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Abstract: Changes in splicing patterns are a characteristic of the aging transcriptome; however, it is unclear whether these age‐related changes in splicing facilitate the progressive functional decline that defines aging. In Drosophila, visual behavior declines with age and correlates with altered gene expression in photoreceptors, including downregulation of genes encoding splicing factors. Here, we characterized the significance of these age‐regulated splicing‐associated genes in both splicing and visual function. To do this, we identified differential splicing events in either the entire eye or photoreceptors of young and old flies. Intriguingly, aging photoreceptors show differential splicing of a large number of visual function genes. In addition, as shown previously for aging photoreceptors, aging eyes showed increased accumulation of circular RNAs, which result from noncanonical splicing events. To test whether proper splicing was necessary for visual behavior, we knocked down age‐regulated splicing factors in photoreceptors in young flies and examined phototaxis. Notably, many of the age‐regulated splicing factors tested were necessary for proper visual behavior. In addition, knockdown of individual splicing factors resulted in changes in both alternative splicing at age‐spliced genes and increased accumulation of circular RNAs. Together, these data suggest that cumulative decreases in splicing factor expression could contribute to the differential splicing, circular RNA accumulation, and defective visual behavior observed in aging photoreceptors. [ABSTRACT FROM AUTHOR]

Drosophila melanogaster has been a key model in developing our current understanding of the molecular mechanisms of ageing. Of particular note is its role in establishing the evolutionary conservation of reduced insulin and IGF-1-like signaling in promoting healthy ageing. Capitalizing on its many advantages for experimentation, more recent work has revealed how precise nutritional and genetic interventions can improve fly lifespan without obvious detrimental side effects. We give a brief summary of these recent findings as well as examples of how they may modify ageing via actions in the gut and muscle. These discoveries highlight how expanding our understanding of metabolic and signaling interconnections will provide even greater insight into how these benefits may be harnessed for anti-ageing interventions. [ABSTRACT FROM AUTHOR]

The impact of disruption of adipokinetic hormone (AKH) signaling was studied during aging in Drosophila in a sexually dimorphic manner. A mutant (Akh¹) producing a non-functional AKH peptide was compared with isogenized wild-type controls (w1118), and Akh-rescue line where AKH was ectopically expressed in the mutant background (EE-Akh). Longevity, fecundity, and locomotor activity rhythms remained unaffected by lack of AKH signaling. While the strength of rhythms declined in general with age across all fly lines tested this was more so in case of Akh¹ flies. Negative geotaxis was significantly impaired in Akh¹ flies. Only young Akh¹ flies of both sexes and old Akh¹ females showed significantly higher body weight compared to age-matched iso-control flies (except in case of EE-Akh). Expression of genes involved in energy homeostasis and aging indicated that dTOR and Akt expression were elevated in Akh¹ flies compared to other genotypes, whereas AMPK and dFoxO expression levels were significantly reduced. Multivariate analysis of the distribution of lipid species revealed a significant accumulation of specific diglyceride (DG) and triglyceride (TG) lipid species, irrespective of sex, attributable in part due to lack of AKH. Moreover, irrespective of lack of AKH, older flies of all genotypes accumulated TGs. Taken together, the results strongly suggest that disruption of AKH has very subtle effects on physiology, behavior and lipid status during aging. [ABSTRACT FROM AUTHOR]

The ejaculatory bulb of Drosophila melanogaster males produces proteins and pheromones that play important roles in reproduction. This tissue is also the final mixing site for the ejaculate before transfer to the female. The ejaculatory bulb’s dynamics remain largely unstudied. By microscopy of the ejaculatory bulb in maturing adult males, we observed that the ejaculatory bulb expands in size as males age. Moreover, we document that when males mate, their ejaculatory bulb expands further as ejaculate transfer begins, and then contracts halfway through the course of mating as ejaculate transfer finishes. Although there is some male-to-male variation in the timing of these changes, ultimately the tissue changes in a predictable pattern that gives insight into the active mating process in Drosophila . [ABSTRACT FROM AUTHOR]

