Extension of life span of Drosophila melanogaster by the inhibitors of tryptophan-kynurenine metabolism

Upregulation of kynurenine (KYN) formation from tryptophan (TRY) was associated with aging in animal and human studies. TRY-KYN metabolism is affected by the activities of TRY 2,3-dioxygenase 2 (TDO) and AT P-binding cassette (ABC) transporter regulating TRY access to intracellular TDO. We studied the effects of TDO inhibitor, alpha-methyl tryptophan (aMT) and ABC transported inhibitor, 5-methyl tryptophan (5MT), on the life span of wild strain female Drosophila flies (Oregon-R). aMT and 5MT prolonged mean and maximum life span (by 27% and 43%, and 21% and 23%, resp.). The present results are the first observation of the extension of life span of Drosophila melanogaster by inhibitors of TRY-KYN metabolism, and in line with literature and previous studies on prolonged life span of TDO- and ABC-deficient female Drosophila mutants. Inhibition of TDO and ABC transporter activity might offer the new target for anti-aging interventions.     

Extension of Drosophila melanogaster life span with a GPCR peptide inhibitor

G protein-coupled receptors (GPCRs) mediate signaling from extracellular ligands to intracellular signal transduction proteins. Methuselah (Mth) is a class B (secretin-like) GPCR, a family typified by their large, ligand-binding, N-terminal extracellular domains. Downregulation of mth increases the life span of Drosophila melanogaster; inhibitors of Mth signaling should therefore enhance longevity. We used mRNA display selection to identify high-affinity (K(d) = 15 to 30 nM) peptide ligands that bind to the N-terminal ectodomain of Mth. The selected peptides are potent antagonists of Mth signaling, and structural studies suggest that they perturb the interface between the Mth ecto- and transmembrane domains. Flies constitutively expressing a Mth antagonist peptide have a robust life span extension, which suggests that the peptides inhibit Mth signaling in vivo. Our work thus provides new life span-extending ligands for a metazoan and a general approach for the design of modulators of this important class of GPCRs.     

Lipid peroxidation and the life span of Drosophila melanogaster

Studies have been made on the relationship between incubation temperature (20-30 degrees C) of D. melanogaster and the life span as well as the content of various products of lipid peroxidation. It was shown that the increase in the environmental temperature results in the decrease in the life span, the content of unsaturated fatty acids and conjugated hydroxyperoxids; ketodienic content increases. Strong correlation was observed between the life span and the content of peroxidation products. As it is indicated by coefficients of bifactorial linear regression with interaction, conjugated hydroperoxids and ketodiens exert negative influence on the life span. Their combined effect on the life span is less significant than the sum of their separate effects, which indicates the existence of common "canals" of their influences on the life span.     

Overexpression of glutamate-cysteine ligase extends life span in Drosophila melanogaster

The hypothesis that overexpression of glutamate-cysteine ligase (GCL), which catalyzes the rate-limiting reaction in de novo glutathione biosynthesis, could extend life span was tested in the fruit fly, Drosophila melanogaster. The GAL4-UAS binary transgenic system was used to generate flies overexpressing either the catalytic (GCLc) or modulatory (GCLm) subunit of this enzyme, in a global or neuronally targeted pattern. The GCL protein content of the central nervous system was elevated dramatically in the presence of either global or neuronal drivers. GCL activity was increased in the whole body or in heads, respectively, of GCLc transgenic flies containing global or neuronal drivers. The glutathione content of fly homogenates was increased by overexpression of GCLc or GCLm, particularly in flies overexpressing either subunit globally, or in the heads of GCLc flies possessing neuronal drivers. Neuronal overexpression of GCLc in a long-lived background extended mean and maximum life spans up to 50%, without affecting the rate of oxygen consumption by the flies. In contrast, global overexpression of GCLm extended the mean life span only up to 24%. These results demonstrate that enhancement of the glutathione biosynthetic capability, particularly in neuronal tissues, can extend the life span of flies, and thus support the oxidative stress hypothesis of aging.     

