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The fat body in Drosophila larvae functions as a reserve tissue and participates in the regulation of organismal growth and homeostasis through its endocrine activity. To better understand its role in growth coordination, we induced fat body atrophy by knocking down several key enzymes of the glycolytic pathway in adipose cells. Our results show that impairing the last steps of glycolysis leads to a drastic drop in adipose cell size and lipid droplet content, and downregulation of the mTOR pathway and REPTOR transcriptional activity. Strikingly, fat body atrophy results in the distant disorganization of body wall muscles and the release of muscle-specific proteins in the hemolymph. Furthermore, we showed that REPTOR activity is required for fat body atrophy downstream of glycolysis inhibition, and that the effect of fat body atrophy on muscles depends on the production of TNF-α/egr and of the insulin pathway inhibitor ImpL2., (© 2024. The Author(s).)

Culex pipiens, the northern house mosquito, is a major vector of West Nile virus. To survive the severe winter, adult mosquitoes enter a diapause programme. Extended lifespan and an increase in lipid storage are key indicators of diapause. Post-translational modifications to histone proteins impact the expression of genes and have been linked to the lifespan and energy utilisation of numerous insects. Here, we investigated the potential contribution of epigenetic alterations in initiating diapause in this mosquito species. Multiple sequence alignment of H3 sequences from other insect species demonstrates a high conservation of the H3 histone in Cx. pipiens throughout evolution. We then compared the levels of histone methylation in the ovaries and fat body tissues of diapausing and non-diapausing Cx. pipiens using western blots. Our data indicate that histone methylation levels in the ovaries of Cx. pipiens do not change during diapause. In contrast, H3K27me2 levels decrease more than twofold in the fat body of diapausing mosquitoes relative to non-diapausing counterparts. H3K27 methylation plays a crucial role in chromosome activation and inactivation during development in many insect species. This is predominantly governed by polycomb repressor complex 2. Intriguingly, a previous ChIP-seq study demonstrated that the transcription factor FOXO (Forkhead box O) targets the genes that comprise this complex. In addition, H3K27me2 exhibits dynamic abundance throughout the diapause programme in Cx. pipiens, suggesting its potential role in the initial activation of the diapause programme. This study expands our understanding of the relationship between alterations in epigenetic regulation and diapause., (© 2023 The Royal Entomological Society.)

All organisms need to balance processes that consume energy against those that produce energy. With an increase in biological complexity over evolutionary time, regulation of this balance has become much more complex, resulting in specialization of metabolic tasks between organelles, cells, organs and, in the case of eusocial organisms, between the individuals that comprise the 'superorganism'. Exemplifying this, nurse honey bees maintain high abdominal lipids, while foragers have very low lipid stores, likely contributing to efficient performance of their social role, and thus to colony fitness. The proximate mechanisms responsible for these metabolic differences remain poorly understood. Here, we investigated the effects of age, worker class and dietary macronutrients on the abdominal activity of fatty acid synthase (FAS), the enzyme responsible for de novo synthesis of fatty acids, as well as the effects of age on lipase activity, enzymes responsible for the breakdown of stored lipids. We found that FAS but not lipase activity declines as bees age past peak nursing age. Feeding both nurses and foragers carbohydrates increased FAS activity compared with starved bees, but, whether fed or starved, nurses had much higher FAS activity than similarly treated foragers, implicating reduced lipid synthesis as one component of foragers' low lipid stores. Finally, we used artificial diets with different amounts of protein and fat to precociously induce low, forager-like FAS activity levels in nurse-age bees deprived of protein. We speculate that reduced protein appetite and consumption during the nurse-forager transition is responsible for suppressed lipid synthesis in foragers., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Spodoptera litura is a significant agricultural pest, and its glutathione S -transferase (GST) plays a crucial role in insecticide resistance. This study aimed to investigate the relationship between the SlGSTe11 gene of S. litura and resistance to cyantraniliprole and nicotine. Transcriptome analysis revealed that SlGSTe11 is highly expressed mainly in fat bodies, with a significant increase in SlGSTe11 gene expression under induction by cyantraniliprole and nicotine. The ectopic expression of the SlGSTe11 gene in transgenic fruit flies resulted in a 5.22-fold increase in the tolerance to cyantraniliprole. Moreover, compared to the UAS- SlGSTe11 line, the Act5C-UAS> SlGSTe11 line laid more eggs and had a lower mortality after nicotine exposure. RNAi-mediated inhibition of SlGSTe11 gene expression led to a significant increase in the mortality of S. litura under cyantraniliprole exposure. In vitro metabolism experiments demonstrated that the recombinant SlGSTe11 protein efficiently metabolizes cyantraniliprole. Molecular docking results indicated that SlGSTe11 has a strong affinity for both cyantraniliprole and nicotine. These findings suggest that SlGSTe11 is involved in the development of resistance to cyantraniliprole and nicotine in S. litura .

Azadirachtin is a widely used botanical pesticide for agricultural pest control worldwide. However, the molecular mechanisms of azadirachtin in insects are not fully understood. In this study, histological analysis and RNA sequencing were conducted to investigate the impact of azadirachtin on the larval development of Spodoptera frugiperda . Under azadirachtin exposure, the development was completely inhibited, and the major internal tissues, fat body, and midgut were strongly damaged under histological analysis. Differential gene expression analysis demonstrated that nutrient absorption and detoxification metabolism-related genes are differentially expressed. Interestingly, the expression of the apoptosis-related gene, caspase- 8, was significantly inhibited under exposure to azadirachtin. In addition, after knocking down the expression of the caspase- 8 gene, the fat body displayed a similar apoptotic phenotype as azadirachtin treatment; the distribution of chromatin and lipid droplets was uneven in the fat body cells. Thus, the results in this study demonstrated that exposure to azadirachtin rapidly activates apoptosis, resulting in innate tissue disruption, ultimately arresting larval development in S. frugiperda .

The interaction between environmental factors affecting honey bees is of growing concern due to their potential synergistic effects on bee health. Our study investigated the interactive impact of Varroa destructor and chlorothalonil on workers' survival, fat body morphology, and the expression of gene associated with detoxification, immunity, and nutrition metabolism during their adult stage. We found that both chlorothalonil and V. destructor significantly decreased workers' survival rates, with a synergistic effect observed when bees were exposed to both stressors simultaneously. Morphological analysis of fat body revealed significant alterations in trophocytes, particularly a reduction in vacuoles and granules after Day 12, coinciding with the transition of the bees from nursing to other in-hive work tasks. Gene expression analysis showed significant changes in detoxification, immunity, and nutrition metabolism over time. Detoxification genes, such as CYP9Q2, CYP9Q3, and GST-D1, were downregulated in response to stressor exposure, indicating a potential impairment in detoxification processes. Immune-related genes, including defensin-1, Dorsal-1, and Kayak, exhibited an initially upregulation followed by varied expression patterns, suggesting a complex immune response to stressors. Nutrition metabolism genes, such as hex 70a, AmIlp2, VGMC, AmFABP, and AmPTL, displayed dynamic expression changes, reflecting alterations in nutrient utilization and energy metabolism in response to stressors. Overall, these findings highlight the interactive and dynamic effects of environmental stressor on honey bees, providing insights into the mechanisms underlying honey bee decline. These results emphasize the need to consider the interactions between multiple stressors in honey bee research and to develop management strategies to mitigate their adverse effects on bee populations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The northern house mosquito, Culex pipiens, employs diapause as an essential survival strategy during winter, inducing important phenotypic changes such as enhanced stress tolerance, lipid accumulation, and extended longevity. During diapause, the cessation of reproductive development represents another distinctive phenotypic change, underlining the need for adjusted modulation of gene expressions within the ovary. Although considerable advancements in screening gene expression profiles in diapausing and non-diapausing mosquitoes, there remains a gap in tissue-specific transcriptomic profiling that could elucidate the complicated formation of diverse diapause features in Cx. pipiens. Here, we filled this gap by utilizing RNA sequencing, providing a detailed examination of gene expression patterns in the fat body and ovary during diapause compared to non-diapause conditions. Functional annotation of upregulated genes identified associations with carbohydrate metabolism, stress tolerance, immunity, and epigenetic regulation. The validation of candidate genes using quantitative real-time PCR verified the differentially expressed genes identified in diapausing mosquitoes. Our findings contribute novel insights into potential regulators during diapause in Cx. pipiens, thereby opening possible avenues for developing innovative vector control strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Ninein (Nin) is a microtubule (MT) anchor at the subdistal appendages of mother centrioles and the pericentriolar material (PCM) of centrosomes that also functions to organize MTs at noncentrosomal MT-organizing centers (ncMTOCs). In humans, the NIN gene is mutated in Seckel syndrome, an inherited developmental disorder. Here, we dissect the protein domains involved in Nin's localization and interactions with dynein and ensconsin (ens/MAP7) and show that the association with ens cooperatively regulates MT assembly in Drosophila fat body cells. We define domains of Nin responsible for its localization to the ncMTOC on the fat body cell nuclear surface, localization within the nucleus, and association with Dynein light intermediate chain (Dlic) and ens, respectively. We show that Nin's association with ens synergistically regulates MT assembly. Together, these findings reveal novel features of Nin function and its regulation of a ncMTOC.