Mutations in TDP-43 are associated with proteinaceous inclusions in neurons and are believed to be causative in neurodegenerative diseases such as frontotemporal dementia or amyotrophic lateral sclerosis. Here we describe a Drosophila system where we have engineered the genome to replace the endogenous TDP-43 orthologue with wild type or mutant human TDP-43(hTDP-43). In contrast to other models, these flies express both mutant and wild type hTDP-43 at similar levels to those of the endogenous gene and importantly, no age-related TDP-43 accumulation observed among all the transgenic fly lines. Immunoprecipitation of TDP-43 showed that flies with hTDP-43 mutations had increased levels of ubiquitination and phosphorylation of the hTDP-43 protein. Furthermore, histologically, flies expressing hTDP-43 M337V showed global, robust neuronal staining for phospho-TDP. All three lines: wild type hTDP-43, -G294A and -M337V were homozygous viable, with no defects in development, life span or behaviors observed. The primary behavioral defect was that flies expressing either hTDP-43 G294A or M337V showed a faster decline with age in negative geotaxis. Together, these observations implied that neurons could handle these TDP-43 mutations by phosphorylation- and ubiquitin-dependent proteasome systems, even in a background without the wild type TDP-43. Our findings suggest that these two specific TDP-43 mutations are not inherently toxic, but may require additional environmental or genetic factors to affect longevity or survival. [ABSTRACT FROM AUTHOR]

Aging has significant effects on the locomotor performance of insects including Drosophila . Using a protocol for the high-throughput analysis of fly locomotion in a circular arena, we examined age-dependent behavioral characteristics in adult flies. There are widely used wild-type and genetically engineered background lines including the Canton-S strain and the w1118 strain, which has a null mutation of the white gene. Under standard rearing conditions, we found similar survival and median lifespans in Canton-S (50 days) and w1118 (54 days) strains, however, w1118 flies maintained stable body mass for up to 43 days, whereas Canton-S flies gained body mass at young age, followed by a gradual decline. We also tested the behavioral performance of young and old flies. Compared with young w1118 flies (5–10 days), old w1118 flies (40–45 days) had an increased boundary preference during locomotion in small circular arenas, and increased speed of locomotor recovery from anoxia. Old Canton-S files, however, exhibited unchanged boundary preference and reduced recovery speed from anoxia relative to young flies. In addition, old w1118 flies showed decreased path length per minute and reduced 0.2 s path increment compared with young flies, whereas old Canton-S flies displayed the same path length per minute and the same 0.2 s path increment compared with young flies. We conclude that age-dependent behavioral and physiological changes differ between Canton-S and w1118 flies. These results illustrate that phenotypic differences between strains can change qualitatively, as well as quantitatively, as the animals age. [ABSTRACT FROM AUTHOR]

The Drosophila GeneSwitch system facilitates the spatial and temporal control of gene expression through dietary supplementation of mifepristone (RU486). Because experimental and control groups differ only by treatment with RU486, confounding results from using flies of different genetic backgrounds are eliminated, making GeneSwitch especially useful in studies of aging. However, the effect of RU486 itself on longevity has not been well characterized, particularly in relation to nutritional states known to affect lifespan. Here, we show that RU486 has dose- and diet-dependent effects on longevity in both sexes. On low nutrient diets, RU486 supplementation reduces total food consumption, perhaps exacerbating undernutrition to shorten life. RU486 also inhibits proboscis extension responses to low nutrient diets, suggesting that RU486 has an aversive taste which leads to decreased food consumption and diminished longevity. RU486 is not detrimental to fly lifespan on high nutrient food, correlating with reduced effects of the drug on palatability and total consumption on rich diets. Our results highlight the critical importance of considering how food palatability and nutrient intake might be altered by dietary or drug manipulations in studies of aging and behavior. [ABSTRACT FROM AUTHOR]

Reduced amino acid availability attenuates mRNA translation in cells and helps to extend lifespan in model organisms. The amino acid deprivation-activated kinase GCN2 mediates this response in part by phosphorylating eIF2α. In addition, the cap-dependent translational inhibitor 4E-BP is transcriptionally induced to extend lifespan in Drosophila melanogaster, but through an unclear mechanism. Here, we show that GCN2 and its downstream transcription factor, ATF4, mediate 4E-BP induction, and GCN2 is required for lifespan extension in response to dietary restriction of amino acids. The 4E-BP intron contains ATF4-binding sites that not only respond to stress but also show inherent ATF4 activity during normal development. Analysis of the newly synthesized proteome through metabolic labeling combined with click chemistry shows that certain stress-responsive proteins are resistant to inhibition by 4E-BP, and gcn2 mutant flies have reduced levels of stress-responsive protein synthesis. These results indicate that GCN2 and ATF4 are important regulators of 4E-BP transcription during normal development and aging. [ABSTRACT FROM AUTHOR]

