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PloS One 2017Chorea-Acanthocytosis is a rare, neurodegenerative disorder characterized by progressive loss of locomotor and cognitive function. It is caused by loss of function...
Chorea-Acanthocytosis is a rare, neurodegenerative disorder characterized by progressive loss of locomotor and cognitive function. It is caused by loss of function mutations in the Vacuolar Protein Sorting 13A (VPS13A) gene, which is conserved from yeast to human. The consequences of VPS13A dysfunction in the nervous system are still largely unspecified. In order to study the consequences of VPS13A protein dysfunction in the ageing central nervous system we characterized a Drosophila melanogaster Vps13 mutant line. The Drosophila Vps13 gene encoded a protein of similar size as human VPS13A. Our data suggest that Vps13 is a peripheral membrane protein located to endosomal membranes and enriched in the fly head. Vps13 mutant flies showed a shortened life span and age associated neurodegeneration. Vps13 mutant flies were sensitive to proteotoxic stress and accumulated ubiquitylated proteins. Levels of Ref(2)P, the Drosophila orthologue of p62, were increased and protein aggregates accumulated in the central nervous system. Overexpression of the human Vps13A protein in the mutant flies partly rescued apparent phenotypes. This suggests a functional conservation of human VPS13A and Drosophila Vps13. Our results demonstrate that Vps13 is essential to maintain protein homeostasis in the larval and adult Drosophila brain. Drosophila Vps13 mutants are suitable to investigate the function of Vps13 in the brain, to identify genetic enhancers and suppressors and to screen for potential therapeutic targets for Chorea-Acanthocytosis.
Topics: Animals; Brain; Drosophila; Drosophila Proteins; Homeostasis; Humans; Mutation; Nerve Tissue Proteins; Vesicular Transport Proteins
PubMed: 28107480
DOI: 10.1371/journal.pone.0170106 -
Biology Open Jul 2022The compartmentalized domains of polarized epithelial cells arise from mutually antagonistic actions between the apical Par complex and the basolateral Scrib module. In...
The compartmentalized domains of polarized epithelial cells arise from mutually antagonistic actions between the apical Par complex and the basolateral Scrib module. In Drosophila, the Scrib module proteins Scribble (Scrib) and Discs-large (Dlg) are required to limit Lgl phosphorylation at the basolateral cortex, but how Scrib and Dlg could carry out such a 'protection' activity is not clear. We tested Protein Phosphatase 1α (PP1) as a potential mediator of this activity, but demonstrate that a significant component of Scrib and Dlg regulation of Lgl is PP1 independent, and found no evidence for a Scrib-Dlg-PP1 protein complex. However, the Dlg SH3 domain plays a role in Lgl protection and, in combination with the N-terminal region of the Dlg HOOK domain, in recruitment of Scrib to the membrane. We identify a 'minimal Dlg' comprised of the SH3 and HOOK domains that is both necessary and sufficient for Scrib localization and epithelial polarity function in vivo. This article has an associated First Person interview with the first author of the paper.
Topics: Animals; Drosophila; Drosophila Proteins; Epithelial Cells; Humans
PubMed: 35722710
DOI: 10.1242/bio.059408 -
Methods in Enzymology 2015The study of circadian behavior in model organisms is almost exclusively confined to the laboratory, where rhythmic phenotypes are studied under highly simplified... (Review)
Review
The study of circadian behavior in model organisms is almost exclusively confined to the laboratory, where rhythmic phenotypes are studied under highly simplified conditions such as constant darkness or rectangular light-dark cycles. Environmental cycles in nature are far more complex, and recent work in rodents and flies has revealed that when placed in natural/seminatural situations, circadian behavior shows unexpected features that are not consistent with laboratory observations. In addition, the recent observations of clockless mutants, both in terms of their circadian behavior and their Darwinian fitness, challenge some of the traditional beliefs derived from laboratory studies about what constitutes an adaptive circadian phenotype. Here, we briefly summarize the results of these newer studies and then describe how Drosophila behavior can be studied in the wild, pointing out solutions to some of the technical problems associated with extending locomotor monitoring to this unpredictable environment. We also briefly describe how to generate sophisticated simulations of natural light and temperature cycles that can be used to successfully mimic the fly's natural circadian behavior. We further clarify some misconceptions that have been raised in recent studies of natural fly behavior and show how these can be overcome with appropriate methodology. Finally, we describe some recent technical developments that will enhance the naturalistic study of fly circadian behavior.