Energy storage and endocrine functions of the Drosophila fat body make it an excellent model for elucidating mechanisms that underlie physiological and pathophysiological organismal metabolism. Combined with Drosophila's robust genetic and immunofluorescence microscopy toolkits, studies of Drosophila fat body function are ripe for cell biological analysis. Unlike the larval fat body, which is easily removed as a single, cohesive sheet of tissue, isolating intact adult fat body proves to be more challenging, thus hindering consistent immunofluorescence labeling even within a single piece of adipose tissue. Here, we describe an improved approach to handling Drosophila abdomens that ensures full access of the adult fat body to solutions generally used in immunofluorescence labeling protocols. In addition, we assess the quality of fluorescence reporter expression and antibody immunoreactivity in response to variations in fixative type, fixation incubation time, and detergent used for cellular permeabilization. Overall, we provide several recommendations for steps in a whole-mount staining protocol that results in consistent and robust immunofluorescence labeling of the adult Drosophila fat body., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

T2 RNases are transferase-type enzymes distributed across phyla, crucial for breaking down single-stranded RNA molecules. In addition to their canonical function, several T2 enzymes exhibit pleiotropic roles, contributing to various biological processes, such as the immune response in invertebrates and vertebrates. This study aims at characterizing RNASET2 in the larvae of black soldier fly (BSF), Hermetia illucens, which are used for organic waste reduction and the production of valuable insect biomolecules for feed formulation and other applications. Given the exposure of BSF larvae to pathogens present in the feeding substrate, it is likely that the mechanisms of their immune response have undergone significant evolution and increased complexity. After in silico characterization of HiRNASET2, demonstrating the high conservation of this T2 homolog, we investigated the expression pattern of the enzyme in the fat body and hemocytes, two districts mainly involved in the insect immune response, in larvae challenged with bacterial infection. While no variation in HiRNASET2 expression was observed in the fat body following infection, a significant upregulation of HiRNASET2 synthesis occurred in hemocytes shortly after the injection of bacteria in the larva. The intracellular localization of HiRNASET2 in lysosomes of plasmatocytes, its extracellular association with bacteria, and the presence of a putative antimicrobial domain in the molecule, suggest its potential role in RNA clean-up and as an alarm molecule promoting phagocytosis activation by hemocytes. These insights contribute to the characterization of the immune response of Hermetia illucens larvae and may facilitate the development of animal feedstuff enriched with highly valuable BSF bioactive compounds., (© 2024 The Author(s). Archives of Insect Biochemistry and Physiology published by Wiley Periodicals LLC.)

Energy metabolism is essential for insect development, reproduction and detoxification. Insects often reallocate energy and resources to manage external stress, balancing the demands of detoxification and reproduction. Glucose transport 4 (Glut4), a glucose transporter, is involved in glucose and lipid metabolism. However, the specific molecular mechanism of Glut4 in insect reproduction, and its role in the response to insecticide-induced oxidative stress remain unclear. In this study, LmGlut4 was identified and analyzed in Locusta migratoria. Silencing of LmGlut4 significantly reduced vitellogenin (Vg) biosynthesis in the fat body and Vg absorption by oocytes, ultimately hindering ovarian development and oocyte maturation. Knockdown of LmGlut4 also inhibited the biosynthesis of key insect hormones, such as juvenile hormone (JH), 20-hydroxyecdysone (20E) and insulin. Furthermore, LmGlut4 knockdown led to reduced triglyceride (TG) and glycogen content in the fat body and ovary, as well as decreased capacity for trehalose biosynthesis in adipocytes. Additionally, dsLmGlut4-treated locusts showed heightened sensitivity to deltamethrin, leading to increased triglyceride depletion during detoxification. This study sheds light on the biological function of LmGlut4 in the ovary and provides potential target genes for exploring biological pest management strategies., Competing Interests: Declaration of competing interest The authors declare that they have no competing financial interests or personal relationships., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Iron homeostasis is of critical importance to living organisms. Drosophila melanogaster has emerged as an excellent model to study iron homeostasis, while the regulatory mechanism of iron metabolism remains poorly understood. Herein, we accidently found that knockdown of juvenile hormone (JH) acid methyltransferase (Jhamt) specifically in the fat body, a key rate-limiting enzyme for JH synthesis, led to iron accumulation locally, resulting in serious loss and dysfunction of fat body. Jhamt knockdown-induced phenotypes were mitigated by iron deprivation, antioxidant and Ferrostatin-1, a well-known inhibitor of ferroptosis, suggesting ferroptosis was involved in Jhamt knockdown-induced defects in the fat body. Further study demonstrated that upregulation of Tsf1 and Malvolio (Mvl, homolog of mammalian DMT1), two iron importers, accounted for Jhamt knockdown-induced iron accumulation and dysfunction of the fat body. Mechanistically, Kr-h1, a key transcription factor of JH, acts downstream of Jhamt inhibiting Tsf1 and Mvl transcriptionally. In summary, the findings indicated that fat body-derived Jhamt is required for the development of Drosophila by maintaining iron homeostasis in the fat body, providing unique insight into the regulatory mechanisms of iron metabolism in Drosophila., (© 2024 Federation of American Societies for Experimental Biology.)

Today, plastic pollution is one of the biggest threats to the environment and public health. In the tissues of exposed species, micro- and nano-fragments accumulate, leading to genotoxicity, altered metabolism, and decreased lifespan. A model to investigate the genotoxic and tumor-promoting potential of nanoplastics (NPs) is Drosophila melanogaster . Here we tested polystyrene, which is commonly used in food packaging, is not well recycled, and makes up at least 30% of landfills. In order to investigate the biological effects and carcinogenic potential of 100 µm polystyrene nanoparticles (PSNPs), we raised Oregon [R] wild-type flies on contaminated food. After prolonged exposure, fluorescent PSNPs accumulated in the gut and fat bodies. Furthermore, PSNP-fed flies showed considerable alterations in weight, developmental time, and lifespan, as well as a compromised ability to recover from starvation. Additionally, we noticed a decrease in motor activity in DNA lig4 mutants fed with PSNPs, which are known to be susceptible to dietary stressors. A qPCR molecular investigation of the larval intestines revealed a markedly elevated expression of the genes drice and p53 , suggesting a response to cell damage. Lastly, we used warts -defective mutants to assess the carcinogenic potential of PSNPs and discovered that exposed flies had more aberrant masses than untreated ones. In summary, our findings support the notion that ingested nanopolystyrene triggers metabolic and genetic modifications in the exposed organisms, eventually delaying development and accelerating death and disease.