Meningiomas may be classified as neurofibromin 2 (NF2)-associated and non-NF2 meningiomas depending on the presence or absence of molecular alterations in the NF2 gene. One of the characteristic histological features of meningiomas is the whorl formation of neoplastic arachnoid cells. NF2 is a human homolog of the Drosophila gene, Merlin (Mer). In humans, NF2 is the gene responsible for the disease neurofibromatosis type II, which results in the development of brain tumors, including acoustic neurinoma and meningioma. The present study aimed to investigate the molecular pathogenesis of spinal meningioma. It was hypothesized that the whorl formation of meningiomas may occur as a result of a disturbance in the planar cell polarity (PCP) of arachnoid cells, thus, genes understood to govern PCP signaling were analyzed for alterations. Whole exome sequencing followed by Sanger sequencing validation was performed for the analysis of spinal meningioma tissue obtained from a 42-year-old Japanese female. The sequencing identified a nonsynonymous mutation of c.3597G>C, resulting in p.Q1199H, in the FAT atypical cadherin 2 (FAT2) gene. FAT2 is homologous to the Drosophila Fat (Ft) gene, which belongs to the cadherin superfamily. Drosophila Fat is involved in PCP, tumor suppression and Hippo (Hpo) signaling, which is associated with Mer. Taken together, the results of the present study concluded that human FAT2 may function as a key molecule that governs not only PCP, but also NF2-Hpo signaling in arachnoid cells; thus, a mutation in this gene may result in spinal meningioma. [ABSTRACT FROM AUTHOR]

Aging is a risk factor for many human pathologies and is characterized by extensive metabolic changes. Using targeted high-throughput metabolite profiling in Drosophila melanogaster at different ages, we demonstrate that methionine metabolism changes strikingly during aging. Methionine generates the methyl donor S-adenosyl-methionine (SAM), which is converted via methylation to S-adenosyl-homocysteine (SAH), which accumulates during aging. A targeted RNAi screen against methionine pathway components revealed significant life span extension in response to down-regulation of two noncanonical Drosophila homologs of the SAH hydrolase Ahcy (S-adenosyl-L-homocysteine hydrolase [SAHH[) , CG9977/dAhcyL1 and Ahcy89E/CG8956/dAhcyL2, which act as dominant-negative regulators of canonical AHCY. Importantly, tissue-specific down-regulation of dAhcyL1/L2 in the brain and intestine extends health and life span. Furthermore, metabolomic analysis of dAhcyL1 -deficient flies revealed its effect on age-dependent metabolic reprogramming and H3K4 methylation. Altogether, reprogramming of methionine metabolism in young flies and suppression of age-dependent SAH accumulation lead to increased life span. These studies highlight the role of noncanonical Ahcy enzymes as determinants of healthy aging and longevity. [ABSTRACT FROM AUTHOR]

Eukaryotic genomes contain transposable elements ( TE) that can move into new locations upon activation. Since uncontrolled transposition of TEs, including the retrotransposons and DNA transposons, can lead to DNA breaks and genomic instability, multiple mechanisms, including heterochromatin-mediated repression, have evolved to repress TE activation. Studies in model organisms have shown that TEs become activated upon aging as a result of age-associated deregulation of heterochromatin. Considering that different organisms or cell types may undergo distinct heterochromatin changes upon aging, it is important to identify pathways that lead to TE activation in specific tissues and cell types. Through deep sequencing of isolated RNAs, we report an increased expression of many retrotransposons in the old Drosophila fat body, an organ equivalent to the mammalian liver and adipose tissue. This de-repression correlates with an increased number of DNA damage foci and decreased level of Drosophila lamin-B in the old fat body cells. Depletion of the Drosophila lamin-B in the young or larval fat body results in a reduction of heterochromatin and a corresponding increase in retrotransposon expression and DNA damage. Further manipulations of lamin-B and retrotransposon expression suggest a role of the nuclear lamina in maintaining the genome integrity of the Drosophila fat body by repressing retrotransposons. [ABSTRACT FROM AUTHOR]

