Tissue-Specific Knockdown of Genes of the Argonaute Family Modulates Lifespan and Radioresistance in Drosophila Melanogaster.

Small RNAs are essential to coordinate many cellular processes, including the regulation of gene expression patterns, the prevention of genomic instability, and the suppression of the mutagenic transposon activity. These processes determine the aging, longevity, and sensitivity of cells and an organism to stress factors (particularly, ionizing radiation). The biogenesis and activity of small RNAs are provided by proteins of the Argonaute family. These proteins participate in the processing of small RNA precursors and the formation of an RNA-induced silencing complex. However, the role of Argonaute proteins in regulating lifespan and radioresistance remains poorly explored. We studied the effect of knockdown of Argonaute genes (AGO1, AGO2, AGO3, piwi) in various tissues on the Drosophila melanogaster lifespan and survival after the γ-irradiation at a dose of 700 Gy. In most cases, these parameters are reduced or did not change significantly in flies with tissue-specific RNA interference. Surprisingly, piwi knockdown in both the fat body and the nervous system causes a lifespan increase. But changes in radioresistance depend on the tissue in which the gene was knocked out. In addition, analysis of changes in retrotransposon levels and expression of stress response genes allow us to determine associated molecular mechanisms.

The role of DNA repair genes in radiation-induced adaptive response in Drosophila melanogaster is differential and conditional.

Studies in human and mammalian cell cultures have shown that induction of DNA repair mechanisms is required for the formation of stimulation effects of low doses of ionizing radiation, named "hormesis". Nevertheless, the role of cellular defense mechanisms in the formation of radiation-induced hormesis at the level of whole organism remains poorly studied. The aim of this work was to investigate the role of genes involved in different mechanisms and stages of DNA repair in radioadaptive response and radiation hormesis by lifespan parameters in Drosophila melanogaster. We studied genes that control DNA damage sensing (D-Gadd45, Hus1, mnk), nucleotide excision repair (mei-9, mus210, Mus209), base excision repair (Rrp1), DNA double-stranded break repair by homologous recombination (Brca2, spn-B, okr) and non-homologous end joining (Ku80, WRNexo), and the Mus309 gene that participates in several mechanisms of DNA repair. The obtained results demonstrate that in flies with mutations in studied genes radioadaptive response and radiation hormesis are absent or appear to a lesser extent than in wild-type Canton-S flies. Chronic exposure of γ-radiation in a low dose during pre-imaginal stages of development leads to an increase in expression of the studied DNA repair genes, which is maintained throughout the lifespan of flies. However, the activation of conditional ubiquitous overexpression of DNA repair genes does not induce resistance to an acute exposure to γ-radiation and reinforces its negative impact.

Genome-Protecting Compounds as Potential Geroprotectors.

Throughout life, organisms are exposed to various exogenous and endogenous factors that cause DNA damages and somatic mutations provoking genomic instability. At a young age, compensatory mechanisms of genome protection are activated to prevent phenotypic and functional changes. However, the increasing stress and age-related deterioration in the functioning of these mechanisms result in damage accumulation, overcoming the functional threshold. This leads to aging and the development of age-related diseases. There are several ways to counteract these changes: 1) prevention of DNA damage through stimulation of antioxidant and detoxification systems, as well as transition metal chelation; 2) regulation of DNA methylation, chromatin structure, non-coding RNA activity and prevention of nuclear architecture alterations; 3) improving DNA damage response and repair; 4) selective removal of damaged non-functional and senescent cells. In the article, we have reviewed data about the effects of various trace elements, vitamins, polyphenols, terpenes, and other phytochemicals, as well as a number of synthetic pharmacological substances in these ways. Most of the compounds demonstrate the geroprotective potential and increase the lifespan in model organisms. However, their genome-protecting effects are non-selective and often are conditioned by hormesis. Consequently, the development of selective drugs targeting genome protection is an advanced direction.

The influence of pro-longevity gene Gclc overexpression on the age-dependent changes in Drosophila transcriptome and biological functions.