Human activities associated with large-scale farms and the monocultures expose honey bees to one type of food. Moreover, there is an ongoing decline of plant species producing pollen and nectar in Europe. A poorly balanced diet affects a number of processes occurring in a bee's body. The fat body and hemolymph are the tissues that participate in all of them. Therefore, the aim of our study was to determine the effect of hazel, pine, rapeseed, buckwheat, phacelia and goldenrod pollen on the morphological parameters of fat body trophocytes, the diameters of cell nuclei in oenocytes and the concentrations of compounds involved in energy metabolism (glucose, glycogen, triglycerides and protein). In the cage tests, the bees were fed from the first day of life with sugar candy (control group) or candy with a 10% addition of one of the 6 pollen types. Hemolymph and fat body from various locations were collected from 1-, 7- and 14-day-old workers. Pollen produced by plant species such as hazel and pine increased glucose concentrations in the bee tissues, especially in the hemolymph. It can therefore be concluded that they are valuable sources of energy (in the form of simple carbohydrates) which are quickly used by bees. Pollen from plants blooming in the summer and autumn increased the concentrations of proteins, glycogen and triglycerides in the fat body, especially that from the third tergite. The accumulation of these compounds was associated with an increased the length and width of trophocytes as well as with enhanced metabolic activity, which was evidenced in the increasing diameter of oenocyte cell nuclei. It seems a balanced multi-pollen diet is more valuable for bees, but it is important to understand the effects of the particular pollen types in the context of a mono-diet. In the future, this will make it possible to produce mixtures that can ensure homeostasis in the apian body., (© 2024. The Author(s).)

Melittin is a powerful toxin present in honeybee venom that is active in a wide range of animals, from insects to humans. Melittin exerts numerous biological, toxicological, and pharmacological effects, the most important of which is destruction of the cell membrane. The phospholipase activity of melittin and its ability to activate phospholipases in the venom contribute to these actions. Using analytical methods, we discovered that the honeybee Apis mellifera produces melittin not only in the venom gland but also in its fat body cells, which remain resistant to this toxin's effects. We suggest that melittin acts as an anti-bacterial agent, since its gene expression is significantly upregulated when honeybees are infected with Escherichia coli and Listeria monocytogenes bacteria; additionally, melittin effectively kills these bacteria in the disc diffusion test. We hypothesize that the chemical and physicochemical properties of the melittin molecule (hydrophilicity, lipophilicity, and capacity to form tetramers) in combination with reactive conditions (melittin concentration, salt concentration, pH, and temperature) are responsible for the targeted destruction of bacterial cells and apparent tolerance towards own tissue cells. Considering that melittin is an important current and, importantly, potential broad-spectrum medication, a thorough understanding of the observed phenomena may significantly increase its use in clinical practice., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Blister beetles of Epicauta impressicornis have attracted attention because they contain a large amount of cantharidin (CTD). To date, however, the synthesis and transfer of CTD in adults of E. impressicornis are largely unknown. Here, we showed that the larvae E. impressicornis are capable of synthesizing CTD and they consume CTD during pupation. Before sexual maturity, both male and female adults synthesized a small amount of CTD, while after sexual maturity, males produced larger amounts of CTD, but females did not. The newly synthesized CTD in males first appeared in the hemolymph and then accumulated in the reproductive system. During the mating, the males transferred CTD to the reproductive system of females. In addition, a farnesyl pyrophosphate synthase (FPPS) gene was identified in male E. impressicornis . RNA-seq analysis, quantitative RT-PCR, and RNA interference analyses were conducted to investigate expression patterns and the functional roles of E. impressicornis FPPS ( EiFPPS ). Our results indicate that EiFPPS is highly expressed in the fat body of males. Moreover, the knock-down of EiFPPS led to a significant decrease in CTD synthesis. The current study indicates that EiFPPS is expressed in the fat body to regulate CTD synthesis in male E. impressicornis blister beetles.

Lipid droplets (LDs) are dynamic organelles that participate in the regulation of lipid metabolism and cellular homeostasis inside of cells. LD-associated proteins, also known as perilipins (PLINs), are a family of proteins found on the surface of LDs that regulate lipid metabolism, immunity, and other functions. In silkworms, pébrine disease caused by infection by the microsporidian Nosema bombycis ( Nb ) is a severe threat to the sericultural industry. Although we found that Nb relies on lipids from silkworms to facilitate its proliferation, the relationship between PLINs and Nb proliferation remains unknown. Here, we found Nb infection caused the accumulation of LDs in the fat bodies of silkworm larvae. The characterized perilipin1 gene ( plin1 ) promotes the accumulation of intracellular LDs and is involved in Nb proliferation. plin1 is similar to perilipin1 in humans and is conserved in all insects. The expression of plin1 was mostly enriched in the fat body rather than in other tissues. Knockdown of plin1 enhanced Nb proliferation, whereas overexpression of plin1 inhibited its proliferation. Furthermore, we confirmed that plin1 increased the expression of the Domeless and Hop in the JAK-STAT immune pathway and inhibited Nb proliferation. Taken together, our current findings demonstrate that plin1 inhibits Nb proliferation by promoting the JAK-STAT pathway through increased expression of Domeless and Hop . This study provides new insights into the complicated connections among microsporidia pathogens, LD surface proteins, and insect immunity.IMPORTANCELipid droplets (LDs) are lipid storage sites in cells and are present in almost all animals. Many studies have found that LDs may play a role in host resistance to pathogens and are closely related to innate immunity. The present study found that a surface protein of insect lipid droplets could not only regulate the morphological changes of lipid droplets but also inhibit the proliferation of a microsporidian pathogen Nosema bombycis ( Nb ) by activating the JAK-STAT signaling pathway. This is the first discovery of the relationship between microsporidian pathogen and insect lipid surface protein perilipin and insect immunity., Competing Interests: The authors declare no conflict of interest.

