These studies reveal previously unknown mechanisms for generation and maintenance of cells with chromosome aberrations after exposure to IR.
Dawson, J. E., Bryant, A., Jordan, T., Ajamu-Johnson, A., Langridge, P. and Malmi-Kakkada, A. N. (2023). Emergent spatiotemporal patterning of synthetic Notch signal transduction in vivo. bioRxiv. PubMed ID: 37503188
Summary:
Cell-cell communication through direct contact is essential during multiple fundamental biological processes. Synthetic forms of contacted-mediated cell-cell communication can generate custom gene expression outputs, making them valuable for tissue engineering and regenerative medicine. Mechanisms underlying the spatiotemporal behavior of synthetic signal outputs in growing tissues, necessary to precisely control the output location and timing, are not well understood. Towards this goal, this study combined theory and quantitative experiments to study patterns of synthetic Notch (synNotch) activation a custom synthetic gene circuit that was implement within growing Drosophila wing imaginal discs. Cell growth, division, output synthesis and degradation are the key minimal parameters that predict the heterogenous spatiotemporal patterns of synNotch activation in tissues. At long times, synNotch output forms a graded exponential spatial profile that extends several cell diameters from the signal source, indicating a role of cell division in signal propagation. Furthermore, it was discovered that the shape of the interface between ligand and receptor cells is important in determining the synNotch output. Overall, this study elucidate key biophysical principles that underlie complex emergent spatiotemporal patterns of synNotch output in growing tissues.
Colizzi, F. S., Veenstra, J. A., Rezende, G. L., Helfrich-Forster, C. and Martinez-Torres, D. (2023). Pigment-dispersing factor is present in circadian clock neurons of pea aphids and may mediate photoperiodic signalling to insulin-producing cells. Open Biol 13(6): 230090. PubMed ID: 37369351
Summary:
The neuropeptide pigment-dispersing factor (PDF) plays a pivotal role in the circadian clock of most Ecdysozoa and is additionally involved in the timing of seasonal responses of several photoperiodic species. The pea aphid, Acyrthosiphon pisum, is a paradigmatic photoperiodic species with an annual life cycle tightly coupled to the seasonal changes in day length. Nevertheless, PDF could not be identified in A. pisum so far. The present identified a PDF-coding gene that has undergone significant changes in the otherwise highly conserved insect C-terminal amino acid sequence. A newly generated aphid-specific PDF antibody stained four neurons in each hemisphere of the aphid brain that co-express the clock protein Period and have projections to the pars lateralis that are highly plastic and change their appearance in a daily and seasonal manner, resembling those of the fruit fly PDF neurons. Most intriguingly, the PDF terminals overlap with dendrites of the insulin-like peptide (ILP) positive neurosecretory cells in the pars intercerebralis and with putative terminals of Cryptochrome (CRY) positive clock neurons. Since ILP has been previously shown to be crucial for seasonal adaptations and CRY might serve as a circadian photoreceptor vital for measuring day length, these results suggest that PDF plays a critical role in aphid seasonal timing.

Boutet, A., Zeledon, C. and Emery, G. (2023). ArfGAP1 regulates the endosomal sorting of guidance receptors to promote directed collective cell migration in vivo. iScience 26(8): 107467. PubMed ID: 37599820
Summary:
. .

Chemotaxis drives diverse migrations important for development and involved in diseases, including cancer progression. Using border cells in the Drosophila egg chamber as a model for collective cell migration, this study characterized the role of ArfGAP1 in regulating chemotaxis during this process. ArfGAP1 was found to be required for the maintenance of receptor tyrosine kinases, the guidance receptors, at the plasma membrane. In the absence of ArfGAP1, the level of active receptors is reduced at the plasma membrane and increased in late endosomes. Consequently, clusters with impaired ArfGAP1 activity lose directionality. Furthermore, it was found that the number and size of late endosomes and lysosomes are increased in the absence of ArfGAP1. Finally, genetic interactions suggest that ArfGAP1 acts on the kinase and GTPase Lrrk to regulate receptor sorting. Overall, these data indicate that ArfGAP1 is required to maintain guidance receptors at the plasma membrane and promote chemotaxis.

Tuesday, September 19th - Transposons and RNAi

Bologa, A. M., Stoica, I., Constantin, N. D. and Ecovoiu, A. A. (2023). The Landscape of the DNA Transposons in the Genome of the Horezu_LaPeri Strain of Drosophila melanogaster. Insects 14(6). PubMed ID: 37367310
Summary:
>Natural transposons (NTs) represent mobile DNA sequences found in both prokaryotic and eukaryotic genomes. Drosophila melanogaster (the fruit fly) is a eukaryotic model organism with NTs standing for about 20% of its genome and has contributed significantly to the understanding of various aspects of transposon biology. This study describes an accurate approach designed to map class II transposons (DNA transposons) in the genome of the Horezu_LaPeri fruit fly strain, consecutive to Oxford Nanopore Technology sequencing. A whole genome bioinformatics analysis was conducted using Genome ARTIST_v2, LoRTE and RepeatMasker tools to identify DNA transposons insertions. Then, a gene ontology enrichment analysis was performed in order to evaluate the potential adaptive role of some DNA transposons insertions. This study describes DNA transposon insertions specific for the Horezu_LaPeri genome and a predictive functional analysis of some insertional alleles. The PCR validation of P-element insertions specific for this fruit fly strain, along with a putative consensus sequence for the KP element, is also reported. Overall, the genome of the Horezu_LaPeri strain contains several insertions of DNA transposons associated with genes known to be involved in adaptive processes. For some of these genes, insertional alleles obtained via mobilization of the artificial transposons were previously reported. This is a very alluring aspect, as it suggests that insertional mutagenesis experiments conducting adaptive predictions for laboratory strains may be confirmed by mirroring insertions which are expected to be found at least in some natural fruit fly strains.
Asif-Laidin, A., Casier, K., Ziriat, Z., Boivin, A., Viode, E., Delmarre, V., Ronsseray, S., Carre, C. and Teysset, L. (2023). Modeling early germline immunization after horizontal transfer of transposable elements reveals internal piRNA cluster heterogeneity. BMC Biol 21(1): 117. PubMed ID: 37226160
Summary:
A fraction of all genomes is composed of transposable elements (TEs) whose mobility needs to be carefully controlled. In gonads, TE activity is repressed by PIWI-interacting RNAs (piRNAs), a class of small RNAs synthesized by heterochromatic loci enriched in TE fragments, called piRNA clusters. Maintenance of active piRNA clusters across generations is secured by maternal piRNA inheritance providing the memory for TE repression. On rare occasions, genomes encounter horizontal transfer (HT) of new TEs with no piRNA targeting them, threatening the host genome integrity. Naive genomes can eventually start to produce new piRNAs against these genomic invaders, but the timing of their emergence remains elusive. Using a set of TE-derived transgenes inserted in different germline piRNA clusters and functional assays, this study has modeled a TE HT in Drosophila melanogaster. The complete co-option of these transgenes by a germline piRNA cluster can occur within four generations, associated with the production of new piRNAs all along the transgenes and the germline silencing of piRNA sensors. Synthesis of new transgenic TE piRNAs is linked to piRNA cluster transcription dependent on Moonshiner and heterochromatin mark deposition that propagates more efficiently on short sequences. Moreover, sequences located within piRNA clusters can have different piRNA profiles and can influence transcript accumulation of nearby sequences. This study reveals that genetic and epigenetic properties, such as transcription, piRNA profiles, heterochromatin, and conversion efficiency along piRNA clusters, could be heterogeneous depending on the sequences that compose them. These findings suggest that the capacity of transcriptional signal erasure induced by the chromatin complex specific of the piRNA cluster can be incomplete through the piRNA cluster loci. Finally, these results have revealed an unexpected level of complexity that highlights a new magnitude of piRNA cluster plasticity fundamental for the maintenance of genome integrity.
Brown, J. S., Zhang, D., Gaylord, O., Chen, W. and Lee, H. C. (2023). Sensitized piRNA reporter identifies multiple RNA processing factors involved in piRNA-mediated gene silencing. Genetics 224(4). PubMed ID: 37210214
Summary:
Metazoans guard their germlines against transposons and other foreign transcripts with PIWI-interacting RNAs (piRNAs). Due to the robust heritability of the silencing initiated by piRNAs in Caenorhabditis elegans (C. elegans), previous screens using C. elegans were strongly biased to uncover members of this pathway in the maintenance process but not in the initiation process. To identify novel piRNA pathway members, a sensitized reporter strain was utilized which detects defects in initiation, amplification, or regulation of piRNA silencing. Using this reporter,Integrator complex subunits, nuclear pore components, protein import components, and pre-mRNA splicing factors were identified as essential for piRNA-mediated gene silencing. The small nuclear processing cellular machine termed the Integrator complex is required for both type I and type II piRNA production. Notably, a role was identied for nuclear pore and nucleolar components NPP-1/Nup54, NPP-6/Nup160, NPP-7/Nup153, and FIB-1 in promoting the perinuclear localization of anti-silencing CSR-1 Argonaute, as well as a role for Importin factor IMA-3 in nuclear localization of silencing Argonaute HRDE-1. Together, this study has shown that piRNA silencing in C. elegans is dependent on evolutionarily ancient RNA processing machinery that has been co-opted to function in the piRNA-mediated genome surveillance pathway.
Deng, T., Su, S., Yuan, X., He, J., Huang, Y., Ma, J. and Wang, J. (2023). Structural mechanism of R2D2 and Loqs-PD synergistic modulation on DmDcr-2 oligomers. Nat Commun 14(1): 5228. PubMed ID: 37633971
Summary:
Small interference RNAs are the key components of RNA interference, a conserved RNA silencing or viral defense mechanism in many eukaryotes. In Drosophila melanogaster, Dicer-2 (DmDcr-2)-mediated RNAi pathway plays important roles in defending against viral infections and protecting genome integrity. During the maturation of siRNAs, two cofactors can regulate DmDcr-2's functions: Loqs-PD that is required for dsRNA processing, and R2D2 that is essential for the subsequent loading of siRNAs into effector Ago2 to form RISC complexes. However, due to the lack of structural information, it is still unclear whether R2D2 and Loqs-PD affect the functions of DmDcr-2 simultaneously. This study presents several cryo-EM structures of DmDcr-2/R2D2/Loqs-PD complex bound to dsRNAs with various lengths by the Helicase domain. These structures revealed that R2D2 and Loqs-PD can bind to different regions of DmDcr-2 without interfering with each other. Furthermore, the cryo-EM results demonstrate that these complexes can form large oligomers and assemble into fibers. The formation and depolymerization of these oligomers are associated with ATP hydrolysis. These findings provide insights into the structural mechanism of DmDcr-2 and its cofactors during siRNA processing.
Aute, R. and Deshmukh, M. V. (2023). Chemical shift assignments of dsRBD1 and linker region of R2D2, a siRNA binding protein in the Drosophila RNAi pathway. Biomol NMR Assign. PubMed ID: 37405581
Summary:
In the model organism Drosophila melanogaster, one of the Dicer homologs, Dcr-2, initiates the RNA interference pathway by cleaving long double-stranded RNA into small interfering RNA (siRNA). The Dcr-2:R2D2 heterodimer subsequently binds to the 21-nucleotide siRNA to form the R2D2:Dcr-2 Initiator (RDI) complex, which is critical for initiating the assembly of the RNA-induced silencing complex containing guide siRNA strand. During RDI complex formation, R2D2 senses the stability of the 5' end of the siRNA and a 5'-phosphate group, although the underlying mechanism of siRNA asymmetry sensing and 5'-phosphate recognition by R2D2 is elusive. This study presents nearly complete chemical shift assignments of the backbone and the side chain of a construct that comprises the N-terminus dsRBD1 and linker of R2D2 (~ 10.3 kDa; henceforth: R2D2D1L). This study would further aid in the structural and functional characterization of R2D2.
Chary, S. and Hayashi, R. (2023). The absence of core piRNA biogenesis factors does not impact efficient transposon silencing in Drosophila. PLoS Biol 21(6): e3002099. PubMed ID: 37279192
Summary:
Organisms require mechanisms to distinguish self and non-self-RNA. This distinction is crucial to initiate the biogenesis of Piwi-interacting RNAs (piRNAs). In Drosophila ovaries, PIWI-guided slicing and the recognition of piRNA precursor transcripts by the DEAD-box RNA helicase Yb are the 2 known mechanisms to licence an RNA for piRNA biogenesis in the germline and the soma, respectively. Both the PIWI proteins and Yb are highly conserved across most Drosophila species and are thought to be essential to the piRNA pathway and for silencing transposons. However, this study found that species closely related to Drosophila melanogaster have lost the yb gene, as well as the PIWI gene Ago3. The precursor RNA is still selected in the absence of Yb to abundantly generate transposon antisense piRNAs in the soma. It was further demonstrated that Drosophila eugracilis, which lacks Ago3, is completely devoid of ping-pong piRNAs and exclusively produces phased piRNAs in the absence of slicing. Thus, core piRNA pathway genes can be lost in evolution while still maintaining efficient transposon silencing.

