Longevity Of Flies Research Articles

Enhanced Age-Dependent Motor Impairment in Males of Drosophila melanogaster Modeling Spinocerebellar Ataxia Type 1 Is Linked to Dysregulation of a Matrix Metalloproteinase

Over the past two decades, Drosophila melanogaster has proven to be successful in modeling the polyglutamine (polyQ) (caused by CAG repeats) family of neurodegenerative disorders, including the faithful recapitulation of pathological features such as polyQ length-dependent formation of protein aggregates and progressive neuronal degeneration. In this study, pan-neuronal expression of human Ataxin-1 with long polyQ repeat of 82 amino acids was driven using an elav-GAL4 driver line. This would essentially model the polyQ disease spinocerebellar ataxia type 1 (SCA1). Longevity and behavioral analysis of male flies expressing human Ataxin-1 revealed compromised lifespan and accelerated locomotor activity deficits both in diurnal activity and negative geotaxis response compared to control flies. Interestingly, this decline in motor response was coupled to an enhancement of matrix metalloproteinase 1 (dMMP1) expression together with declining expression of extracellular matrix (ECM) fibroblast growth factor (FGF) signaling by hedgehog (Hh) and branchless (bnl) and a significant decrease in expression of survival motor neuron gene (dsmn) in old (30 d) flies. Taken together, our results indicate a role for dysregulation of matrix metalloproteinase in polyQ disease with consequent impact on ECM signaling factors, as well as SMN at the neuromuscular junction causing overt physiological and behavioral deficits.

Evaluation of the Entomopathogenic Fungus Beauveria bassiana on Different Stages of Phlebotomus papatasi (Diptera: Psychodidae), Vector of Zoonotic Cutaneous Leishmaniasis in Iran.

Zoonotic cutaneous leishmaniasis is a major public health problem in Iran with the main vector of Phlebotomus papatasi. The use of entomopathogenic fungi for biological control of the vector is a potential substitute for the current methods which are being used. The purpose of the current study was to assess the virulence of two local isolates of Beauveria bassiana (OZ2 and TV) against Ph. papatasi. To perform the bioassay test, fungal suspensions were applied for every stage of the sand fly life cycle. The mortality rate, longevity, and number of eggs laid were determined. Also, the probability of fungal survival on the surface of rodent's body was assessed. The longevity of infected adult sand flies with both isolates of B. bassiana was significantly lower (P< 0.05) in comparison to the negative control. The estimated Lethal concentration 50 (LC50) values for adult female and male sand flies treated with OZ2 isolate were 1.4×106 and 2.2×107 conidia/ml, respectively, while they were 6.8×106 and 2.3×108 conidia/ml for TV isolate, respectively. Both isolates of B. bassiana exhibited nonsignificant mortality rates in sand fly larvae and pupae and fecundity rate (P> 0.05). According to our findings for both isolates, the fungus continued to spread throughout the surface of the rodent's body for 144 hours after spraying. The current study demonstrated that both isolates of B. bassiana have considerable biological control capacity against adult sand flies.

Sirtuins in kidney health and disease.

Sirtuins (SIRTs) are putative regulators of lifespan in model organisms. Since the initial discovery that SIRTs could promote longevity in nematodes and flies, the identification of additional properties of these proteins has led to understanding of their roles as exquisite sensors that link metabolic activity to oxidative states. SIRTs have major roles in biological processes that are important in kidney development and physiological functions, including mitochondrial metabolism, oxidative stress, autophagy, DNA repair and inflammation. Furthermore, altered SIRT activity has been implicated in the pathophysiology and progression of acute and chronic kidney diseases, including acute kidney injury, diabetic kidney disease, chronic kidney disease, polycystic kidney disease, autoimmune diseases and renal ageing. The renoprotective roles of SIRTs in these diseases make them attractive therapeutic targets. A number of SIRT-activating compounds have shown beneficial effects in kidney disease models; however, further research is needed to identify novel SIRT-targeting strategies with the potential to treat and/or prevent the progression of kidney diseases and increase the average human healthspan.