Until recently, chemical pesticides were one of the most effective means of controlling agricultural pests; therefore, the search for insecticide targets for agricultural pests has been an ongoing problem. Estrogen-related receptors (ERRs) are transcription factors that regulate cellular metabolism and energy homeostasis in animals. Silkworms are highly sensitive to chemical pesticides, making them ideal models for pesticide screening and evaluation. In this study, we detected ERR expression in key organs involved in pesticide metabolism in silkworms (Bombyx mori), including the fat body and midgut. Using ChIP-seq technology, many estrogen- related response elements were identified in the 2000-bp promoter region upstream of metabolism-related genes, almost all of which were potential ERR target genes. The ERR inhibitor, XCT-790, and the endocrine disruptor, bisphenol A, significantly inhibited expression of the ERR target genes, BmTreh-1, BmTret-1, BmPK, BmPFK, and BmHK, in the fat bodies of silkworms, resulting in pupation difficulties in silkworm larvae that ultimately lead to death. In addition, based on the clarification that the ERR can bind to XCT-790, as observed through biofilm interferometry, its three-dimensional spatial structure was predicted, and using molecular docking techniques, small-molecule compounds with a stronger affinity for the ERR were identified. In summary, utilizing the powerful metabolic regulatory function of ERR in Lepidoptera pests, the developed small molecule inhibitors of ERR can be used for future control of Lepidoptera pests., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Pancreatic β cells secrete insulin in response to glucose elevation to maintain glucose homeostasis. A complex network of inter-organ communication operates to modulate insulin secretion and regulate glucose levels after a meal. Lipids obtained from diet or generated intracellularly are known to amplify glucose-stimulated insulin secretion, however, the underlying mechanisms are not completely understood. Here, we show that a Drosophila secretory lipase, Vaha (CG8093), is synthesized in the midgut and moves to the brain where it concentrates in the insulin-producing cells in a process requiring Lipid Transfer Particle, a lipoprotein originating in the fat body. In response to dietary fat, Vaha stimulates insulin-like peptide release (ILP), and Vaha deficiency results in reduced circulatory ILP and diabetic features including hyperglycemia and hyperlipidemia. Our findings suggest Vaha functions as a diacylglycerol lipase physiologically, by being a molecular link between dietary fat and lipid amplified insulin secretion in a gut-brain axis., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Lipid distribution in an organism is mediated by the interplay between lipoprotein particles, lipoprotein receptors and class B scavenger receptors of the CD36 family. CD36 is a multifunctional protein mediating lipid uptake, mobilization and signaling at the plasma membrane and inside of the cell. The CD36 protein family has 14 members in Drosophila melanogaster, which allows for the differentiated analysis of their functions. Here, we unravel a role for the so far uncharacterized scavenger receptor Bez in lipid export from Drosophila adipocytes. Bez shares the lipid binding residue with CD36 and is expressed at the plasma membrane of the embryonic, larval and adult fat body. Bez loss of function lowers the organismal availability of storage lipids and blocks the maturation of egg chambers in ovaries. We demonstrate that Bez interacts with the APOB homolog Lipophorin at the plasma membrane of adipocytes and trace the Bez-dependent transfer of an alkyne-labeled fatty acid from adipocytes to Lipophorin. Our study demonstrates how lipids are distributed by scavenger receptor-lipoprotein interplay and contribute to the metabolic control of development., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Bombyx mori triose-phosphate transporter protein (BmTPT) is a member of the solute carrier (SLC) family. Its main function is to transport triose phosphate between intracellular and extracellular. In this study, BmTPT was cloned and characterised from the fat body of the silkworm Bombyx mori, resulting in an open reading frame (ORF) with a full length of 936 bp, which can encode 311 amino acid residues and has eight transmembrane structural domains. BmTPT was distributed throughout the cell and deposited the most in the nucleus, and is expressed in all tissues of Bombyx mori. Bombyx mori nucleopolyhedrovirus (BmNPV) infection significantly up-regulated BmTPT expression in immune tissue fat bodies. In addition, overexpression of BmTPT significantly inhibited BmNPV infection and markedly reduced the expression of enzymes related to the cellular glycolytic pathway; on the contrary, down-regulation of BmTPT expression by RNA interference resulted in robust replication of BmNPV and a significant increase in the expression of enzymes related to the cellular glycolytic pathway. This is the first report that BmTPT has antiviral effect in silkworm, and also could result in a lack of energy and raw materials for BmNPV replication and infection through down-regulation of the cellular glycolytic pathway., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

We explored the metabolic integration of Blattella germanica and its obligate endosymbiont Blattabacterium cuenoti by the transcriptomic analysis of the fat body of quasi-aposymbiotic cockroaches, where the endosymbionts were almost entirely removed with rifampicin. Fat bodies from quasi-aposymbiotic insects displayed large differences in gene expression compared to controls. In quasi-aposymbionts, the metabolism of phenylalanine and tyrosine involved in cuticle sclerotization and pigmentation increased drastically to compensate for the deficiency in the biosynthesis of these amino acids by the endosymbionts. On the other hand, the uricolytic pathway and the biosynthesis of uric acid were severely decreased, probably because the reduced population of endosymbionts was unable to metabolize urea to ammonia. Metabolite transporters that could be involved in the endosymbiosis process were identified. Immune system and antimicrobial peptide (AMP) gene expression was also reduced in quasi-aposymbionts, genes encoding peptidoglycan-recognition proteins, which may provide clues for the maintenance of the symbiotic relationship, as well as three AMP genes whose involvement in the symbiotic relationship will require additional analysis. Finally, a search for AMP-like factors that could be involved in controlling the endosymbiont identified two orphan genes encoding proteins smaller than 200 amino acids underexpressed in quasi-aposymbionts, suggesting a role in the host-endosymbiont relationship.

The European corn borer (Ostrinia nubilalis, Hbn.), enters diapause, a strategy characterized by arrest of development and reproduction, reduction of metabolic rate and the emergence of increased resistance to challenging seasonal conditions as low sub-zero winter temperatures. The aim of this study was to investigate the potential role of inorganic elements in the ecophysiology of O. nubilalis, analysing their content in the whole body, hemolymph and fat body, both metabolically active, non-diapausing and overwintering diapausing larvae by ICP-OES spectrometer following the US EPA method 200.7:2001. O nubilalis as many phytophagous lepidopteran species maintain a very low extracellular sodium concentration and has potassium as dominant cation in hemolymph of their larvae. Changes in hemolymph and the whole body sodium content occur already at the onset of diapause (when the mean environmental temperatures are still high above 0 ºC) and remain stable during the time course of diapause when larvae of this species cope with sub-zero temperatures, it seems that sodium content regulation is rather a part of diapausing program than the direct effect of exposure to low temperatures. Compared to non-diapausing O. nubilalis larvae, potassium levels are much higher in the whole body and fat body of diapausing larvae and substantially increase approaching the end of diapause. The concentration of Ca, Mg, P and S differed in the whole body, hemolymph and fat body between non-diapausing and diapausing larvae without a unique trend during diapause, except an increase in their contents at the end of diapause., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Ticks are blood-feeding arachnids that are known to transmit various pathogenic microorganisms to their hosts. During blood feeding, ticks activate their metabolism and immune system to efficiently utilise nutrients from the host's blood and complete the feeding process. In contrast to insects, in which the fat body is known to be a central organ that controls essential metabolic processes and immune defense mechanisms, the function of the fat body in tick physiology is still relatively unexplored. To fill this gap, we sought to uncover the repertoire of genes expressed in the fat body associated with trachea (FB/Tr) by analyzing the transcriptome of individual, partially fed (previtellogenic) Ixodes ricinus females. The resulting catalog of individual mRNA sequences reveals a broad repertoire of transcripts encoding proteins involved in nutrient storage and distribution, as well as components of the tick immune system. To gain a detailed insight into the secretory products of FB/Tr specifically involved in inter-tissue transport and humoral immunity, the transcriptomic data were complemented with the proteome of soluble proteins in the hemolymph of partially fed female ticks. Among these proteins, the hemolipoglyco-carrier proteins were predominant. When comparing immune peptides and proteins from the fat body with those produced by hemocytes, we found that the fat body serves as a unique producer of certain immune components. Finally, time-resolved transcriptional regulation of selected immune transcripts from the FB/Tr was examined in response to experimental challenges with model microbes and analyzed by RT-qPCR. Overall, our data show that the fat body of ticks, similar to insects, is an important metabolic tissue that also plays a remarkable role in immune defense against invading microbes. These findings improve our understanding of tick biology and its impact on the transmission of tick-borne pathogens., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

In this study, we found that in the adipose tissue of wildtype animals, insulin and TGF-β signalling converge via a BMP antagonist short gastrulation (sog) to regulate ECM remodelling. In tumour bearing animals, Sog also modulates TGF-β signalling to regulate ECM accumulation in the fat body. TGF-β signalling causes ECM retention in the fat body and subsequently depletes muscles of fat body-derived ECM proteins. Activation of insulin signalling, inhibition of TGF-β signalling, or modulation of ECM levels via SPARC, Rab10 or Collagen IV in the fat body, is able to rescue tissue wasting in the presence of tumour. Together, our study highlights the importance of adipose ECM remodelling in the context of cancer cachexia., (© 2023 The Authors. Published under the terms of the CC BY 4.0 license.)