BACKGROUND: Transcriptional changes that contribute to the organism's longevity and prevent the age-dependent decline of biological functions are not well understood. Here, we overexpressed pro-longevity gene encoding glutamate-cysteine ligase catalytic subunit (Gclc) and analyzed age-dependent changes in transcriptome that associated with the longevity, stress resistance, locomotor activity, circadian rhythmicity, and fertility. RESULTS: Here we reproduced the life extension effect of neuronal overexpression of the Gclc gene and investigated its influence on the age-depended dynamics of transcriptome and biological functions such as fecundity, spontaneous locomotor activity and circadian rhythmicity, as well as on the resistance to oxidative, proteotoxic and osmotic stresses. It was shown that Gclc overexpression reduces locomotor activity in the young and middle ages compared to control flies. Gclc overexpression slowed down the age-dependent decline of locomotor activity and circadian rhythmicity, and resistance to stress treatments. Gclc level demonstrated associations with the expression of genes involved in a variety of cellular processes including Jak-STAT, MAPK, FOXO, Notch, mTOR, TGF-beta signaling pathways, translation, protein processing in endoplasmic reticulum, proteasomal degradation, glycolysis, oxidative phosphorylation, apoptosis, regulation of circadian rhythms, differentiation of neurons, synaptic plasticity and transmission. CONCLUSIONS: Our study revealed that Gclc overexpression induces transcriptional changes associated with the lifespan extension and uncovered pathways that may be associated with the age-dependent decline of biological functions.

A comparison of the transcriptome of Drosophila melanogaster in response to entomopathogenic fungus, ionizing radiation, starvation and cold shock.

BACKGROUND: The molecular mechanisms that determine the organism's response to a variety of doses and modalities of stress factors are not well understood. RESULTS: We studied effects of ionizing radiation (144, 360 and 864 Gy), entomopathogenic fungus (10 and 100 CFU), starvation (16 h), and cold shock (+4, 0 and -4°C) on an organism's viability indicators (survival and locomotor activity) and transcriptome changes in the Drosophila melanogaster model. All stress factors but cold shock resulted in a decrease of lifespan proportional to the dose of treatment. However, stress-factors affected locomotor activity without correlation with lifespan. Our data revealed both significant similarities and differences in differential gene expression and the activity of biological processes under the influence of stress factors. CONCLUSIONS: Studied doses of stress treatments deleteriously affect the organism's viability and lead to different changes of both general and specific cellular stress response mechanisms.

Fucoxanthin increases lifespan of Drosophila melanogaster and Caenorhabditis elegans.

The pharmacological activation of stress-defense mechanisms is one of the perspective ways to increase human lifespan. The goal of the present study was to study the effects on lifespan of Drosophila melanogaster and Caenorhabditis elegans of two carotenoids: ß-carotene and fucoxanthin, which are bioactive natural substances in human diet. In addition, the effects of carotenoids on the flies survival were studied under stress conditions, including starvation, thermal stress (35°C), oxidative stress (20 mM paraquat), as well as locomotor activity, fecundity, and genes expression level. Our results demonstrated lifespan extension of flies by both carotenoids. However, the positive effects on the lifespan of C. elegans were revealed only for fucoxanthin. In presence of carotenoids decreased flies' fecundity, increased spontaneous locomotor activity and resistance to oxidative stress were detected.

Polyphenols as Potential Geroprotectors.