Topics: Animals; Circadian Rhythm; Drosophila Proteins; Drosophila melanogaster; Gene Expression; Genetic Association Studies; Genetic Testing; Mutation; Phenotype
PubMed: 25662454
DOI: 10.1016/bs.mie.2014.10.001 -
Cell Death and Differentiation Jan 2022The Drosophila IAP protein, Diap2, is a key mediator of NF-κB signalling and innate immune responses. Diap2 is required for both local immune activation, taking place...
The Drosophila IAP protein, Diap2, is a key mediator of NF-κB signalling and innate immune responses. Diap2 is required for both local immune activation, taking place in the epithelial cells of the gut and trachea, and for mounting systemic immune responses in the cells of the fat body. We have found that transgenic expression of Diap2 leads to a spontaneous induction of NF-κB target genes, inducing chronic inflammation in the Drosophila midgut, but not in the fat body. Drice is a Drosophila effector caspase known to interact and form a stable complex with Diap2. We have found that this complex formation induces its subsequent degradation, thereby regulating the amount of Diap2 driving NF-κB signalling in the intestine. Concordantly, loss of Drice activity leads to accumulation of Diap2 and to chronic intestinal inflammation. Interestingly, Drice does not interfere with pathogen-induced signalling, suggesting that it protects from immune responses induced by resident microbes. Accordingly, no inflammation was detected in transgenic Diap2 flies and Drice-mutant flies reared in axenic conditions. Hence, we show that Drice, by restraining Diap2, halts unwanted inflammatory signalling in the intestine.
Topics: Animals; Drosophila; Drosophila Proteins; Immunity, Innate; Inflammation; Inhibitor of Apoptosis Proteins; Signal Transduction
PubMed: 34262145
DOI: 10.1038/s41418-021-00832-w -
Neuron Mar 2022The nervous and endocrine systems coordinately monitor and regulate nutrient availability to maintain energy homeostasis. Sensory detection of food regulates internal...
The nervous and endocrine systems coordinately monitor and regulate nutrient availability to maintain energy homeostasis. Sensory detection of food regulates internal nutrient availability in a manner that anticipates food intake, but sensory pathways that promote anticipatory physiological changes remain unclear. Here, we identify serotonergic (5-HT) neurons as critical mediators that transform gustatory detection by sensory neurons into the activation of insulin-producing cells and enteric neurons in Drosophila. One class of 5-HT neurons responds to gustatory detection of sugars, excites insulin-producing cells, and limits consumption, suggesting that they anticipate increased nutrient levels and prevent overconsumption. A second class of 5-HT neurons responds to gustatory detection of bitter compounds and activates enteric neurons to promote gastric motility, likely to stimulate digestion and increase circulating nutrients upon food rejection. These studies demonstrate that 5-HT neurons relay acute gustatory detection to divergent pathways for longer-term stabilization of circulating nutrients.
Topics: Animals; Drosophila Proteins; Drosophila melanogaster; Nutrients; Serotonergic Neurons; Taste
PubMed: 35051377
DOI: 10.1016/j.neuron.2021.12.028 -
Developmental Cell Sep 2023The eukaryotic genome is organized to enable the precise regulation of gene expression. This organization is established as the embryo transitions from a fertilized...