Cytochrome P450 plays vital roles in detoxifying xenobiotics. In this study, SlCYP340A and SlCYP340L expression in the Spodoptera litura fat body and SlCYP332A1 , SlCYP6AB12 , SlCYP6AB58 , SlCYP6AB59, and SlCYP6AN4 expression in the Malpighian tubules were significantly upregulated after cyantraniliprole exposure, and SlCYP6AB58 and SlCYP6AB59 expression levels were simultaneously increased in the Malpighian tubules after gossypol treatment. Drosophila ectopically expressing candidate P450 genes showed that SlCYP332A1 , SlCYP6AB12 , SlCYP6AB59 , SlCYP6AN4, and SlCYP340A conferred cyantraniliprole tolerance. The overexpression of SlCYP6AB58 and SlCYP6AB59 in Drosophila increased the number of eggs laid under the gossypol treatment. Moreover, the knockdown of SlCYP332A1 , SlCYP6AB12 , SlCYP6AB59 , SlCYP6AN4, and SlCYP340A increased S. litura mortality under the cyantraniliprole treatment. Homology modeling and molecular docking results suggested that candidate P450 has the potential to bind with cyantraniliprole. These results indicate that the CYP3 and CYP4 genes participate in cyantraniliprole detoxification and that SlCYP6AB59 may be simultaneously involved in the gossypol tolerance of S. litura .

The control of insect moulting and metamorphosis involves ecdysteroids that orchestrate the execution of developmental genetic programs by binding to dimeric hormone receptors consisting of the ecdysone receptor (EcR) and ultraspiracle (USP). In insects, the main ecdysteroids comprise ecdysone (E), which is synthesized in the prothoracic gland and secreted into the haemolymph, and 20-hydroxyecdysone (20E), which is considered the active form by binding to the nuclear receptor of the target cell. While biosynthesis of ecdysteroids has been studied in detail in different insects, the transport systems involved in guiding these steroid hormones across cellular membranes have just recently begun to be studied. By analysing RNAi phenotypes in the red flour beetle, Tribolium castaneum, we have identified three transporter genes, TcABCG-8A, TcABCG-4D and TcOATP4-C1, whose silencing results in phenotypes similar to that observed when the ecdysone receptor gene TcEcRA is silenced, that is, abortive moulting and abnormal development of adult compound eyes during the larval stage. The genes of all three transporters are expressed at higher levels in the larval fat body of T. castaneum. We analysed potential functions of these transporters by combining RNAi and mass spectrometry. However, the analysis of gene functions is challenged by mutual RNAi effects indicating interdependent gene regulation. Based on our findings, we propose that TcABCG-8A, TcABCG-4D and TcOATP4-C1 participate in the ecdysteroid transport in fat body cells, which are involved in E → 20E conversion catalysed by the P450 enzyme TcShade., (© 2023 The Authors. Insect Molecular Biology published by John Wiley & Sons Ltd on behalf of Royal Entomological Society.)

The fat body plays a central role in the regulation of the life cycle of insects and acts as the major site for detoxification, nutrient storage, energy metabolism, and innate immunity. However, the diversity of cell types in the fat body, as well as how these cell subsets respond to virus infection, remains largely unknown. We used single-nucleus RNA sequencing to identify 23 distinct clusters representing adipocyte, hemocyte, epithelial cell, muscle cell, and glial cell types in the fat body of silkworm larvae. Further, by analysis of viral transcriptomes in each cell subset, we reveal that all fat body cells could be infected by Bombyx mori nucleopolyhedrovirus (BmNPV) at 72 h postinfection, and that the majority of infected cells carried at least a medium viral load, whereas most cells infected by BmNPV at 24 h postinfection had only low levels of infection. Finally, we characterize the responses occurring in the fat body cell clusters on BmNPV infection, which, on one hand, mainly reduce their metabolic functions, involving energy, carbohydrates, lipids, and amino acids, but, on the other hand, initiate a strong antiviral response. Our single-nucleus RNA sequencing analysis reveals the diversity of insect fat body cells and provides a resource of gene expression profiles for a systems-level understanding of their response to virus infection., (Copyright © 2023 by The American Association of Immunologists, Inc.)

The steroid hormone 20-hydroxyecdysone (20E) has been described to regulate fat body lipid metabolism in insects, but its accurate regulatory mechanism, especially the crosstalk between 20E-induced lipid metabolism and gluconeogenesis remains largely unclear. Here, we specially investigated the effect of 20E on lipid metabolism and gluconeogenesis in the fat body of Hyphantria cunea larvae, a notorious pest in forestry. Lipidomics analysis showed that a total of 1 907 lipid species were identified in the fat body of H. cunea larvae assigned to 6 groups and 48 lipid classes. The differentially abundant lipids analysis showed a significant difference between 20E-treated and control samples, indicating that 20E caused a remarkable alteration of lipidomics profiles in the fat body of H. cunea larvae. Further studies demonstrated that 20E accelerated fatty acid β-oxidation, inhibited lipid synthesis, and promoted lipolysis. Meanwhile, the activities of pyruvate carboxylase, phosphoenolpyruvate carboxykinase, fructose-1,6-bisphosphatase, and glucose-6-phosphatase were dramatically suppressed by 20E in the fat body of H. cunea larvae. As well, the transcriptions of genes encoding these 4 rate-limiting gluconeogenic enzymes were significantly downregulated in the fat body of H. cunea larvae after treatment with 20E. Taken together, our results revealed that 20E disturbed fat body lipid homeostasis, accelerated fatty acid β-oxidation and promoted lipolysis, but negatively regulated gluconeogenesis in H. cunea larvae. The findings might provide a new insight into hormonal regulation of glucose and lipid metabolism in insect fat body., (© 2022 Institute of Zoology, Chinese Academy of Sciences.)

Hexamerins, the proteins massively stored in the larval haemolymph of insects, are gradually used throughout metamorphosis as a source of raw material and energy for the development of adult tissues. Such behaviour defined hexamerins as storage proteins. Immunofluorescence experiments coupled with confocal microscopy show a hexamerin, HEX 70a, in the nucleus of the brain and fat body cells from honeybee workers, an unexpected localization for a storage protein. HEX 70a colocalizes with fibrillarin, a nucleolar-specific protein and H3 histone, thus suggesting a potential role as a chromatin-binding protein. This was investigated through chromatin immunoprecipitation and high-throughput DNA sequencing (ChIP-seq). The significant HEX 70a-DNA binding sites were mainly localized at the intergenic, promoter and intronic regions. HEX 70a targeted DNA stretches mapped to the genomic regions encompassing genes with relevant functional attributes. Several HEX 70a targeted genes were associated with H3K27ac or/and H3K27me3, known as active and repressive histone marks. Brain and fat body tissues shared a fraction of the HEX 70 targeted genes, and tissue-specific targets were also detected. The presence of overrepresented DNA motifs in the binding sites is consistent with specific HEX 70a-chromatin association. In addition, a search for HEX 70a targets in RNA-seq public libraries of fat bodies from nurses and foragers revealed differentially expressed targets displaying hex 70a-correlated developmental expression, thus supporting a regulatory activity for HEX 70a. Our results support the premise that HEX 70a is a moonlighting protein that binds chromatin and has roles in the brain and fat body cell nuclei, apart from its canonical role as a storage protein., (© 2023 Royal Entomological Society.)

Nuclear receptor-binding SET domain-containing protein 1 (NSD1) inactivation in tumor cells contributes to an immune-cold phenotype, indicating its potential association with immune disturbances. Drosophila NSD is a homolog of the human NSD1. Thus, in this study, we investigated the effect of NSD overexpression in the fat body, the central organ involved in Drosophila immune responses. Upon ectopic expression of NSD in the fat body, the mRNA levels of antimicrobial peptides increased. Using reporter constructs containing deletions of various NF-κB sites in the Attacin-A ( AttA ) promoter, we found that transcriptional activation by NSD is mainly mediated via the IMD pathway by activating Relish. Since the IMD pathway is required to resist Gram-negative bacterial infections, we further examined the effect of fat body-specific NSD overexpression on Drosophila immune defenses. Upon oral ingestion of Gram-negative Pseudomonas entomophila , the survival rate of the NSD-overexpressing larvae was higher than that of the wild type, suggesting a positive role of NSD in immune responses. Taken together, these results suggest the association of NSD with the IMD pathway and is thus expected to contribute to the elucidation of the molecular mechanisms of immune malfunction in various NSD1-associated human diseases.

