<i>orsai</i>, the Drosophila homolog of human ETFRF1, links lipid catabolism to growth control

BACKGROUND: Lipid homeostasis is an evolutionarily conserved process that is crucial for energy production, storage and consumption. Drosophila larvae feed continuously to achieve the roughly 200-fold increase in size and accumulate sufficient reserves to provide all energy and nutrients necessary for the development of the adult fly. The mechanisms controlling this metabolic program are poorly understood. RESULTS: Herein we identified a highly conserved gene, orsai (osi), as a key player in lipid metabolism in Drosophila. Lack of osi function in the larval fat body, the regulatory hub of lipid homeostasis, reduces lipid reserves and energy output, evidenced by decreased ATP production and increased ROS levels. Metabolic defects due to reduced Orsai (Osi) in time trigger defective food-seeking behavior and lethality. Further, we demonstrate that downregulation of Lipase 3, a fat body-specific lipase involved in lipid catabolism in response to starvation, rescues the reduced lipid droplet size associated with defective orsai. Finally, we show that osi-related phenotypes are rescued through the expression of its human ortholog ETFRF1/LYRm5, known to modulate the entry of β-oxidation products into the electron transport chain; moreover, knocking down electron transport flavoproteins EtfQ0 and walrus/ETFA rescues osi-related phenotypes, further supporting this mode of action. CONCLUSIONS: These findings suggest that Osi may act in concert with the ETF complex to coordinate lipid homeostasis in the fat body in response to stage-specific demands, supporting cellular functions that in turn result in an adaptive behavioral response. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12915-022-01417-w

Dual Lipolytic Control of Body Fat Storage and Mobilization in Drosophila

Energy homeostasis is a fundamental property of animal life, providing a genetically fixed balance between fat storage and mobilization. The importance of body fat regulation is emphasized by dysfunctions resulting in obesity and lipodystrophy in humans. Packaging of storage fat in intracellular lipid droplets, and the various molecules and mechanisms guiding storage-fat mobilization, are conserved between mammals and insects. We generated a Drosophila mutant lacking the receptor (AKHR) of the adipokinetic hormone signaling pathway, an insect lipolytic pathway related to ß-adrenergic signaling in mammals. Combined genetic, physiological, and biochemical analyses provide in vivo evidence that AKHR is as important for chronic accumulation and acute mobilization of storage fat as is the Brummer lipase, the homolog of mammalian adipose triglyceride lipase (ATGL). Simultaneous loss of Brummer and AKHR causes extreme obesity and blocks acute storage-fat mobilization in flies. Our data demonstrate that storage-fat mobilization in the fly is coordinated by two lipocatabolic systems, which are essential to adjust normal body fat content and ensure lifelong fat-storage homeostasis

Diacylglycerol triggers Rim101 pathway dependent necrosis in yeast: a model for lipotoxicity

The loss of lipid homeostasis can lead to lipid overload and is associated with a variety of disease states. However, little is known as to how the disruption of lipid regulation or lipid overload affects cell survival. In this study we investigated how excess diacylglycerol (DG), a cardinal metabolite suspected to mediate lipotoxicity, compromises the survival of yeast cells. We reveal that increased DG achieved by either genetic manipulation or pharmacological administration of 1,2-dioctanoyl-sn-glycerol (DOG) triggers necrotic cell death. The toxic effects of DG are linked to glucose metabolism and require a functional Rim101 signaling cascade involving the Rim21 dependent sensing complex and activation of a calpain-like protease. The Rim101 cascade is an established pathway that triggers a transcriptional response to alkaline or lipid stress. We propose that the Rim101 pathway senses DG-induced lipid perturbation and conducts a signaling response that either facilitates cellular adaptation or triggers lipotoxic cell death. Using established models of lipotoxicity i.e. high fat diet in Drosophila and palmitic acid administration in cultured human endothelial cells, we present evidence that the core mechanism underlying this calpain-dependent lipotoxic cell death pathway is phylogenetically conserved