Monday, September 18th - Adult Neural Development, Structure and Function

De, J., Wu, M., Lambatan, V., Hua, Y. and Joiner, W. J. (2023). Re-examining the role of the dorsal fan-shaped body in promoting sleep in Drosophila. Curr Biol. PubMed ID: 37552985
Summary:
The needs fulfilled by sleep are unknown, though the effects of insufficient sleep are manifold. To better understand how the need to sleep is sensed and discharged, much effort has gone into identifying the neural circuits involved in regulating arousal, especially those that promote sleep. In prevailing models, the dorsal fan-shaped body (dFB) plays a central role in this process in the fly brain. This study manipulated various properties of the dFB including its electrical activity, synaptic output, and endogenous gene expression. In each of these experimental contexts it was not possible to identify any effect on sleep that could be unambiguously mapped to the dFB. Furthermore, evidence was found that sleep phenotypes previously attributed to the dFB were caused by genetic manipulations that inadvertently targeted the ventral nerve cord. Expression was examined of two genes whose purported effects have been attributed to functions within a specific subpopulation of dFB neurons. In both cases little to no expression expression was found in the expected cells. Collectively, these results cast doubt on the prevailing hypothesis that the dFB plays a central role in promoting sleep.
Chen, S. F., Hsien, H. L., Wang, T. F. and Lin, M. D. (2023). Drosophila Phosphatase of Regenerating Liver Is Critical for Photoreceptor Cell Polarity and Survival during Retinal Development. Int J Mol Sci 24(14). PubMed ID: 37511262
Summary:
Establishing apicobasal polarity, involving intricate interactions among polarity regulators, is key for epithelial cell function. Though phosphatase of regenerating liver (PRL) proteins are implicated in diverse biological processes, including cancer, their developmental role remains unclear. This study explored the role of Drosophila PRL (dPRL) in photoreceptor cell development. dPRL, requiring a C-terminal prenylation motif, is highly enriched in the apical membrane of developing photoreceptor cells. Moreover, dPRL knockdown during retinal development results in adult Drosophila retinal degeneration, caused by hid-induced apoptosis. dPRL depletion also mislocalizes cell adhesion and polarity proteins like Armadillo, Crumbs, and DaPKC and relocates the basolateral protein, alpha subunit of Na(+)/K(+)-ATPase, to the presumed apical membrane. Importantly, this polarity disruption is not secondary to apoptosis, as suppressing hid expression does not rescue the polarity defect in dPRL-depleted photoreceptor cells. These findings underscore dPRL's crucial role in photoreceptor cell polarity and emphasize PRL's importance in establishing epithelial polarity and maintaining cell survival during retinal development, offering new insights into PRL's role in normal epithelium.
Chen, N., Zhang, Y., Rivera-Rodriguez, E. J., Yu, A. D., Hobin, M., Rosbash, M. and Griffith, L. C. (2023). Widespread posttranscriptional regulation of cotransmission.. Sci Adv 9(22): eadg9836. PubMed ID: 37267358
Summary:
While neurotransmitter identity was once considered singular and immutable for mature neurons, it is now appreciated that one neuron can release multiple neuroactive substances (cotransmission) whose identities can even change over time. To explore the mechanisms that tune the suite of transmitters a neuron releases, this study developed transcriptional and translational reporters for cholinergic, glutamatergic, and GABAergic signaling in Drosophila. Many glutamatergic and GABAergic cells also transcribe cholinergic genes, but fail to accumulate cholinergic effector proteins. Suppression of cholinergic signaling involves posttranscriptional regulation of cholinergic transcripts by the microRNA miR-190; chronic loss of miR-190 function allows expression of cholinergic machinery, reducing and fragmenting sleep. Using a "translation-trap" strategy, this study shows that neurons in these populations have episodes of transient translation of cholinergic proteins, demonstrating that suppression of cotransmission is actively modulated. Posttranscriptional restriction of fast transmitter cotransmission provides a mechanism allowing reversible tuning of neuronal output.
Christenson, M. P., Diez A, S., Heath, S. L., Saavedra-Weisenhaus, M., Adachi, A., Abbott, L. F. and Behnia, R. (2023). Hue selectivity from recurrent circuitry in Drosophila. bioRxiv. PubMed ID: 37502934
Summary:
A universal principle of sensory perception is the progressive transformation of sensory information from broad non-specific signals to stimulus-selective signals that form the basis of perception. To perceive color, human brains must transform the wavelengths of light reflected off objects into the derived quantities of brightness, saturation and hue. Neurons responding selectively to hue have been reported in primate cortex, but it is unknown how their narrow tuning in color space is produced by upstream circuit mechanisms. To enable circuit level analysis of color perception, this study report sthe discovery of neurons in the Drosophila optic lobe with hue selective properties. Using the connectivity graph of the fly brain, a connectomics-constrained circuit model was constructed that accounts for this hue selectivity. Unexpectedly, the model predicts that recurrent connections in the circuit are critical for hue selectivity. Experiments using genetic manipulations to perturb recurrence in adult flies confirms this prediction. These findings reveal the circuit basis for hue selectivity in color vision.
Chen, C. C., Lin, H. W., Feng, K. L., Tseng, D. W., de Belle, J. S. and Chiang, A. S. (2023). A subset of cholinergic mushroom body neurons blocks long-term memory formation in Drosophila. Cell Rep 42(8): 112974. PubMed ID: 37590142
Summary:
Long term memory (LTM) requires learning-induced synthesis of new proteins allocated to specific neurons and synapses in a neural circuit. Not all learned information, however, becomes permanent memory. How the brain gates relevant information into LTM remains unclear. In Drosophila adults, weak learning after a single training session in an olfactory aversive task typically does not induce protein-synthesis-dependent LTM. Instead, strong learning after multiple spaced training sessions is required. This study reports that pre-synaptic active-zone protein synthesis and cholinergic signaling from the early α/β subset of mushroom body (MB) neurons produce a downstream inhibitory effect on LTM formation. When inhibitory signaling was eliminated from these neurons, weak learning was then sufficient to form LTM. This bidirectional circuit mechanism modulates the transition between distinct memory phase functions in different subpopulations of MB neurons in the olfactory memory circuit.
Cury, K. M. and Axel, R. (2023). Flexible neural control of transition points within the egg-laying behavioral sequence in Drosophila. Nat Neurosci 26(6): 1054-1067. PubMed ID: 37217726
Summary:
Innate behaviors are frequently comprised of ordered sequences of component actions that progress to satisfy essential drives. Progression is governed by specialized sensory cues that induce transitions between components within the appropriate context. This study has characterized the structure of the egg-laying behavioral sequence in Drosophila and found significant variability in the transitions between component actions that affords the organism an adaptive flexibility. Distinct classes of interoceptive and exteroceptive sensory neurons were identified that control the timing and direction of transitions between the terminal components of the sequence. A pair of motor neurons was identified that enact the final transition to egg expulsion. These results provide a logic for the organization of innate behavior in which sensory information processed at critical junctures allows for flexible adjustments in component actions to satisfy drives across varied internal and external environments.

Friday, September 15th - Synapse and Vesicles

Cheon, Y., Yoon, S., Lee, J. H., Kim, K., Kim, H. J., Hong, S. W., Yun, Y. R., Shim, J., Kim, S. H., Lu, B., Lee, M. and Lee, S. (2023). A Novel Interaction between MFN2/Marf and MARK4/PAR-1 Is Implicated in Synaptic Defects and Mitochondrial Dysfunction. eNeuro 10(8). PubMed ID: 37550059
Summary:
As cellular energy powerhouses, mitochondria undergo constant fission and fusion to maintain functional homeostasis. The conserved dynamin-like GTPase, Mitofusin2 (MFN2)/mitochondrial assembly regulatory factor (Marf), plays a role in mitochondrial fusion, mutations of which are implicated in age-related human diseases, including several neurodegenerative disorders. However, the regulation of MFN2/Marf-mediated mitochondrial fusion, as well as the pathologic mechanism of neurodegeneration, is not clearly understood. This study identified a novel interaction between MFN2/Marf and microtubule affinity-regulating kinase 4 (MARK4)/PAR-1. In the Drosophila larval neuromuscular junction, muscle-specific overexpression of MFN2/Marf decreased the number of synaptic boutons, and the loss of MARK4/PAR-1 alleviated the synaptic defects of MFN2/Marf overexpression. Downregulation of MARK4/PAR-1 rescued the mitochondrial hyperfusion phenotype caused by MFN2/Marf overexpression in the Drosophila muscles as well as in the cultured cells. In addition, knockdown of MARK4/PAR-1 rescued the respiratory dysfunction of mitochondria induced by MFN2/Marf overexpression in mammalian cells. Together, these results indicate that the interaction between MFN2/Marf and MARK4/PAR-1 is fine-tuned to maintain synaptic integrity and mitochondrial homeostasis, and its dysregulation may be implicated in neurologic pathogenesis.
Asuncion, J. D., Eamani, A., Rohrbach, E. W., Knapp, E. M., Deshpande, S. A., Bonanno, S. L., Murphy, J. E., Lawal, H. O. and Krantz, D. E. (2023). Precise CRISPR-Cas9-mediated mutation of a membrane trafficking domain in the Drosophila vesicular monoamine transporter gene. Curr Res Physiol 6: 100101. PubMed ID: 37409154
Summary:
Monoamine neurotransmitters such as noradrenalin are released from both synaptic vesicles (SVs) and large dense-core vesicles (LDCVs), the latter mediating extrasynaptic signaling. The contribution of synaptic versus extrasynaptic signaling to circuit function and behavior remains poorly understood. To address this question, transgenes were previously used, encoding a mutation in the Drosophila Vesicular Monoamine Transporter (dVMAT) that shifts amine release from SVs to LDCVs. To circumvent the use of transgenes with non-endogenous patterns of expression, this study used CRISPR-Cas9 to generate a trafficking mutant in the endogenous dVMAT gene. To minimize disruption of the dVMAT coding sequence and a nearby RNA splice site, a point mutation was precisely introduced using single-stranded oligonucleotide repair. A predicted decrease in fertility was used as a phenotypic screen to identify founders in lieu of a visible marker. Phenotypic analysis revealed a defect in the ovulation of mature follicles and egg retention in the ovaries. No defects were detected in the contraction of lateral oviducts following optogenetic stimulation of octopaminergic neurons. These findings suggest that release of mature eggs from the ovary is disrupted by changing the balance of VMAT trafficking between SVs and LDCVs. Further experiments using this model will help determine the mechanisms that sensitize specific circuits to changes in synaptic versus extrasynaptic signaling.
Bulgari, D., Cavolo, S. L., Schmidt, B. F., Buchan, K., Bruchez, M. P., Deitcher, D. L. and Levitan, E. S. (2023). Ca2+ and cAMP open differentially dilating synaptic fusion pores. J Cell Sci 136(13). PubMed ID: 37303204
Summary:
Neuronal dense-core vesicles (DCVs) contain neuropeptides and much larger proteins that affect synaptic growth and plasticity. Rather than using full collapse exocytosis that commonly mediates peptide hormone release by endocrine cells, DCVs at the Drosophila neuromuscular junction release their contents via fusion pores formed by kiss-and-run exocytosis. This study used fluorogen-activating protein (FAP) imaging to reveal the permeability range of synaptic DCV fusion pores and then shows that this constraint is circumvented by cAMP-induced extra fusions with dilating pores that result in DCV emptying. These Ca2+-independent full fusions require PKA-R2, a PKA phosphorylation site on Complexin and the acute presynaptic function of Rugose, the homolog of mammalian neurobeachin, a PKA-R2 anchor implicated in learning and autism. Therefore, localized Ca2+-independent cAMP signaling opens dilating fusion pores to release large cargoes that cannot pass through the narrower fusion pores that mediate spontaneous and activity-dependent neuropeptide release. These results imply that the fusion pore is a variable filter that differentially sets the composition of proteins released at the synapse by independent exocytosis triggers responsible for routine peptidergic transmission (Ca2+) and synaptic development (cAMP).
Cheng, G., Chang, J., Gong, H. and Zhou, W. (2023). A distinct Golgi-targeting mechanism of dGM130 in Drosophila neurons. Front Mol Neurosci 16: 1206219. PubMed ID: 37333614
Summary:
GM130 is a matrix protein that is conserved in metazoans and involved in the architecture of the Golgi apparatus. In neurons, Golgi apparatus and dendritic Golgi outposts (GOs) have different compartmental organizations, and GM130 localization is present in both, indicating that GM130 has a unique Golgi-targeting mechanism. This study investigated the Golgi-targeting mechanism of the GM130 homologue, dGM130, using in vivo imaging of Drosophila dendritic arborization (da) neurons. The results showed that two independent Golgi-targeting domains (GTDs) with different Golgi localization characteristics in dGM130, together determined the precise localization of dGM130 in both the soma and dendrites. GTD1, covering the first coiled-coil region, preferentially targeted to somal Golgi rather than GOs; whereas GTD2, containing the second coiled-coil region and C-terminus, dynamically targeted to Golgi in both soma and dendrites. These findings suggest that there are two distinct mechanisms by which dGM130 targets to the Golgi apparatus and GOs, underlying the structural differences between them, and further provides new insights into the formation of neuronal polarity.
Borchers, A. C., Janz, M., Schafer, J. H., Moeller, A., Kümmel, D., Paululat, A., Ungermann, C. and Langemeyer, L. (2023). Regulatory sites in the Mon1-Ccz1 complex control Rab5 to Rab7 transition and endosome maturation. Proc Natl Acad Sci U S A 120(30): e2303750120. PubMed ID: 37463208
Summary:
Maturation from early to late endosomes depends on the exchange of their marker proteins Rab5 to Rab7. This requires Rab7 activation by its specific guanine nucleotide exchange factor (GEF) Mon1-Ccz1. Efficient GEF activity of this complex on membranes depends on Rab5, thus driving Rab-GTPase exchange on endosomes. However, molecular details on the role of Rab5 in Mon1-Ccz1 activation are unclear. This study identified key features in Mon1 involved in GEF regulation. The intrinsically disordered N-terminal domain of Mon1 was shown to autoinhibit Rab5-dependent GEF activity on membranes. Consequently, Mon1 truncations result in higher GEF activity in vitro and alterations in early endosomal structures in Drosophila nephrocytes. A shift from Rab5 to more Rab7-positive structures in yeast suggests faster endosomal maturation. Using modeling, a conserved Rab5-binding site was identified in Mon1. Mutations impairing Rab5 interaction result in poor GEF activity on membranes and growth defects in vivo. This analysis provides a framework to understand the mechanism of Ras-related in brain (Rab) conversion and organelle maturation along the endomembrane system.
Bruelle, C., Pinot, M., Daniel, E., Daude, M., Mathieu, J. and Borgne, R. L. (2023). Cell-intrinsic and -extrinsic roles of the ESCRT-III subunit Shrub in abscission of Drosophila sensory organ precursors. Development 150(10). PubMed ID: 37226981
Summary:
Although the molecular mechanisms governing abscission of isolated cells have largely been elucidated, those underlying the abscission of epithelial progenitors surrounded by epidermal cells (ECs), connected via cellular junctions, remain largely unexplored. This study investigated the remodeling of the paracellular diffusion barrier ensured by septate junctions (SJs) during cytokinesis of Drosophila sensory organ precursors (SOPs). SOP cytokinesis involves the coordinated, polarized assembly and remodeling of SJs in the dividing cell and its neighbors, which remain connected to the former via membrane protrusions pointing towards the SOP midbody. SJ assembly and midbody basal displacement occur faster in SOPs than in ECs, leading to quicker disentanglement of neighboring cell membrane protrusions prior to midbody release. As reported in isolated cells, the endosomal sorting complex required for the transport-III component Shrub/CHMP4B is recruited at the midbody and cell-autonomously regulates abscission. In addition, Shrub is recruited to membrane protrusions and is required for SJ integrity, and alteration of SJ integrity leads to premature abscission. This study uncovers cell-intrinsic and -extrinsic functions of Shrub in coordinating remodeling of the SJs and SOP abscission.