The eukaryotic genome is organized to enable the precise regulation of gene expression. This organization is established as the embryo transitions from a fertilized gamete to a totipotent zygote. To understand the factors and processes that drive genomic organization, we focused on the pioneer factor GAGA factor (GAF) that is required for early development in Drosophila. GAF transcriptionally activates the zygotic genome and is localized to subnuclear foci. This non-uniform distribution is driven by binding to highly abundant GA repeats. At GA repeats, GAF is necessary to form heterochromatin and silence transcription. Thus, GAF is required to establish both active and silent regions. We propose that foci formation enables GAF to have opposing transcriptional roles within a single nucleus. Our data support a model in which the subnuclear concentration of transcription factors acts to organize the nucleus into functionally distinct domains essential for the robust regulation of gene expression.
Topics: Animals; DNA; Drosophila; Drosophila Proteins; Gene Expression Regulation, Developmental; Genome; Transcription Factors; Zygote
PubMed: 37478844
DOI: 10.1016/j.devcel.2023.06.010 -
Insect Biochemistry and Molecular... Jun 2021The Seipin protein is a conserved key component in the biogenesis of lipid droplets (LDs). Recently, a cooperation between human Seipin and the Lipid droplet assembly...
The Seipin protein is a conserved key component in the biogenesis of lipid droplets (LDs). Recently, a cooperation between human Seipin and the Lipid droplet assembly factor 1 (LDAF1) was described. LDAF1 physically interacts with Seipin and the holocomplex safeguards regular LD biogenesis. The function of LDAF1 proteins outside mammals is less clear. In yeast, the lipid droplet organization (LDO) proteins, which also cooperate with Seipin, are the putative homologs of LDAF1. While certain functional aspects are shared between the LDO and mammalian LDAF1 proteins, the relationship between the proteins is under debate. Here, we identify the Drosophila melanogaster protein CG32803, which we re-named to dmLDAF1, as an insect member of this protein family. dmLDAF1 decorates LDs in cultured cells and in vivo and the protein is linked to the fly and mouse Seipin proteins. Altering the dmLDAF1 abundance affects LD size, number and overall lipid storage amounts. Our results suggest that the LDAF1 proteins thus fulfill an evolutionarily conserved function in the biogenesis and biology of LDs.
Topics: Animals; Drosophila Proteins; Drosophila melanogaster; GTP-Binding Protein gamma Subunits; Humans; Lipid Droplets; Lipid Metabolism; Membrane Proteins
PubMed: 33307187
DOI: 10.1016/j.ibmb.2020.103512 -
Methods in Enzymology 2016The cytoskeleton is a dynamic network of filamentous protein polymers required for virtually all cellular processes. It consists of three major classes, filamentous...
The cytoskeleton is a dynamic network of filamentous protein polymers required for virtually all cellular processes. It consists of three major classes, filamentous actin (F-actin), intermediate filaments, and microtubules, all displaying characteristic structural properties, functions, cellular distributions, and sets of interacting regulatory proteins. One unique class of proteins, the spectraplakins, bind, regulate, and integrate the functions of all three classes of cytoskeleton proteins. Spectraplakins are giant, evolutionary conserved multidomain proteins (spanning up to 9000 aa) that are true members of the plakin, spectrin, and Gas2-like protein families. They have OMIM-listed disease links to epidermolysis bullosa and hereditary sensory and autonomic neuropathy. Their role in disease is likely underrepresented since studies in model animal systems have revealed critical roles in polarity, morphogenesis, differentiation and maintenance, migration, signaling, and intracellular trafficking in a variety of tissues. This enormous diversity of spectraplakin function is consistent with the numerous isoforms produced from single genomic loci that combine different sets of functional domains in distinct cellular contexts. To study the broad range of functions and complexity of these proteins, Drosophila is a powerful model. Thus, the fly spectraplakin Short stop (Shot) acts as an actin-microtubule linker and plays important roles in many developmental processes, which provide experimentally amenable and relevant contexts in which to study spectraplakin functions. For these studies, a versatile range of relevant experimental resources that facilitate genetics and transgenic approaches, highly refined genomics tools, and an impressive set of spectraplakin-specific genetic and molecular tools are readily available. Here, we use the example of Shot to illustrate how the various tools and strategies available for Drosophila can be employed to decipher and dissect cellular roles and molecular mechanisms of spectraplakins.