Kaempferol enhances intestinal repair and inhibits the hyperproliferation of aging intestinal stem cells in Drosophila.

Intestinal stem cells (ISCs) are crucial for tissue repair and homeostasis because of their ability to self-renew and differentiate. However, their functionality declines significantly with age, resulting in reduced tissue regeneration and a higher risk of age-related diseases. Addressing this decline in ISC performance during aging presents a substantial challenge. The specific impact of nutrients or dietary elements on ISC adaptive resizing is urgent to explore. Drosophila ISCs are an ideal model for studying development and aging because of their genetic richness, ease of manipulation, and similarity to mammalian tissues. As the primary mitotically active cells in the Drosophila gut, ISCs are flexible in response to dietary and stress signals. Manipulating signaling pathways or dietary restrictions has shown promise in regulating ISC functions and extending lifespan in flies, these approaches face broader applications for aging research. Kaempferol is well-regarded for its antioxidant, anti-inflammatory, and potential anticancer effects. However, its impacts on ISCs and the associated mechanisms remain inadequately understood. Our findings indicate that Kaempferol accelerates gut recovery after damage and improves the organism's stress tolerance. Moreover, Kaempferol suppresses the hyperproliferation of aging ISCs in Drosophila. Further investigation revealed that the regulatory effects of Kaempferol on ISCs are mediated through the reduction of endoplasmic reticulum (ER) stress in aging flies and the modulation of excessive reactive oxygen species (ROS) levels via ER-stress pathways. Furthermore, Kaempferol exerts regulatory effects on the insulin signaling pathway, thereby contributing to the attenuation of ISC senescence. This study reveals that Kaempferol promotes intestinal homeostasis and longevity in aging flies by targeting ER stress and insulin signaling pathways, though the exact molecular mechanisms require further exploration. Future research will aim to dissect the downstream signaling events involved in these pathways to better understand how Kaempferol exerts its protective effects at the molecular level.

Late-life shift in caloric intake affects fly metabolism and longevity

The prevalence of obesity is increasing in older adults and contributes to age-related decline. Caloric restriction (CR) alleviates obesity phenotypes and delays the onset of age-related changes. However, how late in life organisms benefit from switching from a high-(H) to a low-calorie (L) diet is unclear. We transferred male flies from a H to a L (HL) diet or vice versa (LH) at different times during life. Both shifts immediately change fly rate of aging even when applied late in life. HL shift rapidly reduces fly mortality rate to briefly lower rate than in flies on a constant L diet, and extends lifespan. Transcriptomic analysis uncovers that flies aged on H diet have acquired increased stress response, which may have temporal advantage over flies aged on L diet and leads to rapid decrease in mortality rate after HL switch. Conversely, a LH shift increases mortality rate, which is temporarily higher than in flies aged on a H diet, and shortens lifespan. Unexpectedly, more abundant transcriptomic changes accompanied LH shift, including increase in ribosome biogenesis, stress response and growth. These changes reflect protection from sudden release of ROS, energy storage, and use of energy to growth, which all likely contribute to higher mortality rate. As the beneficial effects of CR on physiology and lifespan are conserved across many organisms, our study provides framework to study underlying mechanisms of CR interventions that counteract the detrimental effects of H diets and reduce rate of aging even when initiated later in life.

The adaptor protein 14-3-3zeta modulates intestinal immunity and aging in Drosophila

The proteins that coordinate the complex transcriptional networks of aging have not been completely documented. Protein 14-3-3zeta is an adaptor protein that coordinates signaling and transcription factor networks, but its function in aging is not fully understood. Here, we showed that the protein expression of 14-3-3zeta gradually increased during aging. High levels of 14-3-3zeta led to shortened lifespan and imbalance of intestinal immune homeostasis in Drosophila, but the decrease in 14-3-3zeta protein levels by RNAi was able to significantly promote the longevity and intestinal immune homeostasis of fruit flies. Importantly, we demonstrate that adult-onset administration of TIC10, a compound that reduces the aging-related AKT and extracellular signal-regulated kinase (ERK) signaling pathways, rescues the shortened lifespan of 14-3-3zeta-overexpressing flies. This funding suggests that 14-3-3zeta plays a critical role in regulating the aging process. Our study elucidates the role of 14-3-3zeta in natural aging and provides the rationale for subsequent 14-3-3zeta-based antiaging research.