Metabolic modifications during the developmental period can extend longevity. We found that malic enzyme ( Men ) overexpression during the larval period lengthened the lifespan of Drosophila . Men overexpression by S106-GeneSwitch-Gal4 driver increased pyruvate content and NADPH/NADP + ratio but reduced triglyceride, glycogen, and ATP levels in the larvae. ROS levels increased unexpectedly in Men -overexpressing larvae. Interestingly, adults exposed to larval Men -overexpression maintained ROS tolerance with enhanced expression levels of glutathione-S-transferase D2 and thioredoxin-2. Our results suggest that metabolic changes mediated by Men during development might be related to the control of ROS tolerance and the longevity of Drosophila . [ABSTRACT FROM AUTHOR]

Survival records of longevity experiments are a key component in research on aging. However, surprisingly there have been very few cross-study analyses, besides comparisons of median lifespans or similar summary information. Here, we use a large set of full survival data from various studies to address questions in aging, which are beyond the scope of individual studies. We characterize survival differences between female and male flies of different genetic Drosophila strains, showing significant differences between strains. We further analyse the variation in survival of control cohorts recorded under highly similar conditions within different Drosophila strains. We found that overall transgenic constructs of the UAS/ GAL4 expression system which should have no effect (e.g. a GAL4 construct alone) extend lifespan significantly in the w1118 strain. Using a large data set comprised of various studies, we found no evidence for larger lifespan extensions being associated with shorter lifespans of the control in Drosophila. This demonstrates that lifespan extending treatments are not purely rescuing weak backgrounds. [ABSTRACT FROM AUTHOR]

Highlights: [•] Two NIRS calibration models were developed to estimate the accuracy of age and age structure determination in Drosophila. [•] The composite calibration model was generated by including flies reared under all temperatures and diets. [•] The composite calibration model able to determine flies as less than or equal to 9days, or more than 9days old with 85–97% and 64–99% accuracy, respectively. [•] The simplified calibration models were generated by including flies reared at 23°C on standard diet. [•] Low accuracy rates were observed in simplified calibration models. [ABSTRACT FROM AUTHOR]

Abstract: Methionine sulfoxide reductase B (MsrB) is an enzyme that repairs oxidatively damaged proteins by specifically reducing methionine-R-sulfoxide back to methionine. Three MsrBs, localized in different cellular compartments, are expressed in mammals. However, the physiological roles of each MsrB with regard to its location remain poorly understood. Here, we expressed endoplasmic reticulum (ER)-targeted human MsrB3A (hMsrB3A) in Drosophila and examined its effects on various phenotypes. In two independent transgenic lines, both ubiquitous and neuronal expression of hMsrB3A rendered flies resistant to oxidative stress. Interestingly, these flies also showed significantly enhanced cold and heat tolerance. More strikingly, expression of hMsrB3A in the whole body and nervous system extended the lifespan of fruit flies at 29°C by 43–50% and 12–37%, respectively, suggesting that the targeted expression of MsrB in the ER regulates Drosophila lifespan. A significant increase in lifespan was also observed at 25°C only when hMsrB3A was expressed in neurons. Additionally, hMsrB3A overexpression significantly delayed the age-related decline in locomotor activity and fecundity. Taken together, our data provide evidence that the ER type of MsrB, MsrB3A, plays an important role in protection mechanisms against oxidative, cold and heat stresses and, moreover, in the regulation of fruit fly aging. [Copyright &y& Elsevier]