Thursday, September 14th - Adult Neural Development, Structure and Function

Bonanno, S. L. and Krantz, D. E. (2023). Transcriptional changes in specific subsets of Drosophila neurons following inhibition of the serotonin transporter. Transl Psychiatry 13(1): 226. PubMed ID: 37355701
Summary:
The transcriptional effects of SSRIs and other serotonergic drugs remain unclear, in part due to the heterogeneity of postsynaptic cells, which may respond differently to changes in serotonergic signaling. Relatively simple model systems such as Drosophila afford more tractable microcircuits in which to investigate these changes in specific cell types. This study focused on the mushroom body, an insect brain structure heavily innervated by serotonin and comprised of multiple different but related subtypes of Kenyon cells. Fluorescence-activated cell sorting of Kenyon cells, followed by either bulk or single-cell RNA sequencing were used to explore the transcriptomic response of these cells to SERT inhibition. The effects of two different Drosophila Serotonin Transporter (dSERT) mutant alleles as well as feeding the SSRI citalopram to adult flies were compared. The genetic architecture associated with one of the mutants contributed to significant artefactual changes in expression. Comparison of differential expression caused by loss of SERT during development versus aged, adult flies, suggests that changes in serotonergic signaling may have relatively stronger effects during development, consistent with behavioral studies in mice. Overall, these experiments revealed limited transcriptomic changes in Kenyon cells, but suggest that different subtypes may respond differently to SERT loss-of-function. Further work exploring the effects of SERT loss-of-function in other circuits may be used help to elucidate how SSRIs differentially affect a variety of different neuronal subtypes both during development and in adults.
Chapochnikov, N. M., Pehlevan, C. and Chklovskii, D. B. (2023). Normative and mechanistic model of an adaptive circuit for efficient encoding and feature extraction. Proc Natl Acad Sci U S A 120(29): e2117484120. PubMed ID: 37428907
Summary:
One major question in neuroscience is how to relate connectomes to neural activity, circuit function, and learning. This study offers an answer in the peripheral olfactory circuit of the Drosophila larva, composed of olfactory receptor neurons (ORNs) connected through feedback loops with interconnected inhibitory local neurons (LNs). This study combined structural and activity data and, using a holistic normative framework based on similarity-matching, biologically plausible mechanistic models of the circuit were formulated. In particular, a linear circuit model is considered, for which an exact theoretical solution and a nonnegative circuit model were derived, which was examined through simulations. The latter largely predicts the ORN LN synaptic weights found in the connectome and demonstrates that they reflect correlations in ORN activity patterns. Furthermore, this model accounts for the relationship between ORN LN and LN-LN synaptic counts and the emergence of different LN types. Functionally, it is proposed that LNs encode soft cluster memberships of ORN activity, and partially whiten and normalize the stimulus representations in ORNs through inhibitory feedback. Such a synaptic organization could, in principle, autonomously arise through Hebbian plasticity and would allow the circuit to adapt to different environments in an unsupervised manner. This study thus uncovered a general and potent circuit motif that can learn and extract significant input features and render stimulus representations more efficient. Finally, this study provides a unified framework for relating structure, activity, function, and learning in neural circuits and supports the conjecture that similarity-matching shapes the transformation of neural representations.
Braun, A., Borst, A. and Meier, M. (2023). Disynaptic inhibition shapes tuning of OFF-motion detectors in Drosophila. Curr Biol 33(11): 2260-2269. PubMed ID: 37236181
Summary:
The circuitry underlying the detection of visual motion in Drosophila melanogaster is one of the best studied networks in neuroscience. Lately, electron microscopy reconstructions, algorithmic models, and functional studies have proposed a common motif for the cellular circuitry of an elementary motion detector based on both supralinear enhancement for preferred direction and sublinear suppression for null-direction motion. In T5 cells, however, all columnar input neurons (Tm1, Tm2, Tm4, and Tm9) are excitatory. So, how is null-direction suppression realized there? Using two-photon calcium imaging in combination with thermogenetics, optogenetics, apoptotics, and pharmacology, this study discovered that it is via CT1, the GABAergic large-field amacrine cell, where the different processes have previously been shown to act in an electrically isolated way. Within each column, CT1 receives excitatory input from Tm9 and Tm1 and provides the sign-inverted, now inhibitory input signal onto T5. Ablating CT1 or knocking down GABA-receptor subunit Rdl significantly broadened the directional tuning of T5 cells. It thus appears that the signal of Tm1 and Tm9 is used both as an excitatory input for preferred direction enhancement and, through a sign inversion within the Tm1/Tm9-CT1 microcircuit, as an inhibitory input for null-direction suppression.
Bhattacharya, D., Gorska-Andrzejak, J., Abaquita, T. A. L. and Pyza, E. (2023). Effects of adenosine receptor overexpression and silencing in neurons and glial cells on lifespan, fitness, and sleep of Drosophila melanogaster. Exp Brain Res 241(7): 1887-1904. PubMed ID: 37335362 ID:
Summary:
A single adenosine receptor gene (dAdoR) has been detected in Drosophila melanogaster. However, its function in different cell types of the nervous system is mostly unknown. Therefore, this study overexpressed or silenced the dAdoR gene in eye photoreceptors, all neurons, or glial cells and examined the fitness of flies, the amount and daily pattern of sleep, and the influence of dAdoR silencing on Bruchpilot (BRP) presynaptic protein. Furthermore, the dAdoR and brp gene expression was examined in young and old flies. A higher level of dAdoR was found in the retina photoreceptors, all neurons, and glial cells negatively influenced the survival rate and lifespan of male and female Drosophila in a cell-dependent manner and to a different extent depending on the age of the flies. In old flies, expression of both dAdoR and brp was higher than in young ones. An excess of dAdoR in neurons improved climbing in older individuals. It also influenced sleep by lengthening nighttime sleep and siesta. In turn, silencing of dAdoR decreased the lifespan of flies, although it increased the survival rate of young flies. It hindered the climbing of older males and females, but did not change sleep. Silencing also affected the daily pattern of BRP abundance, especially when dAdoR expression was decreased in glial cells. The obtained results indicate the role of adenosine and dAdoR in the regulation of fitness in flies that is based on communication between neurons and glial cells, and the effect of glial cells on synapses.
Bonheur, M., Swartz, K. J., Metcalf, M. G., Wen, X., Zhukovskaya, A., Mehta, A., Connors, K. E., Barasch, J. G., Jamieson, A. R., Martin, K. C., Axel, R. and Hattori, D. (2023). A rapid and bidirectional reporter of neural activity reveals neural correlates of social behaviors in Drosophila. Nat Neurosci 26(7): 1295-1307. PubMed ID: 37308660
Summary:
Neural activity is modulated over different timescales encompassing subseconds to hours, reflecting changes in external environment, internal state and behavior. Using Drosophila as a model, a rapid and bidirectional reporter was developed that provides a cellular readout of recent neural activity. This reporter uses nuclear versus cytoplasmic distribution of CREB-regulated transcriptional co-activator (CRTC). Subcellular distribution of GFP-tagged CRTC (CRTC::GFP) bidirectionally changes on the order of minutes and reflects both increases and decreases in neural activity. An automated machine-learning-based routine was developed for efficient quantification of reporter signal. Using this reporter, Mating-evoked activation and inactivation of modulatory neurons was demonstrated. The functional role of the master courtship regulator gene fruitless (fru) was further investigated, and it was shown that fru is necessary to ensure activation of male arousal neurons by female cues. Together, these results establish CRTC::GFP as a bidirectional reporter of recent neural activity suitable for examining neural correlates in behavioral contexts.
Bustillo, M. E., Douthit, J., Astigarraga, S. and Treisman, J. E. (2023). Two distinct mechanisms of Plexin A function in Drosophila optic lobe lamination and morphogenesis. bioRxiv. PubMed ID: 37609142
Summary:
Visual circuit development is characterized by subdivision of neuropils into layers that house distinct sets of synaptic connections. This study found that in the Drosophila medulla, this layered organization depends on the axon guidance regulator Plexin A. In plexin A null mutants, synaptic layers of the medulla neuropil and arborizations of individual neurons are wider and less distinct than in controls. Analysis of Semaphorin function indicates that Semaphorin 1a, provided by cells that include Tm5 neurons, is the primary partner for Plexin A in medulla lamination. Removal of the cytoplasmic domain of endogenous Plexin A does not disrupt the formation of medulla layers; however, both null and cytoplasmic domain deletion mutations of plexin A result in an altered overall shape of the medulla neuropil. These data suggest that Plexin A acts as a receptor to mediate morphogenesis of the medulla neuropil, and as a ligand for Semaphorin 1a to subdivide it into layers. Its two independent functions illustrate how a few guidance molecules can organize complex brain structures by each playing multiple roles. The axon guidance molecule Plexin A has two functions in Drosophila medulla development; morphogenesis of the neuropil requires its cytoplasmic domain, but establishing synaptic layers through Semaphorin 1a does not.