Mutations induced by X-rays and some chemical reagents changing Drosophila melanogaster life span

It has been shown that most of Drosophila melanogaster mutant lines obtained as a result of X-rays irradiation (XI) as well as of the combined action of XI and some chemical agents are characterized by decreased indexes of average (7-40 %) and maximal (1-35 %) life span. Insertion-excision processes at the instable genes white and cut are among the reasons of decreased vitality and shortened life span in induced mutants. Collection of neurodegenerative mutants has been obtained under the influence of ENU. Fast dying of flies and decreased vitality correlated with time point of neurodegenerations in brain structure.     

Radioinduced change in life span of laboratory strains of Drosophila melanogaster

Chronic irradiation (accumulated dose 0.6-0.8 Gy) was shown to change the life span in male Drosophila melanogaster. Death was retarded in wild-type strains and accelerated in mutant strains defective in DNA repair and displaying a higher sensitivity to induction of apoptosis.     

Survival analysis of life span quantitative trait loci in Drosophila melanogaster

We used quantitative trait loci (QTL) mapping to evaluate the age specificity of naturally segregating alleles affecting life span. Estimates of age-specific mortality rates were obtained from observing 51,778 mated males and females from a panel of 144 recombinant inbred lines (RILs). Twenty-five QTL were found, having 80 significant effects on life span and weekly mortality rates. Generation of RILs from heterozygous parents enabled us to contrast effects of QTL alleles with the means of RIL populations. Most of the low-frequency alleles increased mortality, especially at younger ages. Two QTL had negatively correlated effects on mortality at different ages, while the remainder were positively correlated. Chromosomal positions of QTL were roughly concordant with estimates from other mapping populations. Our findings are broadly consistent with a mix of transient deleterious mutations and a few polymorphisms maintained by balancing selection, which together contribute to standing genetic variation in life span.     

Overexpression of glucose-6-phosphate dehydrogenase extends the life span of Drosophila melanogaster

The redox state of tissues tends to become progressively more prooxidizing during the aging process. The hypothesis tested in this study was that enhancement of reductive capacity by overexpression of glucose-6-phosphate dehydrogenase (G6PD), a key enzyme for NADPH biosynthesis, could protect against oxidative stress and extend the life span of transgenic Drosophila melanogaster. Overexpression of G6PD was achieved by combining a UAS-G6PD responder transgene at one of four independent loci with either a broad expression (armadillo-GAL4, Tubulin-GAL4, C23-GAL4, and da-GAL4) or a neuronal driver (D42-GAL4 and Appl-GAL4). The mean life spans of G6PD overexpressor flies were extended, in comparison with driver and responder controls, as follows: armadillo-GAL4 (up to 38%), Tubulin-GAL4 (up to 29%), C23-GAL4 (up to 27%), da-GAL4 (up to 24%), D42-GAL4 (up to 18%), and Appl-GAL4 (up to 16%). The G6PD enzymatic activity was increased, as were the levels of NADPH, NADH, and the GSH/GSSG ratio. Resistance to experimental oxidative stress was enhanced. Furthermore, metabolic rates and fertility were essentially the same in G6PD overexpressors and control flies. Collectively, the results demonstrate that enhancement of the NADPH biosynthetic capability can extend the life span of a relatively long-lived strain of flies, which supports the oxidative stress hypothesis of aging.     

Genotype-environment interaction for quantitative trait loci affecting life span in Drosophila melanogaster

The nature of genetic variation for Drosophila longevity in a population of recombinant inbred lines was investigated by estimating quantitative genetic parameters and mapping quantitative trait loci (QTL) for adult life span in five environments: standard culture conditions, high and low temperature, and heat-shock and starvation stress. There was highly significant genetic variation for life span within each sex and environment. In the analysis of variance of life span pooled over sexes and environments, however, the significant genetic variation appeared in the genotype x sex and genotype x environment interaction terms. The genetic correlation of longevity across the sexes and environments was not significantly different from zero in these lines. We estimated map positions and effects of QTL affecting life span by linkage to highly polymorphic roo transposable element markers, using a multiple-trait composite interval mapping procedure. A minimum of 17 QTL were detected; all were sex and/or environment-specific. Ten of the QTL had sexually antagonistic or antagonistic pleiotropic effects in different environments. These data provide support for the pleiotropy theory of senescence and the hypothesis that variation for longevity might be maintained by opposing selection pressures in males and females and variable environments. Further work is necessary to assess the generality of these results, using different strains, to determine heterozygous effects and to map the life span QTL to the level of genetic loci.     