Defective nutrient storage and adipocyte enlargement (hypertrophy) are emerging features of metabolic syndrome and type 2 diabetes. Within adipose tissues, how the cytoskeletal network contributes to adipose cell size, nutrient uptake, fat storage, and signaling remain poorly understood. Utilizing the Drosophila larval fat body (FB) as a model adipose tissue, we show that a specific actin isoform-Act5C-forms the cortical actin network necessary to expand adipocyte cell size for biomass storage in development. Additionally, we uncover a non-canonical role for the cortical actin cytoskeleton in inter-organ lipid trafficking. We find Act5C localizes to the FB cell surface and cell-cell boundaries, where it intimately contacts peripheral LDs (pLDs), forming a cortical actin network for cell architectural support. FB-specific loss of Act5C perturbs FB triglyceride (TG) storage and LD morphology, resulting in developmentally delayed larvae that fail to develop into flies. Utilizing temporal RNAi-depletion approaches, we reveal that Act5C is indispensable post-embryogenesis during larval feeding as FB cells expand and store fat. Act5C-deficient FBs fail to grow, leading to lipodystrophic larvae unable to accrue sufficient biomass for complete metamorphosis. In line with this, Act5C-deficient larvae display blunted insulin signaling and reduced feeding. Mechanistically, we also show this diminished signaling correlates with decreased lipophorin (Lpp) lipoprotein-mediated lipid trafficking, and find Act5C is required for Lpp secretion from the FB for lipid transport. Collectively, we propose that the Act5C-dependent cortical actin network of Drosophila adipose tissue is required for adipose tissue size-expansion and organismal energy homeostasis in development, and plays an essential role in inter-organ nutrient transport and signaling., Competing Interests: RU, ST, JB, AK, BP, WH No competing interests declared, (© 2023, Ugrankar-Banerjee et al.)

Background: 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., Results: Here, we 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., Conclusions: Our results indicated that FOI is critical in coordinating fat body endoreplication and larval growth in Drosophila. Our study provides a novel insight into the relationship between zinc and endoreplication in insects and may provide a reference for relevant mammalian studies., (© 2023. The Author(s).)

The anatomical, physiological, and behavioral characteristics of honey bees are affected by the season as well as division of labor. In this study, we examined the structure, ultrastructure, and gene expression of fat body cells in both long-lived winter and short-lived summer worker bees (the youngest stage of hive bees and forager bees). In contrast to hive bees, foragers and winter bees have a higher metabolism due to intensive muscle activity during their flight (foragers) or endothermic heat production (winter bees). These workers differ from hive bees in the biology of their mitochondria, peroxisomes, and lysosomes as well as in the expression of the genes involved in lipid, carbohydrate, amino acid metabolism, insulin, and TGF- β signaling. Additionally, the expression of genes related to phospholipid metabolism was higher in the hive bees. However, we found no differences between workers in the expression of genes controlling cell organelles, such as the Golgi apparatus, endoplasmic reticulum, ribosomes, nucleus, and vacuoles, as well as genes for DNA replication, cell cycle control, and autophagy. Furthermore, lysosomes, autophagic processes and lipofuscin particles were more frequently observed in winter bees using electron microscopy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Knowledge of adipogenetic mechanisms is essential to understand and treat conditions affecting organismal metabolism and adipose tissue health. In Drosophila, mature adipose tissue (fat body) exists in larvae and adults. In contrast to the well-known development of the larval fat body from the embryonic mesoderm, adult adipogenesis has remained mysterious. Furthermore, conclusive proof of its physiological significance is lacking. Here, we show that the adult fat body originates from a pool of undifferentiated mesodermal precursors that migrate from the thorax into the abdomen during metamorphosis. Through in vivo imaging, we found that these precursors spread from the ventral midline and cover the inner surface of the abdomen in a process strikingly reminiscent of embryonic mesoderm migration, requiring fibroblast growth factor (FGF) signaling as well. FGF signaling guides migration dorsally and regulates adhesion to the substrate. After spreading is complete, precursor differentiation involves fat accumulation and cell fusion that produces mature binucleate and tetranucleate adipocytes. Finally, we show that flies where adult adipogenesis is impaired by knock down of FGF receptor Heartless or transcription factor Serpent display ectopic fat accumulation in oenocytes and decreased resistance to starvation. Our results reveal that adult adipogenesis occurs de novo during metamorphosis and demonstrate its crucial physiological role., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Lei et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The survival of animals during periods of limited nutrients is dependent on the organism's ability to store lipids during times of nutrient abundance. However, the increased availability of food in modern western society has led to an excess storage of lipids resulting in metabolic diseases. To better understand the genes involved in regulating lipid storage, genome-wide RNAi screens were performed in cultured Drosophila cells and one group of genes identified includes mRNA splicing factor genes. Our lab has previously shown that a group of splicing factors important for intron/exon border recognition known as SR proteins are involved in controlling lipid storage in Drosophila; however, how these SR proteins are regulated to control lipid storage is not fully understood. Here, we focus on two SR protein kinases (SRPKs) in Drosophila: SRPK and SRPK79D. Decreasing the expression of these genes specifically in the adult fat body using RNAi resulted in lower levels of triglycerides and this is due to a decrease in the amount of fat stored per cell, despite having more fat cells, when compared to control flies. Decreasing SRPK and SRPK79D levels in the fat body leads to altered splicing of the β-oxidation gene, carnitine palmitoyltransferase 1 (CPT1), resulting in increased production of a more active enzyme, which would increase lipid breakdown and be consistent with the lean phenotype observed in these flies. In addition, flies with decreased SRPK and SRPK79D levels in their fat bodies eat less, which may also contribute to the decreased triglyceride phenotype. Together, these findings provide evidence to support that lipid storage is controlled by the phosphorylation of factors involved in mRNA splicing., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

In Lepidoptera and Diptera, the fat body dissociates into single cells in nondiapause pupae, but it does not dissociate in diapause pupae until diapause termination. Using the cotton bollworm, Helicoverpa armigera , as a model of pupal diapause insects, we illustrated the catalytic mechanism and physiological importance of fat body cell dissociation in regulating pupal development and diapause. In nondiapause pupae, cathepsin L (CatL) activates matrix metalloproteinases (Mmps) that degrade extracellular matrix proteins and cause fat body cell dissociation. Mmp-induced fat body cell dissociation activates lipid metabolism through transcriptional regulation, and the resulting energetic supplies increase brain metabolic activity (i.e., mitochondria respiration and insulin signaling) and thus promote pupal development. In diapause pupae, low activities of CatL and Mmps prevent fat body cell dissociation and lipid metabolism from occurring, maintaining pupal diapause. Importantly, as demonstrated by chemical inhibitor treatments and CRISPR-mediated gene knockouts, Mmp inhibition delayed pupal development and moderately increased the incidence of pupal diapause, while Mmp stimulation promoted pupal development and moderately averted pupal diapause. This study advances our recent understanding of fat body biology and insect diapause regulation.

Defects in how excess nutrients are stored as triglycerides can result in several diseases including obesity, heart disease, and diabetes. Understanding the genes responsible for normal lipid homeostasis will help understand the pathogenesis of these diseases. RNAi screens performed in Drosophila cells identified genes involved in vesicle formation and protein sorting as important for the formation of lipid droplets; however, all of the vesicular trafficking proteins that regulate lipid storage are unknown. Here, we characterize the function of the Drosophila Charged multivesicular protein 1 ( Chmp1 ) gene in regulating fat storage. Chmp1 is a member of the ESCRT-III complex that targets membrane localized signaling receptors to intralumenal vesicles in the multivesicular body of the endosome and then ultimately to the lysosome for degradation. When Chmp1 levels are decreased specifically in the fly fat body, triglyceride accumulates while fat-body-specific Chmp1 overexpression decreases triglycerides. Chmp1 controls triglyceride storage by regulating the number and size of fat body cells produced and not by altering food consumption or lipid metabolic enzyme gene expression. Together, these data uncover a novel function for Chmp1 in controlling lipid storage in Drosophila and supports the role of the endomembrane system in regulating metabolic homeostasis.