Wednesday, September 13th - Cell Cycle

Duan, T., Thyagarajan, S., Amoiroglou, A., Rogers, G. C. and Geyer, P. K. (2023). Analysis of a rare progeria variant of Barrier-to-autointegration factor in Drosophila connects centromere function to tissue homeostasis. Cell Mol Life Sci 80(3): 73. PubMed ID: 36842139
Summary:
Barrier-to-autointegration factor (BAF/BANF) is a nuclear lamina protein essential for nuclear integrity, chromatin structure, and genome stability. Whereas complete loss of BAF causes lethality in multiple organisms, the A12T missense mutation of the BANF1 gene in humans causes a premature aging syndrome, called Nestor-Guillermo Progeria Syndrome (NGPS). This study reports the first in vivo animal investigation of progeroid BAF, using CRISPR editing to introduce the NGPS mutation into the endogenous Drosophila baf gene. Progeroid BAF adults are born at expected frequencies, demonstrating that this BAF variant retains some function. However, tissue homeostasis is affected, supported by studies of the ovary, a tissue that depends upon BAF for stem cell survival and continuous oocyte production. This study found that progeroid BAF causes defects in germline stem cell mitosis that delay anaphase progression and compromise chromosome segregation. These defects were linked to decreased recruitment of centromeric proteins of the kinetochore, indicating dysfunction of cenBAF, a localized pool of dephosphorylated BAF produced by Protein Phosphatase PP4. DNA damage was shown to increase in progenitor germ cells, which causes germ cell death due to activation of the DNA damage transducer kinase Chk2. Mitotic defects appear widespread, as aberrant chromosome segregation and increased apoptosis occur in another tissue. Together, these data highlight the importance of BAF in establishing centromeric structures critical for mitosis. Further, these studies link defects in cenBAF function to activation of a checkpoint that depletes progenitor reserves critical for tissue homeostasis, aligning with phenotypes of NGPS patients.
Baker, C. C., Gallicchio, L., Parsanian, L., Taing, E., Tam, C. and Fuller, M. T. (2023). A cell-type-specific multi-protein complex regulates expression of Cyclin B protein in Drosophila male meiotic prophase. bioRxiv. PubMed ID: 36824933
Summary:
During meiosis, germ cell and stage-specific components impose additional layers of regulation on the core cell cycle machinery to set up an extended G2 period termed meiotic prophase. In Drosophila males, meiotic prophase lasts 3.5 days, during which spermatocytes turn up expression of over 3000 genes and grow 25-fold in volume. Previous work showed that the core cell cycle regulator Cyclin B (CycB) is subject to translational repression in immature Drosophila spermatocytes, mediated by the RNA-binding protein Rbp4 and its partner Fest. This study shows that another spermatocyte-specific protein, Lutin (Lutin (encoded by CG1690), is required for translational repression of cycB in an 8-hour window just before spermatocytes are fully mature. In males mutant for rbp4 or lut, spermatocytes enter and exit the meiotic divisions 6-8 hours earlier than in wild-type. In addition, it was shown that spermatocyte-specific isoforms of Syncrip (Syp) are required for expression of CycB protein and normal entry into the meiotic divisions. Both Lut and Syp interact with Fest in an RNA-independent manner. Thus a complex of spermatocyte-specific regulators choreograph the timing of expression of CycB protein during male meiotic prophase. Expression of a conserved cell cycle component, Cyclin B, is regulated by multiple mechanisms in the Drosophila male germline to dictate the correct timing of meiotic division.
Ji, X., Gao, J., Wei, T., Jin, L. and Xiao, G. (2023). Fear-of-intimacy-mediated zinc transport is required for Drosophila fat body endoreplication. BMC Biol 21(1): 88. PubMed ID: 37069617
Summary:

Endoreplication is involved in the development and function of many organs, the pathologic process of several diseases. However, the metabolic underpinnings and regulation of endoreplication have yet to be well clarified. This study showed that a zinc transporter fear-of-intimacy (foi) is necessary for Drosophila fat body endoreplication. foi knockdown in the fat body led to fat body cell nuclei failure to attain standard size, decreased fat body size and pupal lethality. These phenotypes could be modulated by either altered expression of genes involved in zinc metabolism or intervention of dietary zinc levels. Further studies indicated that the intracellular depletion of zinc caused by foi knockdown results in oxidative stress, which activates the ROS-JNK signaling pathway, and then inhibits the expression of Myc, which is required for tissue endoreplication and larval growth in Drosophila. These results indicated that FOI is critical in coordinating fat body endoreplication and larval growth in Drosophila. This study provides a novel insight into the relationship between zinc and endoreplication in insects and may provide a reference for relevant mammalian studies.

Avellino, A., Peng, C. H. and Lin, M. D. (2023). Cell Cycle Regulation by NF-YC in Drosophila Eye Imaginal Disc: Implications for Synchronization in the Non-Proliferative Region. Int J Mol Sci 24(15). PubMed ID: 37569581
Summary:
Cell cycle progression during development is meticulously coordinated with differentiation. This is particularly evident in the Drosophila 3rd instar eye imaginal disc, where the cell cycle is synchronized and arrests at the G1 phase in the non-proliferative region (NPR), setting the stage for photoreceptor cell differentiation. This study identified the transcription factor Nuclear Factor-YC (NF-YC) as a crucial player in this finely tuned progression, elucidating its specific role in the synchronized movement of the morphogenetic furrow. Depletion of NF-YC leads to extended expression of Cyclin A (CycA) and Cyclin B (CycB) from the FMW to the NPR. Notably, NF-YC knockdown resulted in decreased expression of Eyes absent (Eya) but did not affect Decapentaplegic (Dpp) and Hedgehog (Hh). These findings highlight the role of NF-YC in restricting the expression of CycA and CycB in the NPR, thereby facilitating cell-cycle synchronization. Moreover, this study identified the transcriptional cofactor Eya as a downstream target of NF-YC, revealing a new regulatory pathway in Drosophila eye development. This study expands understanding of NF-YC's role from cell cycle control to encompass developmental processes.
Bar-Cohen, S., Martínez Quiles, M. L., Baskin, A., Dawud, R., Jennings, B. H. and Paroush, Z. (2023). Normal cell cycle progression requires negative regulation of E2F1 by Groucho during S phase and its relief at G2 phase. Development 150(11). PubMed ID: 37260146
Summary:
The cell cycle depends on a sequence of steps that are triggered and terminated via the synthesis and degradation of phase-specific transcripts and proteins. Although much is known about how stage-specific transcription is activated, less is understood about how inappropriate gene expression is suppressed. This study demonstrates that Groucho, the Drosophila orthologue of TLE1 and other related human transcriptional corepressors, regulates normal cell cycle progression in vivo. Although Groucho is expressed throughout the cell cycle, its activity is selectively inactivated by phosphorylation, except in S phase when it negatively regulates E2F1. Constitutive Groucho activity, as well as its depletion and the consequent derepression of e2f1, cause cell cycle phenotypes. The results suggest that Cdk1 contributes to phase-specific phosphorylation of Groucho in vivo. It is proposed that Groucho and its orthologues play a role in the metazoan cell cycle that may explain the links between TLE corepressors and several types of human cancer.
Chakraborty, A., Peterson, N. G., King, J. S., Gross, R. T., Pla, M. M., Thennavan, A., Zhou, K. C., DeLuca, S., Bursac, N., Bowles, D. E., Wolf, M. J. and Fox, D. T. (2023). Conserved chamber-specific polyploidy maintains heart function in Drosophila. Development 150(16). PubMed ID: 37526609
Summary:
Developmentally programmed polyploidy (whole-genome duplication) of cardiomyocytes is common across evolution. Functions of such polyploidy are essentially unknown. In this study, in both Drosophila larvae and human organ donors, distinct polyploidy levels were revealed in cardiac organ chambers. In Drosophila, differential growth and cell cycle signal sensitivity leads the heart chamber to reach a higher ploidy/cell size relative to the aorta chamber. Cardiac ploidy-reduced animals exhibit reduced heart chamber size, stroke volume and cardiac output, and acceleration of circulating hemocytes. These Drosophila phenotypes mimic human cardiomyopathies. These results identify productive and likely conserved roles for polyploidy in cardiac chambers and suggest that precise ploidy levels sculpt many developing tissues. These findings of productive cardiomyocyte polyploidy impact efforts to block developmental polyploidy to improve heart injury recovery.

Tuesday, September 12th - Cytoskeleton

Baez-Cruz, F. A. and Ostap, E. M. (2023). Drosophila class-I myosins that can impact left-right asymmetry have distinct ATPase kinetics. J Biol Chem 299(8): 104961. PubMed ID: 37380077
Summary:
Myosin-1D (myo1D) is important for Drosophila left-right asymmetry, and its effects are modulated by myosin-1C (myo1C). De novo expression of these myosins in nonchiral Drosophila tissues promotes cell and tissue chirality, with handedness depending on the paralog expressed. Remarkably, the identity of the motor domain determines the direction of organ chirality, rather than the regulatory or tail domains. Myo1D, but not myo1C, propels actin filaments in leftward circles in in vitro experiments, but it is not known if this property contributes to establishing cell and organ chirality. To further explore if there are differences in the mechanochemistry of these motors, the ATPase mechanisms of myo1C and myo1D were determined. Myo1D was found to have a 12.5-fold higher actin-activated steady-state ATPase rate, and transient kinetic experiments revealed myo1D has an 8-fold higher MgADP release rate compared to myo1C. Actin-activated phosphate release is rate limiting for myo1C, whereas MgADP release is the rate-limiting step for myo1D. Notably, both myosins have among the tightest MgADP affinities measured for any myosin. Consistent with ATPase kinetics, myo1D propels actin filaments at higher speeds compared to myo1C in in vitro gliding assays. Finally, the ability of both paralogs to transport 50 nm unilamellar vesicles along immobilized actin filaments was tested, and robust transport was found by myo1D and actin binding but no transport by myo1C. These findings support a model where myo1C is a slow transporter with long-lived actin attachments, whereas myo1D has kinetic properties associated with a transport motor.
Bao, M., Dorig, R. E., Vazquez-Pianzola, P. M., Beuchle, D. and Suter, B. (2023). Differential modification of the C-terminal tails of different α-tubulins and their importance for microtubule function in vivo. Elife 12. PubMed ID: 37345829
Summary:
Microtubules (MTs) are built from α-/β-tubulin dimers and used as tracks by kinesin and dynein motors to transport a variety of cargos, such as mRNAs, proteins, and organelles, within the cell. Tubulins are subjected to several post-translational modifications (PTMs). Glutamylation is one of them, and it is responsible for adding one or more glutamic acid residues as branched peptide chains to the C-terminal tails of both α- and β-tubulin. However, very little is known about the specific modifications found on the different tubulin isotypes in vivo and the role of these PTMs in MT transport and other cellular processes in vivo. This study found that in Drosophila ovaries, glutamylation of α-tubulin isotypes occurred clearly on the C-terminal ends of αTub84B and αTub84D (αTub84B/D). In contrast, the ovarian α-tubulin, αTub67C, is not glutamylated. The C-terminal ends of αTub84B/D are glutamylated at several glutamyl sidechains in various combinations. Drosophila TTLL5 is required for the mono- and poly-glutamylation of ovarian αTub84B/D and with this for the proper localization of glutamylated microtubules. Similarly, the normal distribution of kinesin-1 in the germline relies on TTLL5. Next, two kinesin-1-dependent processes, the precise localization of Staufen and the fast, bidirectional ooplasmic streaming, depend on TTLL5, too, suggesting a causative pathway. In the nervous system, a mutation of TTLL5 that inactivates its enzymatic activity decreases the pausing of anterograde axonal transport of mitochondria. These results demonstrate in vivo roles of TTLL5 in differential glutamylation of α-tubulins and point to the in vivo importance of α-tubulin glutamylation for cellular functions involving microtubule transport.
Zapater, I. M. C., Carman, P. J., Soffar, D. B., Windner, S. E., Dominguez, R. and Baylies, M. K. (2023). Drosophila Tropomodulin is required for multiple actin-dependent processes within developing myofibers. Development 150(6). PubMed ID: 36806912
Summary:
Proper muscle contraction requires the assembly and maintenance of sarcomeres and myofibrils. Although the protein components of myofibrils are generally known, less is known about the mechanisms by which they individually function and together synergize for myofibril assembly and maintenance. For example, it is unclear how the disruption of actin filament (F-actin) regulatory proteins leads to the muscle weakness observed in myopathies. This study shows that knockdown of Drosophila Tropomodulin (Tmod), results in several myopathy-related phenotypes, including reduction of muscle cell (myofiber) size, increased sarcomere length, disorganization and misorientation of myofibrils, ectopic F-actin accumulation, loss of tension-mediating proteins at the myotendinous junction, and misshaped and internalized nuclei. These findings support and extend the tension-driven self-organizing myofibrillogenesis model. Like its mammalian counterpart, Drosophila Tmod was shown to cap F-actin pointed-ends, and it is proposed that this activity is crucial for cellular processes in different locations within the myofiber that directly and indirectly contribute to the maintenance of muscle function. These findings provide significant insights to the role of Tmod in muscle development, maintenance and disease.
Niu, X., Mao, C. X., Wang, S., Wang, X., Zhang, Y., Hu, J., Bi, R., Liu, Z. and Shan, J. (2023). α-Tubulin acetylation at lysine 40 regulates dendritic arborization and larval locomotion by promoting microtubule stability in Drosophila. PLoS One 18(2): e0280573. PubMed ID: 36827311
Summary:
Posttranslational modification of tubulin increases the dynamic complexity and functional diversity of microtubules. Acetylation of α-tubulin at Lys-40 is a highly conserved posttranslational modification that has been shown to improve the flexibility and resilience of microtubules. The in vivo functions of α-tubulin acetylation was studied by knocking-out Atat, the Drosophila α-tubulin acetyltransferase, and by mutating Lys-40 to Arg in α1-tubulin. A reduction in the dendritic arborization of larval class I dendritic arborization (da) neurons was found in both mutants. The dendritic developmental defects in atat mutants could be reversed by enhancing the stability of microtubules either through knocking down the microtubule severing protein Katanin 60 or through overexpressing tubulin-specific chaperone E, suggesting that α-tubulin deacetylation impairsed dendritic morphology by decreasing the stability of microtubules. Using time-lapse recordings, this study found that atat and α1-tubulinK40R mutations dramatically increased the number of dendritic protrusions that were likely to be immature dendritic precursors. Finally, it was shown that both Atat and α-tubulin acetylation were required in class I da neurons to control larval locomotion. These findings add novel insight into the current knowledge of the role of α-tubulin acetylation in regulating neuronal development and functions.
Cazzagon, G., Roubinet, C. and Baum, B. (2023). Polarized SCAR and the Arp2/3 complex regulate apical cortical remodeling in asymmetrically dividing neuroblasts. iScience 26(7): 107129. PubMed ID: 37434695
Summary:
Although the formin-nucleated actomyosin cortex has been shown to drive the changes in cell shape that accompany animal cell division in both symmetric and asymmetric cell divisions, the mitotic role of cortical Arp2/3-nucleated actin networks remain unclear. In this study, using asymmetrically dividing Drosophila neural stem cells as a model system, a pool of membrane protrusions was identified that form at the apical cortex of neuroblasts as they enter mitosis. Strikingly, these apically localized protrusions are enriched in SCAR, and depend on SCAR and Arp2/3 complexes for their formation. Because compromising SCAR or the Arp2/3 complex delays the apical clearance of Myosin II at the onset of anaphase and induces cortical instability at cytokinesis, these data point to a role for an apical branched actin filament network in fine-tuning the actomyosin cortex to enable the precise control of cell shape changes during an asymmetric cell division.
Balaghi, N., Erdemci-Tandogan, G., McFaul, C. and Fernandez-Gonzalez, R. (2023). Myosin waves and a mechanical asymmetry guide the oscillatory migration of Drosophila cardiac progenitors. Dev Cell 58(14): 1299-1313. PubMed ID: 37295436
Summary:
Heart development begins with the formation of a tube as cardiac progenitors migrate from opposite sides of the embryo. Abnormal cardiac progenitor movements cause congenital heart defects. However, the mechanisms of cell migration during early heart development remain poorly understood. Using quantitative microscopy, this study found that in Drosophila embryos, cardiac progenitors (cardioblasts) migrated through a sequence of forward and backward steps. Cardioblast steps were associated with oscillatory non-muscle myosin II waves that induced periodic shape changes and were necessary for timely heart tube formation. Mathematical modeling predicted that forward cardioblast migration required a stiff boundary at the trailing edge. Consistent with this, a supracellular actin cable was found at the trailing edge of the cardioblasts that limited the amplitude of the backward steps, thus biasing the direction of cell movement. These results indicate that periodic shape changes coupled with a polarized actin cable produce asymmetrical forces that promote cardioblast migration.