Heat-induced hormesis in longevity is linked to heat-stress sensitivity across laboratory populations from diverse altitude of origin in Drosophila buzzatii.

Heat-induced hormesis in longevity is the increase in life span resulting from the previous exposure to a mild heat stress early in life. Here we examined heat-induced hormesis of Drosophila buzzatii in five mass-mating populations, which were derived from five wild populations along an elevation gradient from 202 to 1855m above sea level in North-Western Argentina. Five day old flies were exposed to 37.5°C for 90min to induce hormesis and its possible variation across altitudinal populations. This heat treatment strongly extended longevity in lowland-derived flies from the most heat-resistant population only. Both heat-induced effects on longevity and heat-knockdown time (heat-stress sensitivity) were negatively correlated to altitude of population of origin. Hormesis was positively correlated to heat-knockdown time across populations. These results indicate that variation in heat-induced hormesis can not be considered as independent of heat-stress sensitivity (or heat-knockdown time) in populations of insects.

Bypassing mitochondrial defects rescues Huntington's phenotypes in Drosophila

Huntington's disease (HD) is a fatal neurodegenerative disease with limited treatment options. Human and animal studies have suggested that metabolic and mitochondrial dysfunctions contribute to HD pathogenesis. Here, we use high-resolution respirometry to uncover defective mitochondrial oxidative phosphorylation and electron transfer capacity when a mutant huntingtin fragment is targeted to neurons or muscles in Drosophila and find that enhancing mitochondrial function can ameliorate these defects. In particular, we find that co-expression of parkin, an E3 ubiquitin ligase critical for mitochondrial dynamics and homeostasis, produces significant enhancement of mitochondrial respiration when expressed either in neurons or muscles, resulting in significant rescue of neurodegeneration, viability and longevity in HD model flies. Targeting mutant HTT to muscles results in larger mitochondria and higher mitochondrial mass, while co-expression of parkin increases mitochondrial fission and decreases mass. Furthermore, directly addressing HD-mediated defects in the fly's mitochondrial electron transport system, by rerouting electrons to either bypass mitochondrial complex I or complexes III-IV, significantly increases mitochondrial respiration and results in a striking rescue of all phenotypes arising from neuronal mutant huntingtin expression. These observations suggest that bypassing impaired mitochondrial respiratory complexes in HD may have therapeutic potential for the treatment of this devastating disorder.

Dietary restriction fails to extend life in stressful environments

Abstract Moderate dietary restriction often prolongs life in laboratory animals, and this response has been interpreted as an adaptive strategy that promotes survival during famine. However, dietary restriction can also increase frailty, and it therefore remains unclear whether restricted diets prolong life under stressful conditions like those experienced by wild animals. We manipulated adult dietary protein of Drosophila melanogaster across a gradient of ambient temperature, and examined effects on survival. To test for trade‐offs, we also quantified reproduction, and performance of F1, F2 and F3 descendants. We found that protein restriction increased longevity of one or both sexes at benign ambient temperatures (25°C and 27°C), but failed to extend longevity of flies maintained in cold (21°C and 23°C) or hot (29°C) conditions. Instead, in females, protein restriction resulted in strongly elevated mortality at cold temperatures. Protein restriction also generally reduced reproductive performance, and did not consistently enhance performance of F1, F2 or F3 descendants. Taken together, our results challenge the long‐held idea that extended longevity of diet‐restricted laboratory animals represents an adaptive survival strategy in natural populations. Our findings suggest instead that this response is an artefact of benign laboratory conditions, and that DR‐induced life extension might not be achieved in the more stressful conditions experienced in the wild. Read the free Plain Language Summary for this article on the Journal blog.