Reliable Drosophila Body Fat Quantification by a Coupled Colorimetric Assay

Factors and mechanisms controlling lipometabolism homeostasis share a remarkable evolutionary conservation between humans and Drosophila flies. Accordingly, the Drosophila model has been successfully used to understand the pathophysiology of human metabolic diseases such as obesity. Body fat stores in species as different as humans and flies consist of neutral lipids, mainly triacylglycerols. Changes in body fat storage are a diagnostic phenotype of lipometabolism imbalances of genetic or environmental origin. Various methods have been developed to quantify Drosophila body fat storage. The most widely used method adopts a commercial coupled colorimetric assay designed for human serum triacylglycerol quantification, which is based on glycerol content determination after enzymatic conversion of glycerides into glycerol. The coupled colorimetric assay is compatible with large-scale genetic screen approaches and has been successfully applied to characterize central regulators of Drosophila lipometabolism. Recently, the applicability of the coupled colorimetric assay for Drosophila storage fat quantification has been questioned in principle. Here we compare the performance of the coupled colorimetric assay on Drosophila samples with thin layer chromatography, the ‘‘gold standard’’ in storage lipid analysis. Our data show that the presented variant of the coupled colorimetric assay reliably discriminate

curled Encodes the Drosophila Homolog of the Vertebrate Circadian Deadenylase Nocturnin

Drosophila melanogaster curled, one of the first fly mutants described by T. H. Morgan >90 years ago, is the founding member of a series of curled wing phenotype mutants widely used as markers in fruit fly genetics. The expressivity of the wing phenotype is environmentally modulated, suggesting that the mutation affects the metabolic status of cells rather than a developmental control gene. However, the molecular identity of any of the curled wing marker mutant genes is still unknown. In a screen for starvation-responsive genes, we previously identified the single fly homolog of the vertebrate nocturnin genes, which encode cytoplasmic deadenylases that act in the post-transcriptional control of genes by poly(A) tail removal of target mRNAs prior to their degradation. Here we show that curled encodes Drosophila Nocturnin and that the gene is required at pupal stage for proper wing morphogenesis after eclosion of the fly. Despite the complex ontogenetic expression pattern of the gene, curled is not expressed in the developing wing, and wing-specific curled knockdown mediated by RNAi does not result in the curled wing phenotype, indicating a tissue-nonautonomous, systemic mode of curled gene function. Our study not only presents an entry point into the functional analysis of invertebrate nocturnins but also paves the way for the identification of the still elusive Nocturnin target mRNAs by genetic suppressor screens on the curled wing phenotype

Characterization of the Drosophila lipid droplet subproteome.

Lipid storage droplets are universal organelles essential for the cellular and organismal lipometabolism including energy homeostasis. Despite their apparently simple design they are proposed to participate in a growing number of cellular processes, raising the question to what extent the functional multifariousness is reflected by a complex organellar proteome composition. Here we present 248 proteins identified in a subproteome analysis using lipid storage droplets of Drosophila melanogaster fat body tissue. In addition to previously known lipid droplet-associated PAT (Perilipin, ADRP, and TIP47) domain proteins and homologues of several mammalian lipid droplet proteins, this study identified a number of proteins of diverse biological function, including intracellular trafficking supportive of the dynamic and multifaceted character of these organelles. We performed intracellular localization studies on selected newly identified subproteome members both in tissue culture cells and in fat body cells directly. The results suggest that the lipid droplets of fat body cells are of combinatorial protein composition. We propose that subsets of lipid droplets within single cells are characterized by a protein "zip code," which reflects functional differences or specific metabolic states

A drosophila in vivo screen identifies store-operated calcium entry as a key regulator of adiposity

To unravel the evolutionarily conserved genetic network underlying energy homeostasis, we performed a systematic in vivo gene knockdown screen in Drosophila. We used a transgenic RNAi library enriched for fly orthologs of human genes to functionally impair about half of all Drosophila genes specifically in adult fat storage tissue. This approach identified 77 genes, which affect the body fat content of the fly, including 58 previously unknown obesity-associated genes. These genes function in diverse biological processes such as lipid metabolism, vesicle-mediated trafficking, and the universal store-operated calciumentry (SOCE). Impairment of the SOCE core component Stromal interaction molecule (Stim), as well as other components of the pathway, causes adiposity in flies. Acute Stim dysfunction in the fat storage tissue triggers hyperphagia via remote control of the orexigenic short neuropeptide F in the brain, which in turn affects the coordinated lipogenic and lipolytic gene regulation, resulting in adipose tissue hypertrophy

Control of triglyceride storage by a WD40/TPR-domain protein

Obesity is a metabolic disorder related to improper control of energy uptake and expenditure, which results in excessive accumulation of body fat. Initial insights into the genetic pathways that regulate energy metabolism have been provided by a discrete number of obesity-related genes that have been identified in mammals. Here, we report the identification of the adipose (adp) gene, the mutation of which causes obesity in Drosophila. Loss of adp activity promotes increased fat storage, which extends the lifespan of mutant flies under starvation conditions. By contrast, adp gain-of-function causes a specific reduction of the fat body in Drosophila. adp encodes an evolutionarily conserved WD40/tetratricopeptide-repeat-domain protein that is likely to represent an intermediate in a novel signalling pathway