Monday, September 11th - Disease Models

Candia, N., Ibacache, A., Medina-Yanez, I., Olivares, G. H., Ramírez, M., Vega-Macaya, F., Couve, A., Sierralta, J. and Olguín, P. (2023). Identification of atlastin genetic modifiers in a model of hereditary spastic paraplegia in Drosophila. Hum Genet 142(8): 1303-1315. PubMed ID: 37368047
Summary:
Hereditary spastic paraplegias (HSPs) are a group of neurodegenerative disorders characterized by progressive dysfunction of corticospinal motor neurons. Mutations in Atlastin1/Spg3, a small GTPase required for membrane fusion in the endoplasmic reticulum, are responsible for 10% of HSPs. Patients with the same Atlastin1/Spg3 mutation present high variability in age at onset and severity, suggesting a fundamental role of the environment and genetic background. This study used a Drosophila model of HSPs to identify genetic modifiers of decreased locomotion associated with atlastin knockdown in motor neurons. First, a screen was performed for genomic regions that modify the climbing performance or viability of flies expressing atl RNAi in motor neurons. 364 deficiencies spanning chromosomes two and three were tested; 35 enhancer and four suppressor regions of the climbing phenotype were found. Candidate genomic regions could also rescue atlastin effects at synapse morphology, suggesting a role in developing or maintaining the neuromuscular junction. Motor neuron-specific knockdown of 84 genes spanning candidate regions of the second chromosome identified 48 genes required for climbing behavior in motor neurons and 7 for viability, mapping to 11 modifier regions. atl was found to interact genetically with Su(z)2, a component of the Polycomb repressive complex 1, suggesting that epigenetic regulation plays a role in the variability of HSP-like phenotypes caused by atl alleles. These results identify new candidate genes and epigenetic regulation as a mechanism modifying neuronal atl pathogenic phenotypes, providing new targets for clinical studies.
Birker, K., Ge, S., Kirkland, N. J., Theis, J. L., Marchant, J., Fogarty, Z. C., Missinato, M. A., Kalvakuri, S., Grossfeld, P., Engler, A. J., Ocorr, K., Nelson, T. J., Colas, A. R., Olson, T. M., Vogler, G. and Bodmer, R. (2023). Mitochondrial MICOS complex genes, implicated in hypoplastic left heart syndrome, maintain cardiac contractility and actomyosin integrity. Elife 12. PubMed ID: 37404133
Summary:
Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease (CHD) with a likely oligogenic etiology, but understanding of the genetic complexities and pathogenic mechanisms leading to HLHS is limited. This study performed whole genome sequencing (WGS) on 183 HLHS patient-parent trios to identify candidate genes, which were functionally tested in the Drosophila heart model. Bioinformatic analysis of WGS data from an index family of a HLHS proband born to consanguineous parents prioritized 9 candidate genes with rare, predicted damaging homozygous variants. Of them, cardiac-specific knockdown (KD) of mitochondrial MICOS complex subunit dCHCHD3/6 resulted in drastically compromised heart contractility, diminished levels of sarcomeric actin and myosin, reduced cardiac ATP levels, and mitochondrial fission-fusion defects. These defects were similar to those inflicted by cardiac KD of ATP synthase subunits of the electron transport chain (ETC), consistent with the MICOS complex's role in maintaining cristae morphology and ETC assembly. Five additional HLHS probands harbored rare, predicted damaging variants in CHCHD3 or CHCHD6. Hypothesizing an oligogenic basis for HLHS, 60 additional prioritized candidate genes from these patients were tested for genetic interactions with CHCHD3/6 in sensitized fly hearts. Moderate KD of CHCHD3/6 in combination with Cdk12 (activator of RNA polymerase II), RNF149 (goliath, E3 ubiquitin ligase), or SPTBN1 (β-Spectrin, scaffolding protein) caused synergistic heart defects, suggesting the likely involvement of diverse pathways in HLHS. Further elucidation of novel candidate genes and genetic interactions of potentially disease-contributing pathways is expected to lead to a better understanding of HLHS and other CHDs.
Borg, R., Herrera, P., Purkiss, A., Cacciottolo, R. and Cauchi, R. J. (2023). Reduced levels of ALS gene DCTN1 induce motor defects in Drosophila. Front Neurosci 17: 1164251. PubMed ID: 37360176
Summary:
Amyotrophic lateral sclerosis (ALS) is a rapidly progressive neuromuscular disease that has a strong genetic component. Deleterious variants in the DCTN1 gene are known to be a cause of ALS in diverse populations. DCTN1 encodes the p150 subunit of the molecular motor dynactin which is a key player in the bidirectional transport of cargos within cells. Whether DCTN1 mutations lead to the disease through either a gain or loss of function mechanism remains unresolved. Moreover, the contribution of non-neuronal cell types, especially muscle tissue, to ALS phenotypes in DCTN1 carriers is unknown. This study shows that gene silencing of Dctn1, the Drosophila main orthologue of DCTN1, either in neurons or muscles is sufficient to cause climbing and flight defects in adult flies. Dred, a protein with high homology to Drosophila Dctn1 and human DCTN1 was identified, that on loss of function also leads to motoric impairments. A global reduction of Dctn1 induced a significant reduction in the mobility of larvae and neuromuscular junction (NMJ) deficits prior to death at the pupal stage. RNA-seq and transcriptome profiling revealed splicing alterations in genes required for synapse organisation and function, which may explain the observed motor dysfunction and synaptic defects downstream of Dctn1 ablation. These findings support the possibility that loss of DCTN1 function can lead to ALS and underscore an important requirement for DCTN1 in muscle in addition to neurons.
Bhatnagar, A., Parmar, V., Barbieri, N., Bearoff, F., Elefant, F. and Kortagere, S. (2023). Novel EAAT2 activators improve motor and cognitive impairment in a transgenic model of Huntington's disease. Front Behav Neurosci 17: 1176777. PubMed ID: 37351153
Summary:
Glutamate excitotoxicity is causal in striatal neurodegeneration underlying motor dysfunction and cognitive deficits in Huntington's disease (HD). Excitatory amino acid transporter 2 (EAAT2), the predominant glutamate transporter accounting for >90% of glutamate transport, plays a key role in preventing excitotoxicity by clearing excess glutamate from the intrasynaptic cleft. Accordingly, EAAT2 has emerged as a promising therapeutic target for prevention of neuronal excitotoxicity underlying HD and other neurodegenerative diseases. Previously novel EAAT2 positive allosteric modulators were designed, GT951, GTS467, and GTS551, with low nanomolar efficacy in glutamate uptake and favorable pharmacokinetic properties. In this study, the neuroprotective abilities of these novel EAAT2 activators was tested in vivo using the robust Drosophila HD transgenic model expressing human huntingtin gene with expanded repeats (Htt128Q). All three compounds significantly restored motor function impaired under HD pathology over a wide dose range. Additionally, treatment with all three compounds significantly improved HD-associated olfactory associative learning and short-term memory defects, while GT951 and GTS551 also improved middle-term memory in low-performing group. Similarly, treatment with GT951 and GTS551 partially protected against early mortality observed in the HD model. Further, treatment with all three EAAT2 activators induced epigenetic expression of EAAT2 Drosophila homolog and several cognition-associated genes. Together, these results highlight the efficacy of GT951, GTS467 and GTS551 in treating motor and cognitive impairments under HD pathology and support their development for treatment of HD.
Blount, J. R., Patel, N. C., Libohova, K., Harris, A. L., Tsou, W. L., Sujkowski, A. and Todi, S. V. (2023). Lysine 117 on ataxin-3 modulates toxicity in Drosophila models of Spinocerebellar Ataxia Type 3. bioRxiv. PubMed ID: 37398109
Summary:
Ataxin-3 (Atxn3) is a deubiquitinase with a polyglutamine (polyQ) repeat tract whose abnormal expansion causes the neurodegenerative disease, Spinocerebellar Ataxia Type 3 (SCA3; also known as Machado-Joseph Disease). The ubiquitin chain cleavage properties of Atxn3 are enhanced when it is ubiquitinated at lysine (K) at position 117. K117-ubiqutinated Atxn3 cleaves poly-ubiquitin more rapidly in vitro compared to its unmodified counterpart and this residue is also important for Atxn3 roles in cell culture and in Drosophila melanogaster. How polyQ expansion causes SCA3 remains unclear. To gather insight into the biology of disease of SCA3, the following question was posited: is K117 important for toxicity caused by Atxn3? Transgenic Drosophila lines were generated that express full-length, human, pathogenic Atxn3 with 80 polyQ with an intact or mutated K117. K117 mutation mildly enhances the toxicity and aggregation of pathogenic Atxn3 in Drosophila. An additional transgenic line that expresses Atxn3 without any K residues confirms increased aggregation of pathogenic Atxn3 whose ubiquitination is perturbed. These findings suggest Atxn3 ubiquitination as a regulatory step of SCA3, in part by modulating its aggregation.
Campesan, S., Del Popolo, I., Marcou, K., Straatman-Iwanowska, A., Repici, M., Boytcheva, K. V., Cotton, V. E., Allcock, N., Rosato, E., Kyriacou, C. P. and Giorgini, F. (2023). Bypassing mitochondrial defects rescues Huntington's phenotypes in Drosophila. Neurobiol Dis 185: 106236. PubMed ID: 37495179
Summary:
Huntington's disease (HD) is a fatal neurodegenerative disease with limited treatment options. Human and animal studies have suggested that metabolic and mitochondrial dysfunctions contribute to HD pathogenesis. This study used high-resolution respirometry to uncover defective mitochondrial oxidative phosphorylation and electron transfer capacity when a mutant huntingtin fragment is targeted to neurons or muscles in Drosophila and find that enhancing mitochondrial function can ameliorate these defects. In particular, it was found that co-expression of parkin, an E3 ubiquitin ligase critical for mitochondrial dynamics and homeostasis, produces significant enhancement of mitochondrial respiration when expressed either in neurons or muscles, resulting in significant rescue of neurodegeneration, viability and longevity in HD model flies. Targeting mutant HTT to muscles results in larger mitochondria and higher mitochondrial mass, while co-expression of parkin increases mitochondrial fission and decreases mass. Furthermore, directly addressing HD-mediated defects in the fly's mitochondrial electron transport system, by rerouting electrons to either bypass mitochondrial complex I or complexes III-IV, significantly increases mitochondrial respiration and results in a striking rescue of all phenotypes arising from neuronal mutant huntingtin expression. These observations suggest that bypassing impaired mitochondrial respiratory complexes in HD may have therapeutic potential for the treatment of this devastating disorder.