Induced overexpression of mitochondrial Mn-superoxide dismutase extends the life span of adult Drosophila melanogaster

A transgenic system ("FLP-out") based on yeast FLP recombinase allowed induced overexpression of MnSOD enzyme in adult Drosophila melanogaster. With FLP-out a brief heat pulse (HP) of young, adult flies triggered the rearrangement and subsequent expression of a MnSOD transgene throughout the adult life span. Control (no HP) and overexpressing (HP) flies had identical genetic backgrounds. The amount of MnSOD enzyme overexpression achieved varied among six independent transgenic lines, with increases up to 75%. Life span was increased in proportion to the increase in enzyme. Mean life span was increased by an average of 16%, with some lines showing 30-33% increases. Maximum life span was increased by an average of 15%, with one line showing as much as 37% increase. Simultaneous overexpression of catalase with MnSOD had no added benefit, consistent with previous observations that catalase is present in excess in the adult fly with regard to life span. Cu/ZnSOD overexpression also increases mean and maximum life span. For both MnSOD and Cu/ZnSOD lines, increased life span was not associated with decreased metabolic activity, as measured by O2 consumption.     

High-resolution mapping of quantitative trait loci affecting increased life span in Drosophila melanogaster

Limited life span and senescence are near-universal characteristics of eukaryotic organisms, controlled by many interacting quantitative trait loci (QTL) with individually small effects, whose expression is sensitive to the environment. Analyses of mutations in model organisms have shown that genes affecting stress resistance and metabolism affect life span across diverse taxa. However, there is considerable segregating variation for life span in nature, and relatively little is known about the genetic basis of this variation. Replicated lines of Drosophila that have evolved increased longevity as a correlated response to selection for postponed senescence are valuable resources for identifying QTL affecting naturally occurring variation in life span. Here, we used deficiency complementation mapping to identify at least 11 QTL on chromosome 3 that affect variation in life span between five old (O) lines selected for postponed senescence and their five base (B) population control lines. Most QTL were sex specific, and all but one affected multiple O lines. The latter observation is consistent with alleles at intermediate frequency in the base population contributing to the response to selection for postponed senescence. The QTL were mapped with high resolution and contained from 12 to 170 positional candidate genes.     

Ethanol extract of Cassia siamea L. increases life span in Drosophila melanogaster

The stem of Cassia siamea L. (Fabaceae) has been used in traditional Thai medicine as a longevity remedy. The objective of this study was to investigate the effect of ethanolic stem extract of C. siamea (CSE) on the life span of Drosophila melanogaster. The results showed that a diet containing 10 mg/mL CSE could significantly extend the mean life span of D. melanogaster by 14% compared with the control diet (P < 0.01). The maximum life span was 74, 78, and 84 days in control, CSE (5 mg/mL) and CSE (10 mg/mL) groups, respectively. Supplementation of CSE at 10 mg/mL also significantly increases the activity of superoxide dismutase (SOD) and catalase (CAT) at days 25 and 40 compared with the control diet. Treatment of CSE at 5 and 10 mg/mL significantly increased the climbing ability of D. melanogaster both on days 25 and 40 compared with the control flies. Paraquat and H₂O₂ challenge test showed that flies fed with CSE at 10 mg/mL had a longer survival time than the control flies (P < 0.01). This study provides supportive evidence that supplementation with CSE prolonged life span and reduced oxidative stress in D. melanogaster.     

High-frequency generation of conditional mutations affecting Drosophila melanogaster development and life span.