Damage from reactive oxygen species (ROS) is thought to be a cause of organismal aging. Reactive oxygen species have also been proposed to be responsible for several age-associated phenotypes, including age-related memory impairment (AMI). However, it has not previously been tested whether increasing ROS affects AMI onset. Here we examined the effects of feeding hydrogen peroxide, and the ROS-generating agent, paraquat, on olfactory aversive memory in Drosophila at young ages and during AMI onset. Reactive oxygen species feeding greatly reduced fly survival, and increased oxidized proteins and transcripts of an antioxidant enzyme, catalase (Cat) and a stress-responsive chaperone, heat-shock protein 22 (Hsp22) in fly heads. However, feeding did not impair memory in young wild-type flies, nor did it exacerbate the memory deficits in flies at the onset of AMI. Strikingly ROS feeding did disrupt memory at young ages and accelerated AMI onset was observed when expression of genes involved in the defense system to ROS, including antioxidant enzymes and Hsp22, was reduced in the mushroom bodies, neural centers required for olfactory memory. These results implicate that although ROS production increases upon aging, neuronal functions required for memory processes are sufficiently protected by the defense system to ROS even at the age of AMI onset. Thus we propose that ROS production does not affect AMI onset in Drosophila. [ABSTRACT FROM AUTHOR]

Abstract: Dietary restriction (DR) extends lifespan in diverse organisms and, in animal and cellular models, can delay a range of aging-related diseases including Alzheimer''s disease (AD). A better understanding of the mechanisms mediating these interactions, however, may reveal novel pathways involved in AD pathogenesis, and potential targets for disease-modifying treatments and biomarkers for disease progression. Drosophila models of AD have recently been developed and, due to their short lifespan and susceptibility to genetic manipulation, we have used the fly to investigate the molecular connections among diet, aging and AD pathology. DR extended lifespan in both Arctic mutant Aβ42 and WT 4R tau over-expressing flies, but the underlying molecular pathology was not altered and neuronal dysfunction was not prevented by dietary manipulation. Our data suggest that DR may alter aging through generalised mechanisms independent of the specific pathways underlying AD pathogenesis in the fly, and hence that lifespan-extending manipulations may have varying effects on aging and functional declines in aging-related diseases. Alternatively, our analysis of the specific effects of DR on neuronal toxicity downstream of Aβ and tau pathologies with negative results may simply confirm that the neuro-protective effects of DR are upstream of the initiating events involved in the pathogenesis of AD. [Copyright &y& Elsevier]

Abstract: A remarkable discovery of recent years is that, despite the complexity of ageing, simple genetic interventions can increase lifespan and improve health during ageing in laboratory animals. The pathways involved have often proved to sense nutrients and to match costly activities of organisms, such as growth, metabolism and reproduction, to nutrient status. For instance, the insulin/insulin-like growth factor and Target of Rapamycin signalling network has proved to play a function in ageing, from yeast to mammals, seemingly including humans. In the fruit fly Drosophila, altered activity of several components of this network can increase lifespan and improve locomotor and cardiac function during ageing. The fly brain, fat body (equivalent of mammalian liver and white adipose tissue) and the germ line are important in determination of lifespan, with considerable communication between different tissues. Cellular detoxification pathways, increased autophagy and altered protein synthesis have all been implicated in increased lifespan from reduced IIS/TOR activity, with the role of defence against oxidative stress unresolved. Reduced IIS/TOR signalling can alter or block the response of lifespan to dietary restriction. Reduced IIS can act acutely to lower death rate, implying that it may ameliorate the effects of ageing-related damage, rather than preventing it. [Copyright &y& Elsevier]

Abstract: Since their discovery in Drosophila, the heat shock proteins (Hsps) have been shown to regulate both stress resistance and life-span. Aging is characterized by increased oxidative stress and the accumulation of abnormal (malfolded) proteins, and these stresses induce Hsp gene expression through the transcription factor HSF. In addition, a subset of Hsps is induced by oxidative stress through the JNK signaling pathway and the transcription factor Foxo. The Hsps counteract the toxicity of abnormal proteins by facilitating protein refolding and turnover, and through other mechanisms including inhibition of apoptosis. The Hsps are up-regulated in tissue-specific patterns during aging, and their expression correlates with, and sometimes predicts, life span, making them ideal biomarkers of aging. The tools available for experimentally manipulating gene function and assaying healthspan in Drosophila provides an unparalleled opportunity to further study the role of Hsps in aging. [Copyright &y& Elsevier]