Friday, September 8th - Chromatin and DNA Replication, and Chromosome Dynamics

Wang, H., Langlais, D. and Nijnik, A. (2023). Histone H2A deubiquitinases in the transcriptional programs of development and hematopoiesis: a consolidated analysis. Int J Biochem Cell Biol 157: 106384. PubMed ID: 36738766
Summary:
Monoubiquitinated lysine 119 of histone H2A (H2AK119ub) is a highly abundant epigenetic mark, associated with gene repression and deposited on chromatin by the polycomb repressor complex 1 (PRC1), which is an essential regulator of diverse transcriptional programs in mammalian development and tissue homeostasis. This study addresses H2A-DUB functions in epigenetic regulation of mammalian development and tissue maintenance by conducting a meta-analysis of 248 genomics datasets from 32 independent studies, focusing on the mouse model and covering embryonic stem cells (ESCs), hematopoietic, and immune cell lineages. This covers all the publicly available datasets that map genomic H2A-DUB binding and H2AK119ub distributions (ChIP-Seq), and all datasets assessing dysregulation in gene expression in the relevant H2A-DUB knockout models (RNA-Seq). Many accessory datasets for PRC1-2 and DUB-interacting proteins are also analyzed and interpreted, as well as further data assessing chromatin accessibility (ATAC-Seq) and transcriptional activity (RNA-seq). This study reports co-localization in the binding of H2A-DUBs BAP1, USP16, and to a lesser extent others that is conserved across different cell-types, and also the enrichment of antagonistic PRC1-2 protein complexes at the same genomic locations. Such conserved sites enriched for the H2A-DUBs and PRC1-2 are proximal to transcriptionally active genes that engage in housekeeping cellular functions. Nevertheless, they exhibit H2AK119ub levels significantly above the genomic average that can undergo further increase with H2A-DUB knockout. This indicates a cooperation between H2A-DUBs and PRC1-2 in the modulation of housekeeping transcriptional programs, conserved across many cell types, likely operating through their antagonistic effects on H2AK119ub and the regulation of local H2AK119ub turnover. This study further highlights existing knowledge gaps and discusses important directions for future work.
Brown, J. L., Price, J. D., Erokhin, M. and Kassis, J. A. (2023). Context-dependent role of Pho binding sites in Polycomb complex recruitment in Drosophila. Genetics 224(4). PubMed ID: 37216193
Summary:
Polycomb group (PcG) proteins maintain the silenced state of key developmental genes, but how these proteins are recruited to specific regions of the genome is still not completely understood. In Drosophila, PcG proteins are recruited to Polycomb response elements (PREs) comprised of a flexible array of sites for sequence-specific DNA binding proteins, "PcG recruiters," including Pho, Spps, Cg, and GAF. Pho is thought to play a central role in PcG recruitment. Early data showed that mutation of Pho binding sites in PREs in transgenes abrogated the ability of those PREs to repress gene expression. In contrast, genome-wide experiments in pho mutants or by Pho knockdown showed that PcG proteins can bind to PREs in the absence of Pho. This study directly addressed the importance of Pho binding sites in 2 engrailed (en) PREs at the endogenous locus and in transgenes. The results show that Pho binding sites are required for PRE activity in transgenes with a single PRE. In a transgene, 2 PREs together lead to stronger, more stable repression and confer some resistance to the loss of Pho binding sites. Making the same mutation in Pho binding sites has little effect on PcG-protein binding at the endogenous en gene. Overall, these data support the model that Pho is important for PcG binding but emphasize how multiple PREs and chromatin environment increase the ability of PREs to function in the absence of Pho. This supports the view that multiple mechanisms contribute to PcG recruitment in Drosophila.
Brennan, K. J., Weilert, M., Krueger, S., Pampari, A., Liu, H. Y., Yang, A. W. H., Morrison, J. A., Hughes, T. R., Rushlow, C. A., Kundaje, A. and Zeitlinger, J. (2023). Chromatin accessibility in the Drosophila embryo is determined by transcription factor pioneering and enhancer activation. Dev Cell. PubMed ID: 37557175
Summary:
Chromatin accessibility is integral to the process by which transcription factors (TFs) read out cis-regulatory DNA sequences, but it is difficult to differentiate between TFs that drive accessibility and those that do not. Deep learning models that learn complex sequence rules provide an unprecedented opportunity to dissect this problem. Using zygotic genome activation in Drosophila as a model, this study analyzed high-resolution TF binding and chromatin accessibility data with interpretable deep learning and performed genetic validation experiments. A hierarchical relationship was identified between the pioneer TF Zelda and the TFs involved in axis patterning. Zelda consistently pioneers chromatin accessibility proportional to motif affinity, whereas patterning TFs augment chromatin accessibility in sequence contexts where they mediate enhancer activation. It is concluded that chromatin accessibility occurs in two tiers: one through pioneering, which makes enhancers accessible but not necessarily active, and the second when the correct combination of TFs leads to enhancer activation.
Yheskel, M., Sidoli, S. and Secombe, J. (2023). Proximity labeling reveals a new in vivo network of interactors for the histone demethylase KDM5. Epigenetics Chromatin 16(1): 8. PubMed ID: 36803422
Summary:
KDM5 family proteins are multi-domain regulators of transcription that when dysregulated contribute to cancer and intellectual disability. KDM5 proteins can regulate transcription through their histone demethylase activity in addition to demethylase-independent gene regulatory functions that remain less characterized. To expand understanding of the mechanisms that contribute to KDM5-mediated transcription regulation, TurboID proximity labeling was used to identify KDM5-interacting proteins. Using Drosophila melanogaster, biotinylated proteins were enriched for from KDM5-TurboID-expressing adult heads using a newly generated control for DNA-adjacent background in the form of dCas9:TurboID. Mass spectrometry analyses of biotinylated proteins identified both known and novel candidate KDM5 interactors, including members of the SWI/SNF and NURF chromatin remodeling complexes, the NSL complex, Mediator, and several insulator proteins. Combined, these data shed new light on potential demethylase-independent activities of KDM5. In the context of KDM5 dysregulation, these interactions may play key roles in the alteration of evolutionarily conserved transcriptional programs implicated in human disorders.
Bruckner, D. B., Chen, H., Barinov, L., Zoller, B. and Gregor, T. (2023). Stochastic motion and transcriptional dynamics of pairs of distal DNA loci on a compacted chromosome. Science 380(6652): 1357-1362. PubMed ID: 37384691
Summary:
Chromosomes in the eukaryotic nucleus are highly compacted. However, for many functional processes, including transcription initiation, the pairwise motion of distal chromosomal elements such as enhancers and promoters is essential and necessitates dynamic fluidity. This study used a live-imaging assay to simultaneously measure the positions of pairs of enhancers and promoters and their transcriptional output while systematically varying the genomic separation between these two DNA loci. This analysis reveals the coexistence of a compact globular organization and fast subdiffusive dynamics. These combined features cause an anomalous scaling of polymer relaxation times with genomic separation leading to long-ranged correlations. Thus, encounter times of DNA loci are much less dependent on genomic distance than predicted by existing polymer models, with potential consequences for eukaryotic gene expression.
Salzler, H. R., Vandadi, V., McMichael, B. D., Brown, J. C., Boerma, S. A., Leatham-Jensen, M. P., Adams, K. M., Meers, M. P., Simon, J. M., Duronio, R. J., McKay, D. J. and Matera, A. G. (2023). Distinct roles for canonical and variant histone H3 lysine-36 in Polycomb silencing. Sci Adv 9(9): eadf2451. PubMed ID: 36857457
Summary:
Polycomb complexes regulate cell type-specific gene expression programs through heritable silencing of target genes. Trimethylation of histone H3 lysine 27 (H3K27me3) is essential for this process. Perturbation of H3K36 is thought to interfere with H3K27me3. This study showa that mutants of Drosophila replication-dependent (H3.2(K36R)) or replication-independent (H3.3(K36R)) histone H3 genes generally maintain Polycomb silencing and reach later stages of development. In contrast, combined (H3.3(K36R)H3.2(K36R)) mutants display widespread Hox gene misexpression and fail to develop past the first larval stage. Chromatin profiling revealed that the H3.2(K36R) mutation disrupts H3K27me3 levels broadly throughout silenced domains, whereas these regions are mostly unaffected in H3.3(K36R) animals. Analysis of H3.3 distributions showed that this histone is enriched at presumptive Polycomb response elements located outside of silenced domains but relatively depleted from those inside. It is concluded that H3.2 and H3.3 K36 residues collaborate to repress Hox genes using different mechanisms.