Upper thermal limits of Rhagoletis indifferens (Diptera: Tephritidae) pupae and pteromalid parasitoids (Hymenoptera: Pteromalidae) inside fly puparia.

Determining upper thermal limits of tephritid fly pupae can have practical implications for disinfesting soils and for predicting differential impacts of global warming on flies and their parasites. Here, upper thermal limits of Rhagoletis indifferens Curran (Diptera: Tephritidae) pupae and pteromalid wasps (Hymenoptera: Pteromalidae) inside puparia were determined. Puparia receiving sufficient chill to terminate pupal diapause were exposed to temperatures ramped linearly over 6 h from 21 °C to 47.8, 49.4, 51.1, 55.0, or 60.0 °C for a 0-h hold time. Flies eclosed when pupae were exposed to 47.8 °C but not to 49.4, 51.1, 55.0, or 60.0 °C nor in a separate test to 47.8 °C for 1-3 h hold times. All fly pupae in treatments where no eclosion occurred were dead based on puparial dissections. In contrast, adult wasps eclosed when puparia were exposed to 49.4 and 51.1 °C for 0 h and to 47.8 °C for 1- and 2-h hold times. Despite the greater upper thermal limits of wasps, heat delayed eclosion times of both adult flies and wasps, in 47.8 and 51.1 °C treatments, respectively. In separate tests, longevity of flies exposed as pupae to 47.3-48.6 °C was greater than of control flies, while longevity of control wasps and wasps exposed as immatures to 47.8-51.1 °C did not differ. Flies exposed as pupae to 47.2-48.6 °C produced as many eggs and puparia as control flies. Results suggest heat could be used to disinfest soils of puparia while sparing parasitoids. In addition, extreme heat waves due to global warming might be more detrimental to fly pupae than immature wasps.

Towards a Sustainable Management of the Spotted-Wing Drosophila: Disclosing the Effects of Two Spider Venom Peptides on Drosophila suzukii.

The spotted-wing drosophila (Drosophila suzukii) is a polyphagous pest that causes severe damage and economic losses to soft-skinned fruit production. Current control methods are dominated by inefficient cultural practices and broad-spectrum insecticides that, in addition to having toxic effects on non-target organisms, are becoming less effective due to acquired resistance. The increasing awareness of the real impact of insecticides on health and the environment has promoted the exploration of new insecticidal compounds, addressing novel molecular targets. This study explores the efficacy of two orally delivered spider venom peptides (SVPs), J-atracotoxin-Hv1c (Hv1c) and µ-theraphotoxin-Hhn2b (TRTX), to manage D. suzukii, through survival assays and the evaluation of gene expression associated with detoxification pathways. Treatment with TRTX at 111.5 µM for 48 h enhanced fly longevity compared with the control group. Gene expression analysis suggests that detoxification and stress-related mechanisms, such as expression of P450 proteins and apoptotic stimuli signaling, are triggered in D. suzukii flies in response to these treatments. Our results highlight the potential interest of SVPs to control this pest, shedding light on how to ultimately develop improved target-specific formulations.

Genetic Targeting of dSAMTOR, A Negative dTORC1 Regulator, during Drosophila Aging: A Tissue-Specific Pathology.