Abstract: Cellular responses to extrinsic and intrinsic insults have to be carefully regulated to properly coordinate cytoprotection, repair processes, cell proliferation and apoptosis. Stress signaling pathways, most prominently the Jun-N-terminal Kinase (JNK) pathway, are critical regulators of such cellular responses and have accordingly been implicated in the regulation of lifespan in various organisms. JNK signaling promotes cytoprotective gene expression, but also interacts with the insulin signaling pathway to influence growth, metabolism, stress tolerance and regeneration. Here, we review recent studies in Drosophila that elucidate the tissue-specific and systemic consequences of JNK activation that ultimately impact lifespan of the organism. [Copyright &y& Elsevier]

Abstract: Understanding how alterations in mitochondrial function in different cells and tissues impacts the aging process remains one of the greatest challenges facing biogerontologists. Here, we discuss the recent upsurge in research in this area using the fruit fly Drosophila melanogaster as a model system. Topics that are discussed include age-related changes in mitochondrial function, mitochondrial oxidative stress and lifespan, life extension mediated by moderate knock-down of genes important for mitochondrial electron transport chain (ETC) function, and the relationship between dietary restriction and ETC activity. Finally, we review recent approaches to supplement the endogenous fly ETC with a single-subunit mitochondrial respiratory enzyme from yeast. [Copyright &y& Elsevier]

Abstract: The hypothesis, that structural deterioration of cytochrome c oxidase (CcO) is a causal factor in the age-related decline in mitochondrial respiratory activity and an increase in H2O2 generation, was tested in Drosophila melanogaster. CcO activity and the levels of seven different nuclear DNA-encoded CcO subunits were determined at three different stages of adult life, namely, young-, middle-, and old-age. CcO activity declined progressively with age by 33%. Western blot analysis, using antibodies specific to Drosophila CcO subunits IV, Va, Vb, VIb, VIc, VIIc, and VIII, indicated that the abundance these polypeptides decreased, ranging from 11% to 40%, during aging. These and previous results suggest that CcO is a specific intra-mitochondrial site of age-related deterioration, which may have a broad impact on mitochondrial physiology. [ABSTRACT FROM AUTHOR]

Abstract: Rab11 mo , a P insertion line of Rab11 showed degenerated ommatidia and excess cell death in larval/pupal eyes. Here, we demonstrate that Rab11 is essential for normal organization of ommatidial cells and their survival in Drosophila, and a mutation in this gene results in cytoskeleton disruption and activation of JNK signaling in the eye. The spatial organization of various cell types in compound eye, viz., cone, photoreceptor, pigment and bristle cells, were disrupted in Rab11 mutants as revealed by immunostaining of F-actin and adherens and septate junction proteins. Genetic interaction studies indicated that mutation in Rab11 upregulates Drosophila apoptotic genes, rpr, hid and grim. In order to study the pathway that causes excessive cell death in Rab11 mutants, the JNK pathway was chosen and genetic interaction analyses were carried out between Rab11 and candidates of the JNK signaling pathway. A downregulation of JNK signaling rescued the phenotype in Rab11 mutant eyes significantly while overexpression of JNK in the eyes using UAS-eiger, UAS-dtak1 or EP(2)0578, resulted in enhancement of the eye phenotype indicating a link between Rab11 and the JNK signaling pathway. [Copyright &y& Elsevier]

Summary Helicase DHX36 plays essential roles in cell development and differentiation at least partially by resolving G-quadruplex (G4) structures. Here we report crystal structures of the Drosophila homolog of DHX36 (DmDHX36) in complex with RNA and a series of DNAs. By combining structural, small-angle X-ray scattering, molecular dynamics simulation, and single-molecule fluorescence studies, we revealed that positively charged amino acids in RecA2 and OB-like domains constitute an elaborate structural pocket at the nucleic acid entrance, in which negatively charged G4 DNA is tightly bound and partially destabilized. The G4 DNA is then completely unfolded through the 3′-5′ translocation activity of the helicase. Furthermore, crystal structures and DNA binding assays show that G-rich DNA is preferentially recognized and in the presence of ATP, specifically bound by DmDHX36, which may cooperatively enhance the G-rich DNA translocation and G4 unfolding. On the basis of these results, a conceptual G4 DNA-resolving mechanism is proposed. [ABSTRACT FROM AUTHOR]