Topics: Animals; Cell Line; Drosophila; Drosophila Proteins; Mice; Microfilament Proteins; NIH 3T3 Cells; Primary Cell Culture
PubMed: 26778568
DOI: 10.1016/bs.mie.2015.06.022 -
PloS One 2019Spermatogenesis in Drosophila melanogaster is characterized by a specific transcriptional program during the spermatocyte stage. Transcription of thousands of genes is...
Spermatogenesis in Drosophila melanogaster is characterized by a specific transcriptional program during the spermatocyte stage. Transcription of thousands of genes is regulated by the interaction of several proteins or complexes, including a tTAF-containing TFIID variant, tMAC, Mediator, and chromatin interactors, e.g., bromodomain proteins. We addressed how distinct subsets of target genes are selected. We characterized the highly similar proteins tPlus3a and tPlus3b, which contain a Plus3 domain and are enriched in the testis, mainly in spermatocytes. In tPlus3a and tplus3b deletion mutants generated using the CRISPR/Cas9 system, fertility was severely reduced and sperm showed defects during individualization. tPlus3a and tPlus3b heterodimerized with the bromodomain protein tBRD-1. To elucidate the role of the tPlus3a and tPlus3b proteins in transcriptional regulation, we determined the transcriptomes of tplus3a-tplus3b and tbrd-1 deletion mutants using next-generation sequencing (RNA-seq) and compared them to that of the wild-type. tPlus3a and tPlus3b positively or negatively regulated the expression of nearly 400 genes; tBRD-1 regulated 1,500 genes. Nearly 200 genes were regulated by both tPlus3a and tPlus3b and tBRD-1. tPlus3a and tPlus3b activated the Y-chromosomal genes kl-3 and kl-5, which indicates that tPlus3a and tPlus3b proteins are required for the function of distinct classes of genes. tPlus3a and tPlus3b and tBRD-1 repress genes relevant for seminal fluid and heat shock. We hypothesize that tPlus3a and tPlus3b proteins are required to specify the general transcriptional program in spermatocytes.
Topics: Animals; Dimerization; Drosophila Proteins; Drosophila melanogaster; Fertility; Heat-Shock Proteins; High-Throughput Nucleotide Sequencing; Male; RNA Interference; Sequence Analysis, RNA; Spermatocytes; Transcription, Genetic; Y Chromosome
PubMed: 30845228
DOI: 10.1371/journal.pone.0213177 -
Pathology, Research and Practice Mar 2023Hippo pathway has been initially recognized as a regulatory mechanism for modulation of organ size in fruitfly. Subsequently, its involvement in the regulation of... (Review)
Review
Hippo pathway has been initially recognized as a regulatory mechanism for modulation of organ size in fruitfly. Subsequently, its involvement in the regulation of homeostasis and tumorigenesis has been identified. This pathway contains some tumor suppressor genes such as hippo (hpo) and warts (wts), as well as a number of oncogenic ones such as yorkie (yki). Recent studies have shown participation of Hippo pathway in the lung carcinogenesis. This pathway can affect lung cancer via different mechanisms. The interaction between some miRNAs and Hippo pathway is a possible mechanism for carcinogenic processes. Moreover, some other types of non-coding RNAs including PVT1, SFTA1P, NSCLCAT1 and circ_0067741 are implicated in this process. Besides, anti-cancer effects of gallic acid, icotinib hydrochloride, curcumin, ginsenoside Rg3, cryptotanshinone, nitidine chloride, cucurbitacin E, erlotinib, verteporfin, sophoridine, cisplatin and verteporfin in lung cancer are mediated through modulation of Hippo pathway. Here, we summarize the results of recent studies that investigated the role of Hippo signaling in the progression of lung cancer, the impact of non-coding RNAs on this pathway and the effects of anti-cancer agents on Hippo signaling in the context of lung cancer.
Topics: Humans; Hippo Signaling Pathway; Signal Transduction; Protein Serine-Threonine Kinases; Verteporfin; Drosophila Proteins; Lung Neoplasms
PubMed: 36736143
DOI: 10.1016/j.prp.2023.154339