mTORC1 regulates mammalian cell metabolism and growth in response to diverse environmental stimuli. Nutrient signals control the localization of mTORC1 onto lysosome surface scaffolds that are critically implicated in its amino acid-dependent activation. Arginine, leucine and S-adenosyl-methionine (SAM) can serve as major mTORC1-signaling activators, with SAM binding to SAMTOR (SAM + TOR), a fundamental SAM sensor, preventing the protein's (SAMTOR's) inhibitory action(s) against mTORC1, thereby triggering its (mTORC1) kinase activity. Given the lack of knowledge regarding the role of SAMTOR in invertebrates, we have identified the Drosophila SAMTOR homologue (dSAMTOR) in silico and have, herein, genetically targeted it through the utilization of the GAL4/UAS transgenic tool. Survival profiles and negative geotaxis patterns were examined in both control and dSAMTOR-downregulated adult flies during aging. One of the two gene-targeted schemes resulted in lethal phenotypes, whereas the other one caused rather moderate pathologies in most tissues. The screening of head-specific kinase activities, via PamGene technology application, unveiled the significant upregulation of several kinases, including the dTORC1 characteristic substrate dp70S6K, in dSAMTOR-downregulated flies, thus strongly supporting the inhibitory dSAMTOR action(s) upon the dTORC1/dp70S6K signaling axis in Drosophila brain settings. Importantly, genetic targeting of the Drosophila BHMT bioinformatics counterpart (dBHMT), an enzyme that catabolizes betaine to produce methionine (the SAM precursor), led to severe compromises in terms of fly longevity, with glia-, motor neuron- and muscle-specific dBHMT downregulations exhibiting the strongest effects. Abnormalities in wing vein architectures were also detected in dBHMT-targeted flies, thereby justifying their notably reduced negative geotaxis capacities herein observed mainly in the brain-(mid)gut axis. In vivo adult fly exposure to clinically relevant doses of methionine revealed the mechanistic synergism of decreased dSAMTOR and increased methionine levels in pathogenic longevity, thus rendering (d)SAMTOR an important component in methionine-associated disorders, including homocystinuria(s).

A window into fly sex: mating increases female but reduces male longevity in black soldier flies

A window into fly sex: mating increases female but reduces male longevity in black soldier flies

Effects of two Ayurvedic formulations, Dhanwantaram Kashaya and Saraswatarishta on life history parameters and toxic aggregates in Drosophila models of Huntington’s and Alzheimer's disease

Ethnopharmacological relevanceAyurveda, the traditional healthcare system native to India, employs dosage forms containing multiple herbs in treating various clinical conditions. Dhanwantaram Kashaya (DK) and Saraswatarishta (SA) are two such formulations containing multiple herbs in varied proportions. Kashaya is a liquid decoction while Arishta is fermented liquid with permissible quantity of self-generated alcohol in it. Both the formulations have been reported to have clinical efficacy in age-related memory impairment. Other mental disorders having clinical presentations similar to psychoses are the other indications for these tested formulations. Aim of the studyThe present study was performed to evaluate the efficacy of two Rasayana formulations i.e., DK and SA, used by clinicians in different neurodegenerative conditions. We tested these formulations in Alzheimer's (AD) and Huntington's disease (HD) models of Drosophila melanogaster. Materials and methodInitial experiments looking for life-history parameters in wild-type larvae were carried out in three sets with hundred larvae in each set. These parameters were also studied in diseased models in four sets with eighty larvae in each set. Aβ plaques and polyQ aggregates were looked at with the help of immunostaining technique and images were captured using confocal microscopy. ResultsThe results revealed that 0.25% concentration of both the formulations improve longevity in wild-type flies. Larval development and adult lifespan in Eye-GAL4>Aβ42 (AD) and GMR-GAL4>127Q (HD) larvae/flies reared on 0.25% & 0.50% DK and 0.25% & 1.00% SA improved substantially. Reduced Aβ plaques and polyQ aggregates indicate disease suppression. ConclusionDK and SA enhanced longevity in Drosophila melanogaster. Suppression of disease aggregates suggests their potential utility in treating AD and HD. Further clinical and pharmaceutical studies are required to confirm these results, however, this is a workable model to test multi-herbal formulations of Ayurveda in the forms they are clinically used.

Heterotrophic culture of Spirulina platensis improved its growth and the study of its nutritional effect