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., (© 2022. The Author(s).)

Diet-induced obesity leads to dysfunctional feeding behavior. However, the precise molecular nodes underlying diet-induced feeding motivation dysregulation are poorly understood. The fruit fly is a simple genetic model system yet displays significant evolutionary conservation to mammalian nutrient sensing and energy balance. Using a longitudinal high-sugar regime in Drosophila , we sought to address how diet-induced changes in adipocyte lipid composition regulate feeding behavior. We observed that subjecting adult Drosophila to a prolonged high-sugar diet degrades the hunger-driven feeding response. Lipidomics analysis reveals that longitudinal exposure to high-sugar diets significantly alters whole-body phospholipid profiles. By performing a systematic genetic screen for phospholipid enzymes in adult fly adipocytes, we identify Pect as a critical regulator of hunger-driven feeding. Pect is a rate-limiting enzyme in the phosphatidylethanolamine (PE) biosynthesis pathway and the fly ortholog of human PCYT2. We show that disrupting Pect activity only in the Drosophila fat cells causes insulin resistance, dysregulated lipoprotein delivery to the brain, and a loss of hunger-driven feeding. Previously human studies have noted a correlation between PCYT2/Pect levels and clinical obesity. Now, our unbiased studies in Drosophila provide causative evidence for adipocyte Pect function in metabolic homeostasis. Altogether, we have uncovered that PE phospholipid homeostasis regulates hunger response., Competing Interests: KK, MA, CS, AB, ZG, MP, JD, AR No competing interests declared, (© 2022, Kelly, Alassaf et al.)

Autophagy is a cellular self-digestion process. It delivers cargo to the lysosomes for degradation in response to various stresses, including starvation. The malfunction of autophagy is associated with aging and multiple human diseases. The autophagy machinery is highly conserved-from yeast to humans. The larval fat body of Drosophila melanogaster, an analog for vertebrate liver and adipose tissue, provides a unique model for monitoring autophagy in vivo. Autophagy can be easily induced by nutrient starvation in the larval fat body. Most autophagy-related genes are conserved in Drosophila. Many transgenic fly strains expressing tagged autophagy markers have been developed, which facilitates the monitoring of different steps in the autophagy process. The clonal analysis enables a close comparison of autophagy markers in cells with different genotypes in the same piece of tissue. The current protocol details procedures for (1) generating somatic clones in the larval fat body, (2) inducing autophagy via amino acid starvation, and (3) dissecting the larval fat body, aiming to create a model for analyzing differences in autophagy using an autophagosome marker (GFP-Atg8a) and clonal analysis.

The fat body originates from mesoderm during embryogenesis and it exists through the developmental stages in insects. It is equivalent to vertebrate adipose tissue and liver because it has multiple metabolic and storage functions. The fat body controls the synthesis, storage and metabolism of glycogen, lipid and protein, and it plays a major role in immune and endocrine systems and detoxification processes. Main cells of fat body, which accomplish these vital functions are trophocytes. In this study, we aimed to determine the reserve molecules like glycogen, lipid, protein and uric acid accumulated in the fat body at postembryonic developmental stages of Bombyx mori. For this purpose, we used specific histochemical techniques to determine glycogen, lipid, protein and uric acid molecules. We determined that glycogen contents are stored from the 3rd larval stage while proteins and uric acids are stored from the 4th larval stage. We also detected that the amount of glycogen, lipid, protein and uric acid increase gradually throughout the larval stage and then these molecules decrease gradually as they are used in the pupal stage. Fat body biology may require further investigations on the underlying function of the fat body formation throughout the developmental stages. It can also be used as a model in research of metabolic disorders and immune diseases., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Tumor-suppressive cell competition is an evolutionarily conserved process that selectively removes precancerous cells to maintain tissue homeostasis. Using the polarity-deficiency-induced cell competition model in Drosophila, we identify Toll-6, a Toll-like receptor family member, as a driver of tension-mediated cell competition through α-Spectrin (α-Spec)-Yorkie (Yki) cascade. Toll-6 aggregates along the boundary between wild-type and polarity-deficient clones, where Toll-6 physically interacts with the cytoskeleton network protein α-Spec to increase mechanical tension, resulting in actomyosin-dependent Hippo pathway activation and the elimination of scrib mutant cells. Furthermore, we show that Spz5 secreted from fat body, the key innate organ in fly, facilitates the elimination of scrib clones by binding to Toll-6. These findings uncover mechanisms by which fat bodies remotely regulate tumor-suppressive cell competition of polarity-deficient tumors through inter-organ crosstalk and identified the Toll-6-α-Spec axis as an essential guardian that prevents tumorigenesis via tension-mediated cell elimination., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

The protozoan parasite Babesia spp. invades into tick oocytes and remains in the offspring. The transovarial transmission phenomenon of Babesia in ticks has been demonstrated experimentally, but the molecular mechanisms remain unclear. Babesia invasion into oocytes occurs along with the progression of oogenesis. In the present study, to find the key tick factor(s) for Babesia transmission, we focused on molecules involved in yolk protein precursor (vitellogenin, Vg) synthesis and Vg uptake, which are crucial events in tick oogenesis. With a Haemaphysalis longicornis tick- Babesia ovata experimental model, the expression profiles of Akt , target of rapamycin , S6K , GATA , and Vg , Vg synthesis-related genes, and Vg receptor ( VgR ) and autophagy-related gene 6 ( ATG6 ), Vg uptake-related genes, were analyzed using real-time PCR using tissues collected during the preovipositional period in Babesia -infected ticks. The expression levels of H. longicornis Vg-2 ( HlVg-2 ) and HlVg-3 decreased in the fat body of Babesia -infected ticks 1 day after engorgement. In the ovary, HlVg-2 mRNA expression was significantly higher in Babesia -infected ticks than in uninfected ticks 1 and 2 days after engorgement and decreased 3 days after engorgement. HlVgR expression was significantly lower in Babesia -infected ticks than in uninfected ticks 2 and 4 days after engorgement. HlATG6 had a lower gene expression in Babesia -infected ticks compared to uninfected ticks 2 days after engorgement. Additionally, western blot analysis using protein extracts from each collected tissue revealed that H. longicornis Vg-2 (HlVg-2) accumulate in the fat body and hemolymph of Babesia -infected ticks. These results suggest that Vg uptake from the hemolymph to the ovary was suppressed in the presence of B. ovata . Moreover, HlVg-2 knockdown ticks had a lower detection rate of B. ovata DNA in the ovary and a significant reduction of B. ovata DNA in the hemolymph compared with control ticks. Taken together, our results suggest that accumulated HlVg-2 is associated with Babesia infection or transmission in the tick body. These findings, besides previous reports on VgR, provide important information to elucidate the transovarial transmission mechanisms of pathogens in tick vectors., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kuniyori, Sato, Yokoyama, Kawazu, Xuan, Suzuki, Fujisaki and Umemiya-Shirafuji.)

Zinc deficiency is commonly attributed to inadequate absorption of the metal. Instead, we show that body zinc stores in Drosophila melanogaster depend on tryptophan consumption. Hence, a dietary amino acid regulates zinc status of the whole insect—a finding consistent with the widespread requirement of zinc as a protein cofactor. Specifically, the tryptophan metabolite kynurenine is released from insect fat bodies and induces the formation of zinc storage granules in Malpighian tubules, where 3-hydroxykynurenine and xanthurenic acid act as endogenous zinc chelators. Kynurenine functions as a peripheral zinc-regulating hormone and is converted into a 3-hydroxykynurenine–zinc–chloride complex, precipitating within the storage granules. Thus, zinc and the kynurenine pathway—well-known modulators of immunity, blood pressure, aging, and neurodegeneration—are physiologically connected.