Thursday, September 7th - Adult, Physiololgy, and Metabloism

Amatobi, K. M., Ozbek-Unal, A. G., Schabler, S., Deppisch, P., Helfrich-Forster, C., Mueller, M. J., Wegener, C. and Fekete, A. (2023). The circadian clock is required for rhythmic lipid transport in Drosophila in interaction with diet and photic condition. J Lipid Res: 100417. PubMed ID: 37481037
Summary:
Modern lifestyle is often at odds with endogenously driven rhythmicity, which can lead to circadian disruption and metabolic syndrome. One signature for circadian disruption is a reduced or altered metabolite cycling in the circulating tissue reflecting the current metabolic status. Drosophila is a well-established model in chronobiology, but day-time dependent variations of transport metabolites in the fly circulation are poorly characterized. This study sampled fly hemolymph throughout the day and analysed diacylglycerols (DGs), phosphoethanolamines (PEs) and phosphocholines (PCs) using LC-MS. In wildtype flies kept on sugar-only medium under a light-dark cycle, all transport lipid species showed a synchronized bimodal oscillation pattern with maxima at the beginning and end of the light phase which were impaired in period01 clock mutants. In wildtype flies under constant dark conditions, the oscillation became monophasic with a maximum in the middle of the subjective day. In strong support of clock-driven oscillations, levels of DGs, PEs and PCs peaked once in the middle of the light phase under time-restricted feeding independent of the time of food intake. When wildtype flies were reared on full standard medium, the rhythmic alterations of hemolymph lipid levels were greatly attenuated. These data suggest that the circadian clock aligns daily oscillations of DGs, PEs and PCs in the hemolymph to the anabolic siesta phase, with a strong influence of light on phase and modality. This finding raises the question of whether and to what extent the circadian regulation of transport lipid levels in the hemolymph contributes to the health of the fly.
Dietary compounds activate an insect gustatory receptor on enteroendocrine cells to elicit myosuppressin secretion. Mang, D., Toyama, T., Yamagishi, T., Sun, J., Purba, E. R., Endo, H., Matthews, M. M., Ito, K., Nagata, S. and Sato, R. (2023). Insect Biochem Mol Biol 155: 103927. PubMed ID: 36871864
Summary:
Sensing of midgut internal contents is important for ensuring appropriate hormonal response and digestion following the ingestion of dietary components. Studies in mammals have demonstrated that taste receptors (TRs), a subgroup of G protein-coupled receptors (GPCRs), are expressed in gut enteroendocrine cells (EECs) to sense dietary compounds and regulate the production and/or secretion of peptide hormones. Although progress has been made in identifying expression patterns of gustatory receptors (GRs) in gut EECs, it is currently unknown whether these receptors, which act as ligand-gated ion channels, serve similar functions as mammalian GPCR TRs to elicit hormone production and/or secretion. A Bombyx mori Gr, BmGr6, has been demonstrated to express in cells by oral sensory organs, midgut and nervous system; and to sense isoquercitrin and chlorogenic acid, which are non-nutritional secondary metabolites of host mulberry. This study shows that BmGr6 co-expresses with Bommo-myosuppressin (BMS) in midgut EECs, responds to dietary compounds and is involved in regulation of BMS secretion. The presence of dietary compounds in midgut lumen after food intake resulted in an increase of BMS secretions in hemolymph of both wild-type and BmGr9 knockout larvae, but BMS secretions in BmGr6 knockout larvae decreased relative to wild-type. In addition, loss of BmGr6 led to a significant decrease in weight gain, excrement, hemolymph carbohydrates levels and hemolymph lipid levels. Interestingly, although BMS is produced in both midgut EECs and brain neurosecretory cells (NSCs), BMS levels in tissue extracts suggested that the increase in hemolymph BMS during feeding conditions is primarily due to secretion from midgut EECs. These studies indicate that BmGr6 expressed in midgut EECs responds to the presence of dietary compounds in the lumen by eliciting BMS secretion in B. mori larvae.
Bozkurt, B., Terlemez, G. and Sezgin, E. (2023). Basidiomycota species in Drosophila gut are associated with host fat metabolism. Sci Rep 13(1): 13807. PubMed ID: 37612350
Summary:
The importance of bacterial microbiota on host metabolism and obesity risk is well documented. However, the role of fungal microbiota on host storage metabolite pools is largely unexplored. This study aimed to investigate the role of microbiota on D. melanogaster fat metabolism, and examine interrelatedness between fungal and bacterial microbiota, and major metabolic pools. Fungal and bacterial microbiota profiles, fat, glycogen, and trehalose metabolic pools are measured in a context of genetic variation represented by whole genome sequenced inbred Drosophila Genetic Reference Panel (DGRP) samples. Increasing Basidiomycota, Acetobacter persici, Acetobacter pomorum, and Lactobacillus brevis levels correlated with decreasing triglyceride levels. Host genes and biological pathways, identified via genome-wide scans, associated with Basidiomycota and triglyceride levels were different suggesting the effect of Basidiomycota on fat metabolism is independent of host biological pathways that control fungal microbiota or host fat metabolism. Although triglyceride, glycogen and trehalose levels were highly correlated, microorganisms' effect on triglyceride pool were independent of glycogen and trehalose levels. Multivariate analyses suggested positive interactions between Basidiomycota, A. persici, and L. brevis that collectively correlated negatively with fat and glycogen pools. In conclusion, fungal microbiota can be a major player in host fat metabolism. Interactions between fungal and bacterial microbiota may exert substantial control over host storage metabolite pools and influence obesity risk.
Asiimwe, O. H., Rubaihayo, J., Sulaiman, S. O., Osuwat, L. O. and Kasozi, K. I. (2023). A protein restricted diet induces a stable increased fat storage phenotype in flies. Toxicol Rep 10: 706-713. PubMed ID: 37396850
Summary:
Scientific evidence has revealed possible confounders in diet induced obesity models of Drosophila melanogaster. High Sugar Diet (HSD) induction of obesity in flies has been associated with fly hyperosmolarity and glucotoxicity, while High Fat Diet (HFD) induction has been associated with lipotoxicity. The objective of this study was to assess for a healthy obesity phenotype by comparison of fly survival, physio-chemical and biochemical changes associated with HSD, HFD and Protein Restricted Diet (PRD) obesity induction models of male Drosophila melanogaster. This study provides information on a PRD as the plausible option in obesity research not involving cancer, diabetes, glucotoxicity and lipotoxicity studies. Obesity was induced by exposing Drosophila melanogaster white mutant w(1118) to four experimental diets for four weeks. Group 1 was fed regular food (control), group 2 was fed a 0.5% less yeast than in regular feed (PRD), group 3 was fed a 30% w/v sucrose to regular cornmeal food (HSD) and group 4 was fed a 10% w/v food-grade coconut oil to regular cornmeal food (HFD). Peristaltic waves were measured on 3rd instar larvae of all experimental groups. Negative geotaxis, fly survival, body mass, catalase activity, triglycerides (TG/TP), sterol, and total protein were measured in adult Drosophila melanogaster after four weeks. Triglycerides (TG/TP) and total protein levels were significantly higher in HSD phenotype. Sterols were higher in HFD phenotype. Though catalase enzyme activity was highest in PRD phenotype, this activity was not statistically significant when compared to that of HSD and HFD phenotypes. However, PRD phenotype had the lowest mass, highest survival rate and the highest negative geotaxis, thus demonstrating a balanced, stable and more viable metabolic status in the experimental model. A protein restricted diet induces a stable increased fat storage phenotype in Drosophila melanogaster.
Mariano, V., Kanellopoulos, A. K., Aiello, G., Lo, A. C., Legius, E., Achsel, T. and Bagni, C. (2023). SREBP modulates the NADP(+)/NADPH cycle to control night sleep in Drosophila. Nat Commun 14(1): 763. PubMed ID: 36808152
Summary:
Sleep behavior is conserved throughout evolution, and sleep disturbances are a frequent comorbidity of neuropsychiatric disorders. However, the molecular basis underlying sleep dysfunctions in neurological diseases remains elusive. Using a model for neurodevelopmental disorders (NDDs), the Drosophila Cytoplasmic FMR1 interacting protein haploinsufficiency (Cyfip(85.1/+)), this study identified a mechanism modulating sleep homeostasis. Increased activity of the sterol regulatory element-binding protein (SREBP) in Cyfip(85.1/+) flies induces an increase in the transcription of wakefulness-associated genes, such as the malic enzyme (Men), causing a disturbance in the daily NADP(+)/NADPH ratio oscillations and reducing sleep pressure at the night-time onset. Reduction in SREBP or Men activity in Cyfip(85.1/+) flies enhances the NADP(+)/NADPH ratio and rescues the sleep deficits, indicating that SREBP and Men are causative for the sleep deficits in Cyfip heterozygous flies. This work suggests modulation of the SREBP metabolic axis as a new avenue worth exploring for its therapeutic potential in sleep disorders.
Brand, J. A., Yee, W. K. W., Aitkenhead, I. J., Martin, J. M., Polverino, G., Chown, S. L., Wong, B. B. M. and Dowling, D. K. (2023). Temperature change exerts sex-specific effects on behavioural variation. Proc Biol Sci 290(2002): 20230110. PubMed ID: 37403505
Summary:
Temperature is a key factor mediating organismal fitness and has important consequences for species' ecology. While the mean effects of temperature on behaviour have been well-documented in ectotherms, how temperature alters behavioural variation among and within individuals, and whether this differs between the sexes, remains unclear. Such effects likely have ecological and evolutionary consequences, given that selection acts at the individual level. This study investigated the effect of temperature on individual-level behavioural variation and metabolism in adult male and female Drosophila melanogaster (n = 129), by taking repeated measures of locomotor activity and metabolic rate at both a standard temperature (25°C) and a high temperature (28°C). Males were moderately more responsive in their mean activity levels to temperature change when compared to females. However, this was not true for either standard or active metabolic rate, where no sex differences in thermal metabolic plasticity were found. Furthermore, higher temperatures increased both among- and within-individual variation in male, but not female, locomotor activity. Given that behavioural variation can be critical to population persistence, it is suggest edthat future studies test whether sex differences in the amount of behavioural variation expressed in response to temperature change may result in sex-specific vulnerabilities to a warming climate.

Wednesday, September 6th - Evolution

Wilcox, A. S., Vea, I. M., Frankino, W. A. and Shingleton, A. W. (2023). Genetic variation of morphological scaling in Drosophila melanogaster. Heredity (Edinb). PubMed ID: 36878946
Summary:
Morphological scaling relationships between the sizes of individual traits and the body captures the characteristic shape of a species, and their evolution is the primary mechanism of morphological diversification. However, almost no knowledge is available of the genetic variation of scaling, which is critical if how scaling evolves is to be understood. This study explored the genetics of population scaling relationships (scaling relationships fit to multiple genetically-distinct individuals in a population) by describing the distribution of individual scaling relationships (genotype-specific scaling relationships that are unseen or cryptic). These individual scaling relationships harbor the genetic variation in the developmental mechanisms that regulate trait growth relative to body growth, and theoretical studies suggest that their distribution dictates how the population scaling relationship will respond to selection. Using variation in nutrition to generate size variation within 197 isogenic lineages of Drosophila melanogaster, extensive variation was revealed in the slopes of the wing-body and leg-body individual scaling relationships among genotypes. This variation reflects variation in the nutritionally-induced size plasticity of the wing, leg, and body. Surprisingly, it was found that variation in the slope of individual scaling relationships primarily results from variation in nutritionally-induced plasticity of body size, not leg or wing size. These data allow prediction of how different selection regimes affect scaling in Drosophila, and is the first step in identifying the genetic targets of such selection. More generally, this approach provides a framework for understanding the genetic variation of scaling, an important prerequisite to explaining how selection changes scaling and morphology.
Blunk, S., Garcia-Verdugo, H., O'Sullivan, S., Camp, J., Haines, M., Coalter, T., Williams, T. A. and Nagy, L. M. (2023). Functional Divergence of the Tribolium castaneum engrailed and invected Paralogs. Insects 14(8). PubMed ID: 37623401
Summary:
Engrailed (en) and invected (inv) encode paralogous transcription factors found as a closely linked tandem duplication within holometabolous insects. Drosophila en mutants segment normally, then fail to maintain their segments. Loss of Drosophila inv is viable, while loss of both genes results in asegmental larvae. Surprisingly, the knockdown of Oncopeltus inv can result in the loss or fusion of the entire abdomen and en knockdowns in Tribolium show variable degrees of segmental loss. The consequence of losing or knocking down both paralogs on embryogenesis has not been studied beyond Drosophila. To further investigate the relative functions of each paralog and the mechanism behind the segmental loss, Tribolium double and single knockdowns of en and inv were analyzed. The most common cuticular phenotype of the double knockdowns was small, limbless, and open dorsally, with all but a single, segmentally iterated row of bristles. Less severe knockdowns had fused segments and reduced appendages. The Tribolium paralogs appear to act synergistically: the knockdown of either Tribolium gene alone was typically less severe, with all limbs present, whereas the most extreme single knockdowns mimic the most severe double knockdown phenotype. Morphological abnormalities unique to either single gene knockdown were not found. inv expression was not affected in the Tribolium en knockdowns, but hh expression was unexpectedly increased midway through development. Thus, while the segmental expression of en/inv is broadly conserved within insects, the functions of en and inv are evolving independently in different lineages.
Andrianova, E. P., Marmion, R. A., Shvartsman, S. Y. and Zhulin, I. B. (2023). Evolutionary history of MEK1 illuminates the nature of deleterious mutations. Proc Natl Acad Sci U S A 120(34): e2304184120. PubMed ID: 37579140
Summary:
Mutations in signal transduction pathways lead to various diseases including cancers. MEK1 kinase, encoded by the human MAP2K1 gene, is one of the central components of the MAPK pathway and more than a hundred somatic mutations in the MAP2K1 gene were identified in various tumors. Germline mutations deregulating MEK1 also lead to congenital abnormalities, such as the cardiofaciocutaneous syndrome and arteriovenous malformation. Evaluating variants associated with a disease is a challenge, and computational genomic approaches aid in this process. Establishing evolutionary history of a gene improves computational prediction of disease-causing mutations; however, the evolutionary history of MEK1 is not well understood. In this study, by revealing a precise evolutionary history of MEK1, a well-defined dataset of MEK1 metazoan orthologs was construct, that provides sufficient depth to distinguish between conserved and variable amino acid positions. Known and predicted disease-causing and benign mutations were matched to evolutionary changes observed in corresponding amino acid positions and found that all known and many suspected disease-causing mutations are evolutionarily intolerable. Several variants were selected that cannot be unambiguously assessed by automated prediction tools but that are confidently identified as "damaging" by this approach, for experimental validation in Drosophila. In all cases, evolutionary intolerant variants caused increased mortality and severe defects in fruit fly embryos confirming their damaging nature. It is anticipated that this analysis will serve as a blueprint to help evaluate known and novel missense variants in MEK1 and that this approach will contribute to improving automated tools for disease-associated variant interpretation.
Schang, K., Garant, R. and Long, T. A. F. (2023). Phenotypic extremes or extreme phenotypes? On the use of large and small-bodied "phenocopied" Drosophila melanogaster males in studies of sexual selection and conflict. Curr Res Insect Sci 3: 100052. PubMed ID: 36794123
Summary:
In the fruit fly, Drosophila melanogaster, variation in body size is influenced by a number of different factors and may be strongly associated with individual condition, performance and success in reproductive competitions. Consequently, intra-sexual variation in size in this model species has been frequently explored in order to better understand how sexual selection and sexual conflict may operate and shape evolutionary trajectories. However, measuring individual flies can often be logistically complicated and inefficient, which can result in limited sample sizes. Instead, many experiments use large and/or small body sizes that are created by manipulating the developmental conditions experienced during the larval stages, resulting in "phenocopied" flies whose phenotypes resemble what is seen at the extremes of a population's size distribution. While this practice is fairly common, there has been remarkedly few direct tests to empirically compare the behaviour or performance of phenocopied flies to similarly-sized individuals that grew up under typical developmental conditions. Contrary to assumptions that phenocopied flies are reasonable approximations, this study found that both large and small-bodied phenocopied males frequently differed from their standard development equivalents in their mating frequencies, their lifetime reproductive successes, and in their effects on the fecundity of the females they interacted with. These results highlight the complicated contributions of environment and genotype to the expression of body size phenotypes and lead the authors to strongly urge caution in the interpretation of studies solely replying upon phenocopied individuals.
Kutzer, M. A. M., Gupta, V., Neophytou, K., Doublet, V., Monteith, K. M. and Vale, P. F. (2023). Intraspecific genetic variation in host vigour, viral load and disease tolerance during Drosophila C virus infection. Open Biol 13(3): 230025. PubMed ID: 36854375
Summary:
Genetic variation for resistance and disease tolerance has been described in a range of species. In Drosophila melanogaster, genetic variation in mortality following systemic Drosophila C virus (DCV) infection is driven by large-effect polymorphisms in the restriction factor pastrel (pst). However, it is unclear if pst contributes to disease tolerance. Systemic DCV challenges spanning nine orders of magnitude were investigated in males and females of 10 Drosophila Genetic Reference Panel lines carrying either a susceptible (S) or resistant (R) pst allele. Among-line variation in fly survival, viral load and disease tolerance were measured both as the ability to maintain survival (mortality tolerance) and reproduction (fecundity tolerance). This study further uncover novel effects of pst on host vigour, as flies carrying the R allele exhibited higher survival and fecundity even in the absence of infection. Finally, significant genetic variation was found in the expression of the JAK-STAT ligand upd3 and the epigenetic regulator of JAK-STAT G9a. However, while G9a has been previously shown to mediate tolerance of DCV infection, this study found no correlation between the expression of either upd3 or G9a on fly tolerance or resistance. This work highlights the importance of both resistance and tolerance in viral defence.
Veenstra, J. A. (2023). Different neuroendocrine cell types in the pars intercerebralis of Periplaneta americana produce their own specific IGF-related peptides. Gen Comp Endocrinol 335: 114233. PubMed ID: 36791825
Summary:
Of the nine genes of the American cockroach, Periplaneta americana, coding for peptides related to insulin and insulin-like growth factor, seven show significant expression in the central nervous system as demonstrated by the polymerase chain reaction on reverse transcribed RNA. In situ hybridisation shows that five of those are expressed by cells in the pars intercerebralis. Antisera raised to the predicted peptides show that these cells are neuroendocrine in nature and project to the corpora cardiaca. Interestingly, there are at least three cell types that each express different genes. This contrasts with Drosophila where a single cell type expresses a number of genes expressing several such peptides. Whereas in Drosophila the neuroendocrine cells producing insulin-like peptides also express sulfakinins, the arthropod orthologs of gastrin and cholecystokinin, in Periplaneta the sulfakinins are produced by different cells. Other neuropeptides known to be produced by the pars intercerebralis in Periplaneta and other insect species, such as the CRF-like diuretic hormone, neuroparsin, leucokinin or myosuppressin, neither colocalize with an insulin-related peptide. The separate cellular localization of these peptides and the existence of multiple insulin receptors in this species implies a more complex regulation by insulin and IGF-related peptides in cockroaches than in the fruit fly.