Significance: Currently, a large amount of evidence of beneficial effects of diets enriched with polyphenols on various aspects of health has been accumulated. These phytochemicals have a geroprotective potential slowing down the pathological processes associated with aging and ensuring longevity. In this study, a comprehensive analysis was conducted to determine the adherence of individual polyphenols to geroprotector criteria. Data from experimental models, clinical trials, and epidemiological studies were analyzed. Recent Advances: Sixty-two polyphenols have been described to increase the life span and improve biomarkers of aging in animal models. They act via evolutionarily conserved molecular mechanisms, including hormesis and maintenance of redox homeostasis, epigenetic regulation, response to cellular damage, metabolic control, and anti-inflammatory and senolytic activity. Epidemiological and clinical studies suggest that certain polyphenols have a potential for prevention and treatment of various diseases, including cancer, metabolic disorders, and cardiovascular conditions in humans. Critical Issues: Among the reviewed phytochemicals, chlorogenic acid, quercetin, epicatechin, genistein, resveratrol, and curcumin were identified as compounds with the highest geroprotective potential. However, there is a lack of unambiguous information on the effectiveness and safety of polyphenols for increasing health span, preventing and treating aging-associated diseases in humans. Future Directions: Further research is needed to fully understand the effects of polyphenols considering their long-term consumption, metabolic modification and bioavailability, complex interactions between different groups of polyphenols and with other phytochemicals, as well as their effects on individuals with different health status. Antioxid. Redox Signal. 40, 564-593.

The critical impacts of small RNA biogenesis proteins on aging, longevity and age-related diseases.

Small RNAs and enzymes that provide their biogenesis and functioning are involved in the organism development and coordination of biological processes, including metabolism, maintaining genome integrity, immune and stress responses. In this review, we focused on the role of small RNA biogenesis proteins in determining the aging and longevity of animals and human. A number of studies have revealed that changes in expression profiles of key enzymes, in particular proteins of the Drosha, Dicer and Argonaute families, are associated with the aging process, as well as with some age-related diseases and progeroid syndromes. Down-regulation of small RNA biogenesis proteins leads to global alterations in the expression of regulatory RNAs, disruption of key molecular, cellular and systemic processes, which leads to a lifespan shortening. In contrast, overexpression of Dicer prolongs lifespan and improves cellular defense. Additionally, the role of small RNA biogenesis proteins in the pathogenesis of age-related diseases, including cancer, inflammaging, neurodegeneration, cardiovascular, metabolic and immune disorders, has been conclusively evidenced. Recent advances in biomedicine allow using these proteins as diagnostic and prognostic biomarkers and therapeutic targets.

The Resistance of Drosophila melanogaster to Oxidative, Genotoxic, Proteotoxic, Osmotic Stress, Infection, and Starvation Depends on Age According to the Stress Factor.

We studied how aging affects the ability of Drosophila melanogaster to tolerate various types of stress factors. Data were obtained on the resistance of D. melanogaster to oxidative and genotoxic (separately paraquat, Fe3+, Cu2+, and Zn2+ ions), proteotoxic (hyperthermia, Cd2+ ions), and osmotic (NaCl) stresses, starvation, and infection with the pathological Beauveria bassiana fungus at different ages. In all cases, we observed a strong negative correlation between age and stress tolerance. The largest change in the age-dependent decline in survival occurred under oxidative and osmotic stress. In most experiments, we observed that young Drosophila females have higher stress resistance than males. We checked whether it is possible to accurately assess the biological age of D. melanogaster based on an assessment of stress tolerance. We have proposed a new approach for assessing a biological age of D. melanogaster using a two-parameter survival curve model. For the model, we used an algorithm that evaluated the quality of age prediction for different age and gender groups. The best predictions were obtained for females who were exposed to CdCl2 and ZnCl2 with an average error of 0.32 days and 0.36 days, respectively. For males, the best results were observed for paraquat and NaCl with an average error of 0.61 and 0.68 days, respectively. The average accuracy for all stresses in our model was 1.73 days.

Terpenoids as Potential Geroprotectors.

Terpenes and terpenoids are the largest groups of plant secondary metabolites. However, unlike polyphenols, they are rarely associated with geroprotective properties. Here we evaluated the conformity of the biological effects of terpenoids with the criteria of geroprotectors, including primary criteria (lifespan-extending effects in model organisms, improvement of aging biomarkers, low toxicity, minimal adverse effects, improvement of the quality of life) and secondary criteria (evolutionarily conserved mechanisms of action, reproducibility of the effects on different models, prevention of age-associated diseases, increasing of stress-resistance). The number of substances that demonstrate the greatest compliance with both primary and secondary criteria of geroprotectors were found among different classes of terpenoids. Thus, terpenoids are an underestimated source of potential geroprotectors that can effectively influence the mechanisms of aging and age-related diseases.