In seasonal breeders, such as amphibians, testicular functions depend on complex processes that change according to seasonality, including Leydig cell (LC) differentiation and lipid-dependent steroidogenesis, extracellular proteins remodeling and actin-dependent cellular dynamics. Speculating that fat bodies (FB) could support some of these processes in L. catesbeianus, we evaluated bilaterally the FB weights, correlating them to testicular parameters such as weight, testosterone (T) immunoexpression, mast cells (MC) number, vascularization and structural proteins. In an attempt to better understand the testicular asymmetry in amphibians, correlations between these different testicular parameters were also established. Right testes (RT), left testes (LT) and associated FB of bullfrogs were weighed, and testes were processed for light and transmission electron microscopy. Collagen content (COL) and MC number were quantified. T and actin immunoexpressions and vascular areas were measured. Statistical analyses and Pearson's correlation were performed. The LT and its associated FB were heavier than the right ones, and showed intense T and actin immunoexpressions, numerous lipid-rich LC, and greater MC number, COL and vascularization than the RT. Positive correlations were detected between: a) FB and testis weights, b) T immunoexpression and testis and FB weights, c) T and actin immunoexpressions and COL. Otherwise, MC number was inversely correlated to T immunoexpression and COL. In right and left sides, the proportional correlation between T immunoexpression and FB weight suggests that FB-stored lipid amount depends on the steroidogenic demand of its associated testis. Thus, the asymmetry in the testes and FB may be associated, at least in part, to the LC steroidogenic activity, which tends to be more intense in LT than in RT. The results also point to a role of COL and mast cells in the LC differentiation and steroidogenesis. Actin was also greater in LT and correlated with T immunoexpression, indicating that the amount of this structural protein depends on androgenic control. Therefore, the testicular asymmetry in bullfrogs seems to be associated to different morphofunctional processes occurring, bilaterally, at different intensities. In this case, there is a tendency of LT, in association with its FB, to be more active than RT. The findings highlight the FB-testis interplay for the comprehension of reproduction in amphibians., (Copyright © 2022 Elsevier GmbH. All rights reserved.)

The major reason of human morbidity and mortality is obesity and related diseases. Brown adipose tissue (BAT) is associated with low total adipose tissue content and a lower risk of type 2 diabetes mellitus. Studies have shown that exercise and cold expose may induce browning. In this study, we verified (1) whether exercise and/or cold exposure can improve the expression level of ucp4c, serca, ampkα, camkII, sirt1, octβ3r , and hamlet ; (2) if these interventions can save cardiac dysfunction induced by a high-fat diet (HFD) in Drosophila. w1118 (wild-type) virgin female flies collected within 8 h after eclosion were divided into eight groups: the normal feed control group (NFD-C), the normal feed exercise group (NFD-E), the normal feed cold exposure group (NFD-CA), the normal feed exercise/cold exposure group (NFD-EC), the HFD control group (HFD-C), the HFD exercise group (HFD-E), the HFD cold exposure group (HFD-CA), and the HFD exercise/cold exposure group (HFD-EC). After exercise and/or cold exposure for 7 days, the mRNA expression levels of ucp4c, serca, ampkα, camk II, sirt1, octβ3r , and hamlet were tested by qRT-PCR, and m-mode was used to assess cardiac function. In addition, we assessed the triacylglycerol (TAG) levels, motor ability, fat mass (by Oil Red O [ORO] staining), and morphological features. The results of TAG, ORO staining, and morphological features all indicate that after interventions, body size of Drosophila was smaller compared with the control group, irrespective of the feeding patterns. The mRNA expression levels of ucp4c, serca, octβ3r, hamlet, ampkα, camkII, and sirt1 were changed to varying degrees under different intervention states (exercise and/or cold exposure). Cold exposure and exercise/cold exposure partly improved cardiac function and the normal fruit flies' cardiac function and exercise ability. However, after exercise intervention, exercise ability and heart function were improved in both HFD and normal-fat diet (NFD) fruit flies. In conclusion, different intervention states (exercise and/or cold exposure) can change the mRNA expression levels of ucp4c, serca, octβ3r, hamlet, ampkα, camkII, and sirt1 . Exercise is the most effective way to restore HFD-induced cardiac dysfunction., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Huang, Jian, Liu, Zheng, Li, Meng, Wang, Zhang, Liu, Guan and Feng.)

For in vivo functional analysis of a protein of interest (POI), multiple transgenic strains with a POI that harbors different tags are needed but generation of these strains is still labor-intensive work. To overcome this, we have developed a versatile Drosophila toolkit with a genetically encoded single-chain variable fragment for the HA epitope tag: 'HA Frankenbody'. This system allows various analyses of HA-tagged POI in live tissues by simply crossing an HA Frankenbody fly with an HA-tagged POI fly. Strikingly, the GFP-mCherry tandem fluorescent-tagged HA Frankenbody revealed a block in autophagic flux and an accumulation of enlarged autolysosomes in the last instar larval and prepupal fat body. Mechanistically, lysosomal activity was downregulated at this stage, and endocytosis, but not autophagy, was indispensable for the swelling of lysosomes. Furthermore, forced activation of lysosomes by fat body-targeted overexpression of Mitf, the single MiTF/TFE family gene in Drosophila, suppressed the lysosomal swelling and resulted in pupal lethality. Collectively, we propose that downregulated lysosomal function in the fat body plays a role in the metamorphosis of Drosophila., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

After a meal, excess nutrients are stored within adipose tissue as triglycerides in structures called lipid droplets. Previous genome-wide RNAi screens have identified that mRNA splicing factor genes are required for normal lipid droplet formation in Drosophila cells. We have previously shown that mRNA splicing factors called serine/arginine-rich (SR) proteins are important for triglyceride storage in the Drosophila fat body. SR proteins shuttle in and out of the nucleus with the help of proteins called Transportins (Tnpo-SR); however, whether this transport is important for SR protein-mediated regulation of lipid storage is unknown. The purpose of this study is to characterize the role of Tnpo-SR proteins in regulating lipid storage in the Drosophila fat body. Decreasing Tnpo-SR in the adult fat body resulted in an increase in triglyceride storage and consistent with this phenotype, Tnpo-SR-RNAi flies also have increased starvation resistance. In addition, the lipid accumulation in Tnpo-SR-RNAi flies is the result of increased triglyceride stored in each fat body cell and not due to increased food consumption. Interestingly, the splicing of CPT1, an enzyme important for the β-oxidation of fatty acids, is altered in Tnpo-SR-RNAi fat bodies. The isoform that produces the less catalytically active form of CPT1 accumulates in fat bodies where Tnpo-SR levels are decreased, suggesting a decrease in lipid breakdown, potentially causing the excess triglyceride storage observed in these flies. Together, these data suggest that the transport of splicing proteins in and out of the nucleus is important for proper triglyceride storage in the Drosophila fat body., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Drosophila has become a popular model for examining the metabolic wasting syndrome, cachexia, characterized by degradation of muscles and fat. Here we present a protocol for quick and consistent scoring of muscle detachment, fat body lipid droplet size, and extracellular matrix (ECM) quantifications in Drosophila larvae. We detail the procedures for dissecting, staining, and imaging third instar Drosophila larval muscle fillets and fat body, and how to utilize FIJI macros for robust quantification of cachectic phenotypes in these dissected tissues. For complete details on the use and execution of this protocol, please refer to Lodge et al. (2021)., Competing Interests: The authors declare no competing interests., (© 2022 The Author(s).)