Tuesday, September 5th - Adult neural development and function

Au, D. D., Liu, J. C., Park, S. J., Nguyen, T. H., Dimalanta, M., Foden, A. J. and Holmes, T. C. (2023). Drosophila photoreceptor systems converge in arousal neurons and confer light responsive robustness. Front Neurosci 17: 1160353. PubMed ID: 37274190
Summary:
Lateral ventral neurons (LNvs) in the fly circadian neural circuit mediate behaviors other than clock resetting, including light-activated acute arousal. Converging sensory inputs often confer functional redundancy. The LNvs have three distinct light input pathways: (1) cell autonomously expressed cryptochrome (CRY), (2) rhodopsin 7 (Rh7), and (3) synaptic inputs from the eyes and other external photoreceptors that express opsins and CRY. This study explored the relative photoelectrical and behavioral input contributions of these three photoreceptor systems to determine their functional impact in flies. Patch-clamp electrophysiology measuring light evoked firing frequency (FF) was performed on large LNvs (l-LNvs) in response to UV (365 nm), violet (405 nm), blue (450 nm), or red (635 nm) LED light stimulation, testing controls versus mutants that lack photoreceptor inputs gl60j, cry-null, rh7-null, and double mutant gl60j-cry-null flies. For UV, violet, and blue short wavelength light inputs, all photoreceptor mutants show significantly attenuated action potential FF responses measured in the l-LNv. In contrast, red light FF responses are only significantly attenuated in double mutant gl60j-cry-null flies. A light-pulse arousal assay was used to compare behavioral responses to UV, violet, blue and red light of control and light input mutants, measuring the awakening arousal response of flies during subjective nighttime at two different intensities to capture potential threshold differences (10 and 400 μW/cm(2)). The light arousal behavioral results are similar to the electrophysiological results, showing significant attenuation of behavioral light responses for mutants compared to control. These results show that the different LNv convergent photoreceptor systems are integrated and together confer functional redundancy for light evoked behavioral arousal.
Armour, E. M., Thomas, C. M., Greco, G., Bhatnagar, A. and Elefant, F. (2023). Experience-dependent Tip60 nucleocytoplasmic transport is regulated by its NLS/NES sequences for neuroplasticity gene control. Mol Cell Neurosci: 103888. PubMed ID: 37598897
Summary:
Nucleocytoplasmic transport (NCT) in neurons is critical for enabling proteins to enter the nucleus and regulate plasticity genes in response to environmental cues. Such experience-dependent (ED) neural plasticity is central for establishing memory formation and cognitive function and can influence the severity of neurodegenerative disorders like Alzheimer's disease (AD). ED neural plasticity is driven by histone acetylation (HA) mediated epigenetic mechanisms that regulate dynamic activity-dependent gene transcription profiles in response to neuronal stimulation. Yet, how histone acetyltransferases (HATs) respond to extracellular cues in the in vivo brain to drive HA-mediated activity-dependent gene control remains unclear. It has been previously demonstrated that extracellular stimulation of rat hippocampal neurons in vitro triggers Tip60 HAT nuclear import with concomitant synaptic gene induction. This study focused on investigating Tip60 HAT subcellular localization and NCT specifically in neuronal activity-dependent gene control by using the learning and memory mushroom body (MB) region of the Drosophila brain as a powerful in vivo cognitive model system. Immunohistochemistry (IHC) was used to compare the subcellular localization of Tip60 HAT in the Drosophila brain under normal conditions and in response to stimulation of fly brain neurons in vivo either by genetically inducing potassium channels activation or by exposure to natural positive ED conditions. Furthermore, this study found that both inducible and ED condition-mediated neural induction triggered Tip60 nuclear import with concomitant induction of previously identified Tip60 target genes and that Tip60 levels in both the nucleus and cytoplasm were significantly decreased in the well-characterized Drosophila AD model. Mutagenesis of a putative nuclear localization signal (NLS) sequence and nuclear export signal (NES) sequence that this this study identified in the Drosophila Tip60 protein revealed that both are functionally required for appropriate Tip60 subcellular localization. These results support a model by which neuronal stimulation triggers Tip60 NCT via its NLS and NES sequences to promote induction of activity-dependent neuroplasticity gene transcription and that this process may be disrupted in AD.
Zhao, J., Zhang, X., Zhao, B., Hu, W., Diao, T., Wang, L., Zhong, Y. and Li, Q. (2023). Genetic dissection of mutual interference between two consecutive learning tasks in Drosophila. Elife 12. PubMed ID: 36897069
Summary:
Animals can continuously learn different tasks to adapt to changing environments and, therefore, have strategies to effectively cope with inter-task interference, including both proactive interference (Pro-I) and retroactive interference (Retro-I). Many biological mechanisms are known to contribute to learning, memory, and forgetting for a single task, however, mechanisms involved only when learning sequential different tasks are relatively poorly understood. This study dissected the respective molecular mechanisms of Pro-I and Retro-I between two consecutive associative learning tasks in Drosophila. Pro-I is more sensitive to an inter-task interval (ITI) than Retro-I. They occur together at short ITI (<0 min), while only Retro-I remains significant at ITI beyond 20 min. Acutely overexpressing Corkscrew (CSW), an evolutionarily conserved protein tyrosine phosphatase SHP2, in mushroom body (MB) neurons reduces Pro-I, whereas acute knockdown of CSW exacerbates Pro-I. Such function of CSW is further found to rely on the γ subset of MB neurons and the downstream Raf/MAPK pathway. In contrast, manipulating CSW does not affect Retro-I as well as a single learning task. Interestingly, manipulation of Rac1, a molecule that regulates Retro-I, does not affect Pro-I. Thus, these findings suggest that learning different tasks consecutively triggers distinct molecular mechanisms to tune proactive and retroactive interference.
An, H., Yu, Y., Ren, X., Zeng, M., Bai, Y., Liu, T., Zheng, H., Sang, R., Zhang, F., Cai, Y. and Xi, Y. (2023). Pipsqueak family genes dan/danr antagonize nuclear Pros to prevent neural stem cell aging in Drosophila larval brains. Front Mol Neurosci 16: 1160222. PubMed ID: 37266371
Summary:
Neural stem cell aging is a fundamental question in neurogenesis. Premature nuclear Prospero (Pros) is considered as an indicator of early neural stem cell aging in Drosophila. The underlying mechanism of how neural stem cells prevent premature nuclear Prospero (Pros) remains largely unknown. This study identified that two pipsqueak family genes, distal antenna (dan) and distal antenna-related (danr), promote the proliferation of neural stem cells (also called neuroblasts, NBs) in third instar larval brains. In the absence of Dan and Danr (dan/danr), the NBs produce fewer daughter cells with smaller lineage sizes. The larval brain NBs in dan/danr clones show premature accumulation of nuclear Pros, which usually appears in the terminating NBs at early pupal stage. The premature nuclear Pros leads to NBs cell cycle defects and NB identities loss. Removal of Pros from dan/danr MARCM clones prevents lineage size shrinkage and rescues the loss of NB markers. It is proposed that the timing of nuclear Pros is after the downregulation of dan/danr in the wt terminating NBs. dan/danr and nuclear Pros are mutually exclusive in NBs. In addition, dan/danr are also required for the late temporal regulator, Grainyhead (Grh), in third instar larval brains. This study uncovers the novel function of dan/danr in NBs cell fate maintenance. dan/danr antagonize nuclear Pros to prevent NBs aging in Drosophila larval brains.
Bengochea, M., Sitt, J. D., Izard, V., Preat, T., Cohen, L. and Hassan, B. A. (2023). Numerical discrimination in Drosophila melanogaster. Cell Rep 42(7): 112772. PubMed ID: 37453418
Summary:
Sensitivity to numbers is a crucial cognitive ability. The lack of experimental models amenable to systematic genetic and neural manipulation has precluded discovering neural circuits required for numerical cognition. This study demonstrates that Drosophila flies spontaneously prefer sets containing larger numbers of objects. This preference is determined by the ratio between the two numerical quantities tested, a characteristic signature of numerical cognition across species. Individual flies maintained their numerical choice over consecutive days. Using a numerical visual conditioning paradigm, it was found that flies are capable of associating sucrose with numerical quantities and can be trained to reverse their spontaneous preference for large quantities. Finally, this study showed that silencing lobula columnar neurons (LC11) reduces the preference for more objects, thus identifying a neuronal substrate for numerical cognition in invertebrates. This discovery paves the way for the systematic analysis of the behavioral and neural mechanisms underlying the evolutionary conserved sensitivity to numerosity.
Ahmed, M., Rajagopalan, A. E., Pan, Y., Li, Y., Williams, D. L., Pedersen, E. A., Thakral, M., Previero, A., Close, K. C., Christoforou, C. P., Cai, D., Turner, G. C. and Clowney, E. J. (2023). Input density tunes Kenyon cell sensory responses in the Drosophila mushroom body. Curr Biol 33(13): 2742-2760.e2712. PubMed ID: 37348501
Summary:
The ability to discriminate sensory stimuli with overlapping features is thought to arise in brain structures called expansion layers, where neurons carrying information about sensory features make combinatorial connections onto a much larger set of cells. For 50 years, expansion coding has been a prime topic of theoretical neuroscience, which seeks to explain how quantitative parameters of the expansion circuit influence sensory sensitivity, discrimination, and generalization. This study investigated the developmental events that produce the quantitative parameters of the arthropod expansion layer, called the mushroom body. Using Drosophila melanogaster as a model, this study employ genetic and chemical tools to engineer changes to circuit development. These allow production of living animals with hypothesis-driven variations on natural expansion layer wiring parameters. Then the functional and behavioral consequences were tested. By altering the number of expansion layer neurons (Kenyon cells) and their dendritic complexity, it was found that input density, but not cell number, tunes neuronal odor selectivity. Simple odor discrimination behavior is maintained when the Kenyon cell number is reduced and augmented by Kenyon cell number expansion. Animals with increased input density to each Kenyon cell show increased overlap in Kenyon cell odor responses and become worse at odor discrimination tasks. Home page: The Interactive Fly© 2020 Thomas B. Brody, Ph.D.

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