Genome sequencing reveals that a large percentage of Drosophila genes have homologs in humans, including many human disease genes. The goal of this research was to develop methods to efficiently test Drosophila genes for functions in vivo. An important challenge is the fact that many genes function at more than one point during development and during the life cycle. Conditional expression systems such as promoters regulated by tetracycline (or its derivative doxycycline) are often ideal for testing gene functions. However, generation of transgenic animals for each gene of interest is impractical. Placing the doxycycline-inducible ("tet-on") promoter directed out of the end of the P transposable element produced a mobile, doxycycline-inducible promoter element, named PdL. PdL was mobilized to 228 locations in the genome and was found to generate conditional (doxycycline-dependent), dominant mutations at high frequency. The temporal control of gene overexpression allowed generation of mutant phenotypes specific to different stages of the life cycle, including metamorphosis and aging. Mutations characterized included inserts in the alpha-mannosidase II (dGMII), ash1, and pumilio genes. Novel phenotypes were identified for each gene, including specific developmental defects and increased or decreased life span. The PdL system should facilitate testing of a large fraction of Drosophila genes for overexpression and misexpression phenotypes at specific developmental and life cycle stages.     

Genes regulating the development and functioning of the nervous system determine life span in Drosophila melanogaster

Earlier, it has been shown that genes responsible for differences in longevity between wild-type Drosophila melanogaster lines 2b and Oregon are localized in region 7A6-B2, 36E4-37B9, 37B9-D2, and 64C-65C. Quantitative complementation tests were conducted between the gene mutations localized in these regions and involved in catecholamine biosynthesis (iav (inactive), Catsup (Catecholamines up), amd (alpha methyl dopa resistant), Dox-A2 (Diphenol oxidase A2), pie (pale)) and neuron development control (Fas3 (Fascyclin 3), tup (tail up), Lim3), on the one hand, and two different normal alleles of these genes in lines 2b and Oregon, on the other. Complementation was found for genes iav, Fas3, amd and ple. The remaining genes (Catsup, Dox-A2, tup, and Lim3) are candidate genes for controlling differences in longevity between lines 2b and Oregon.     

Neuronal expression of p53 dominant-negative proteins in adult Drosophila melanogaster extends life span

Hyperactivation of p53 leads to a reduction in tumor formation and an unexpected shortening of life span in two different model systems . The decreased life span occurs with signs of accelerated aging, such as osteoporosis, reduction in body weight, atrophy of organs, decreased stress resistance, and depletion of hematopoietic stem cells. These observations suggest a role for p53 in the determination of life span and the speculation that decreasing p53 activity may result in positive effects on some aging phenotypes . In this report, we show that expression of dominant-negative versions of Drosophila melanogaster p53 in adult neurons extends life span and increases genotoxic stress resistance in the fly. Consistent with this, a naturally occurring allele with decreased p53 activity has been associated with extended survival in humans . Expression of the dominant-negative Drosophila melanogaster p53 constructs does not further increase the extended life span of flies that are calorie restricted, suggesting that a decrease in p53 activity may mediate a component of the calorie-restriction life span-extending pathway in flies.     

Longevity and metabolism in Drosophila melanogaster: genetic correlations between life span and age-specific metabolic rate in populations artificially selected for long life

We measured age-specific metabolic rates in 2861 individual Drosophila melanogaster adult males to determine how genetic variation in metabolism is related to life span. Using recombinant inbred (RI) lines derived from populations artificially selected for long life, resting metabolic rates were measured at 5, 16, 29, and 47 days posteclosion, while life spans were measured in the same genotypes in mixed-sex population cages and in single-sex vials. We observed much heritable variation between lines in age-specific metabolic rates, evidence for genotype x age interaction, and moderate to large heritabilities at all ages except the youngest. Four traits exhibit evidence of coordinate genetic control: day 16 and day 29 metabolic rates, life span in population cages, and life span in vials. Quantitative trait loci (QTL) for those traits map to the same locations on three major chromosomes, and additive genetic effects are all positively correlated. In contrast, metabolic rates at the youngest and oldest ages are unrelated to metabolic rates at other ages and to survival. We suggest that artificial selection for long life via delayed reproduction also selects for increased metabolism at intermediate ages. Contrary to predictions of the "rate of living" theory, we find no evidence that metabolic rate varies inversely with survival, at the level of either line means or additive effects of QTL.     