With the development of human civilization, many people in the world still suffer from hunger due to wars, epidemics, and other causes such as not being able to afford a healthy diet. As a food, raw material with huge market potential, Spirulina platensis can be used as a new resource for food and has been in large-scale industrialized production worldwide. However, one of the biggest limitations to growing microalgae on an industrial scale is the high cost of the culturing medium. To reduce the culture cost, we hypothesized that S. platensis could grow better in a medium supplemented with glucose. In this study, we investigated the effects of Zarrouk’s medium, Central Food Technological Research Institute (CFTRI) medium, and CFTRI with glucose on the growth rate and biomass of S. platensis. Our results showed that compared with the expensive Zarrouk’s medium containing trace elements, adding a small amount of glucose to the low-cost CFTRI medium significantly increased S. platensis’s growth rate and biomass. Then, to further reduce breeding costs, we continued to explore the growth of S. platensis in a CFTRI medium supplemented with glucose under dark conditions. Our statistical data showed that the heterotrophic glucose consumption of S. platensis was affected by light, and the growth of S. platensis was limited in the absence of light. Finally, we used Drosophila melanogaster to verify the function of S. platensis and found that S. platensis supplementation increased longevity and reproductive ability in fruit flies.

Puerarin extends the lifespan of Drosophila melanogaster by activating autophagy.

Lifespan longevity has attracted increasing attention with societal development. To counter the effects of aging on longevity, we focused on the natural chemicals of plants. In this study, we investigated the effects of puerarin supplementation on the lifespan of Drosophila melanogaster. Puerarin supplementation significantly extended the lifespan of D. melanogaster at 60 μM and 120 μM by upregulating proteasome subunit beta 5 (prosbeta5) and sirtuin-1 (Sirt1). However, puerarin-induced longevity of male flies (F0 generation) may not be passed on to descendants. Additionally, a puerarin diet for 10 and 25 days did not influence the body weight and food intake of male Canton-S flies. Puerarin significantly improved the climbing ability, starvation resistance, and oxidation resistance of male flies by upregulating the expression of Shaker, catalase (CAT), superoxide dismutase 1 (SOD1), and Methuselah, and downregulating poly [ADP-ribose] polymerase (PARP-1) and major heat shock 70 kDa protein Aa (HSP70). Moreover, 120 μM puerarin supplementation for 25 days significantly increased adenosine 5' triphosphate (ATP) content by increasing adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) levels. Additionally, the puerarin diet for 25 days suppressed male fecundity in male flies by decreasing the levels of Bam and Punt. Mechanistically, puerarin enhanced lysosome-involved autophagy by promoting the expression of lysosome markers [β-galactosidase and lysosomal associated membrane protein 1 (LAMP1)], and elevating the levels of autophagy-related genes, including autophagy-associated gene 1 (ATG1), ATG5, and ATG8b. However, puerarin decreased the phosphorylation of the target of rapamycin (TOR) protein. In conclusion, puerarin is a promising compound for improving the longevity of D. melanogaster by activating autophagy.

Longevity of Lucilia sericata (Meigen, 1826) used as pollinator

• Crop growers reliant on insect pollination need additional pollinating species. • The longevity of Lucilia sericata as a pollinator was compared under different enclosure conditions. • Provisioning greenhouses with a sugar + water solution increased fly longevity. Pollinators play an important role in the production of many agricultural products. Honeybees, Apis mellifera L., are leading pollinators, but the number of honeybees in the world is declining. Finding alternatives is beginning to be important. In the present study, we compared the longevity of Lucilia sericata (Meigen, 1826) , which is used as a pollinator, in cages in the laboratory and in a vinyl greenhouse under controlled feeding conditions. First, we showed that the longevity of the flies was significantly extended (c.a. 40 days) in the laboratory when water and sugar were supplied compared to the cases without sugar and/or water. Second, we found that the average longevity was 20 days with water and sugar in cages kept in a vinyl greenhouse during the summer. Finally, we released marked flies into a vinyl greenhouse where strawberries were cultivated without feeding water or sugar in the spring. As a result, fewer than 10% of the flies survived 10 days after release. Based on the result, we discuss the use of this species as a pollinator in the context of its survival rate.

Sex-specific Feeding Behavior of Adult House Flies, Musca domestica L. (Diptera: Muscidae).

House flies, Musca domestica L., (Diptera: Muscidae) mechanically vector diverse disease-causing microorganisms while foraging for food in agricultural and urban habitats. Although flies are diverse feeders, nutrient composition of food is important for both fly longevity and reproduction, especially for anautogenous females who require protein for egg production. We investigated whether fly sex and/or mating status influenced their preference for foods with varying macronutrient composition. Presumably mated or unmated male and female flies were separated by sex and offered four food, each in 10% solution offered on cotton wicks: sugar (carbohydrate-rich), fat-free milk (protein-rich, moderate carbohydrate), egg-yolk (protein and lipid-rich), and water (no macronutrients). Foods were colored with nontoxic dyes, which were rotated between replicates. After 4h exposure, flies were dissected to determine the type of food(s) ingested. The interaction of house fly sex and food type significantly influenced food preference, where females preferred milk (protein and carbohydrate-rich food), and males preferred mainly sugar (carbohydrate-rich). Furthermore, 32.8% of females and 10.6% of males foraged on multiple foods. While interaction of sex and mating status had no effect on food preference, milk preference was significantly higher in presumably mated than unmated females. We also tested whether food color influenced fly feeding preference, and found that color was most significant when flies were offered one food type, but negligible when multiple food types were present. This study suggests that bait-based fly control strategies should consider sex-specific preferences for various food attractants if aiming to target and control both male and female house flies.

Restoration of the GTPase activity and cellular interactions of Gαo mutants by Zn2+ in GNAO1 encephalopathy models.

De novo point mutations in GNAO1, gene encoding the major neuronal G protein Gαo, have recently emerged in patients with pediatric encephalopathy having motor, developmental, and epileptic dysfunctions. Half of clinical cases affect codons Gly203, Arg209, or Glu246; we show that these mutations accelerate GTP uptake and inactivate GTP hydrolysis through displacement Gln205 critical for GTP hydrolysis, resulting in constitutive GTP binding by Gαo. However, the mutants fail to adopt the activated conformation and display aberrant interactions with signaling partners. Through high-throughput screening of approved drugs, we identify zinc pyrithione and Zn2+ as agents restoring active conformation, GTPase activity, and cellular interactions of the encephalopathy mutants, with negligible effects on wild-type Gαo. We describe a Drosophila model of GNAO1 encephalopathy where dietary zinc restores the motor function and longevity of the mutant flies. Zinc supplements are approved for diverse human neurological conditions. Our work provides insights into the molecular etiology of GNAO1 encephalopathy and defines a potential therapy for the patients.

The Role of GCN2 Kinase in Mediating the Effects of Amino Acids on Longevity and Feeding Behaviour in Drosophila.

Restriction of amino acids in the diet can extend lifespan in diverse species ranging from flies to mammals. However, the role of individual amino acids and the underlying molecular mechanisms are only partially understood. The evolutionarily conserved serine/threonine kinase General Control Nonderepressible 2 (GCN2) is a key sensor of amino acid deficiency and has been implicated in the response of lifespan to dietary restriction (DR). Here, we generated a novel Drosophila GCN2 null mutant and analyzed its response to individual amino acid deficiency. We show that GCN2 function is essential for fly development, longevity and feeding behaviour under long-term, but not short-term, deprivation of all individual essential amino acids (EAAs) except for methionine. GCN2 mutants were longer-lived than control flies and showed normal feeding behaviour under methionine restriction. Thus, in flies at least two systems regulate these responses to amino acid deprivation. Methionine deprivation acts via a GCN2-independent mechanism, while all other EAA are sensed by GCN2. Combined deficiency of methionine and a second EAA blocked the response of GCN2 mutants to methionine, suggesting that these two pathways are interconnected. Wild type flies showed a short-term rejection of food lacking individual EAA, followed by a long-term compensatory increase in food uptake. GCN2 mutants also showed a short-term rejection of food deprived of individual EAA, but were unable to mount the compensatory long-term increase in food uptake. Over-expression of the downstream transcription factor ATF4 partially rescued the response of feeding behaviour in GCN2 mutants to amino acid deficiency. Phenotypes of GCN2 mutants induced by leucine and tryptophan, but not isoleucine, deficiency were partially rescued by ATF4 over-expression. The exact function of GCN2 as an amino acid sensor in vivo and the downstream action of its transcription factor effector ATF4 are thus context-specific with respect to the EAA involved.