Within-population variation in cytoplasmic genes affects female life span and aging in Drosophila melanogaster

It has been suggested that mitochondrial DNA (mtDNA) may play an important role in aging. Yet, few empirical studies have tested this hypothesis, partly because the degree of sequence polymorphism in mtDNA is assumed to be low. However, low sequence variation may not necessarily translate into low phenotypic variation. Here, we report an experiment that tests whether there is within-population variation in cytoplasmic genes for female longevity and senescence. To achieve this, we randomly selected 25 "mitochondrial founders" from a single, panmictic population of Drosophila melanogaster and used these founders to generate distinct "mt" lines in which we controlled for the nuclear background by successive backcrossing. Potential confounding effects of cytoplasmically transmitted bacteria were eliminated by tetracycline treatment. The mt lines were then assayed for differences in longevity, Gompertz intercept (frailty), and demographic rate of change in mortality with age (rate-of-senescence) in females. We found significant cytoplasmic effects on all three variables. This provides evidence that genetic variation in cytoplasmic genes, presumably mtDNA, contributes to variation in female mortality and aging.     

Juvenile hormone as a regulator of the trade-off between reproduction and life span in Drosophila melanogaster

Trade-offs between reproduction and life span are ubiquitous, but little is known about their underlying mechanisms. Here we combine treatment with the juvenile hormone analog (JHa) methoprene and experimental evolution in Drosophila melanogaster to study the potential role of juvenile hormone (JH) in mediating such trade-offs at both the physiological and evolutionary level. Exposure to JHa in the larval medium (and up to 24 h posteclosion) increased early life fecundity but reduced life span of normal (unselected) flies, supporting the physiological role of JH in mediating the trade-off. This effect was much smaller for life span, and not detectable for fecundity, in fly lines previously bred for 19 generations on a medium containing JHa. Furthermore, these selection lines lived longer than unselected controls even in the absence of JHa treatment, without a detectable reduction in early life fecundity. Thus, selection for resistance to JHa apparently induced some evolutionary changes in JH metabolism or signaling, which led to longer life span as a correlated response. This supports the hypothesis that JH may mediate evolution of longer life span, but--contrary to our expectation-this apparently does not need to trade--off with fecundity.     

Biotin deficiency decreases life span and fertility but increases stress resistance in Drosophila melanogaster

Biotin deficiency is associated with fetal malformations and activation of cell survival pathways in mammals. In this study we determined whether biotin status affects life span, stress resistance, and fertility in the fruit fly Drosophila melanogaster. Male and female flies of the Canton-S strain had free access to diets containing 6.0 (control), 4.8, 2.5, or 0 pmol biotin/100 mg. Biotin concentrations in diets correlated with activities of biotin-dependent propionyl-CoA carboxylase and biotin concentrations in fly homogenates, but not with biotinylation of histones (DNA-binding proteins). Propionyl-CoA carboxylase activities and biotin concentrations were lower in males than in females fed diets low in biotin. The life span of biotin-deficient males and females was up to 30% shorter compared to biotin-sufficient controls. Exposure to oxidative stress reversed the effects of biotin status on survival in male flies: survival times increased by 40% in biotin-deficient males compared to biotin-sufficient controls. Biotin status did not affect survival of females exposed to oxidative stress. Exposure of flies to cold, heat, and oxidative stress was associated with mobilization of biotin from yet unknown sources. Biotin deficiency decreased fertility of flies. When biotin-deficient males and females were mated, the hatching rate (larvae hatched per egg) decreased by about 28% compared to biotin-sufficient controls. These findings are consistent with the hypothesis that biotin affects life span, stress resistance, and fertility in fruit flies.