Effects of unpaired 1 gene overexpression on the lifespan of Drosophila melanogaster.

BACKGROUND: The JAK/STAT signaling pathway is involved in many aging-related cellular functions. However, effects of overexpression of genes controlling JAK/STAT signal transduction on longevity of model organisms have not been studied. Here we evaluate the effect of overexpression of the unpaired 1 (upd1) gene, which encodes an activating ligand for JAK/STAT pathway, on the lifespan of Drosophila melanogaster. RESULTS: Overexpression of upd1 in the intestine caused a pronounced shortening of the median lifespan by 54.1-18.9%, and the age of 90% mortality by 40.9-19.1% in males and females, respectively. In fat body and in nervous system of male flies, an induction of upd1 overexpression increased the age of 90% mortality and median lifespan, respectively. An increase in upd1 expression enhanced mRNA levels of the JAK/STAT target genes domeless and Socs36E. CONCLUSIONS: Conditional overexpression of upd1 in different tissues of Drosophila imago induces pro-aging or pro-longevity effects in tissue-dependent manner. The effects of upd1 overexpression on lifespan are accompanied by the transcription activation of genes for the components of JAK/STAT pathway.

The Neuronal Overexpression of Gclc in Drosophila melanogaster Induces Life Extension With Longevity-Associated Transcriptomic Changes in the Thorax.

Some effects of aging in animals are tissue-specific. In D. melanogaster neuronal overexpression of Gclc increases lifespan and improves certain physiological parameters associated with health benefits such as locomotor activity, circadian rhythmicity, and stress resistance. Our previous transcriptomic analyses of Drosophila heads, primarily composed of neuronal tissue, revealed significant changes in expression levels of genes involved in aging-related signaling pathways (Jak-STAT, MAPK, FOXO, Notch, mTOR, TGF-beta), translation, protein processing in endoplasmic reticulum, proteasomal degradation, glycolysis, oxidative phosphorylation, apoptosis, regulation of circadian rhythms, differentiation of neurons, synaptic plasticity, and transmission. Considering that various tissues age differently and age-related gene expression changes are tissue-specific, we investigated the effects of neuronal Gclc overexpression on gene expression levels in the imago thorax, which is primarily composed of muscles. A total of 58 genes were found to be differentially expressed between thoraces of control and Gclc overexpressing flies. The Gclc level demonstrated associations with expression of genes involved in the circadian rhythmicity, the genes in categories related to the muscle system process and the downregulation of genes involved in proteolysis. Most of the functional categories altered by Gclc overexpression related to metabolism including Drug metabolism, Metabolism of xenobiotics by cytochrome P450, Glutathione metabolism, Starch and sucrose metabolism, Citrate cycle (TCA cycle), One carbon pool by folate. Thus, the transcriptomic changes caused by neuron-specific Gclc overexpression in the thorax were less pronounced than in the head and affected pathways also differed from previous results. Although these pathways don't belong to the canonical longevity pathways, we suggest that they could participate in the delay of aging of Gclc overexpressing flies.

Overexpression of CBS and CSE genes affects lifespan, stress resistance and locomotor activity in Drosophila melanogaster.

Recent experimental studies highlighted the role of hydrogen sulfide (H2S) in aging and longevity. The cystathionine ß-synthase (CBS) and cystathionine γ-lyase (CSE) are the key enzymes responsible for H2S production. Here we investigated the geroprotective effects of CSE and CBS overexpression in Drosophila. Overexpression of CSE did not affect a lifespan and decrease (mitochondrial form of CSE) or increase (cytoplasmic form of CSE) age dynamics of locomotor activity, while overexpression of CBS increase median (by 12.5%) and maximum (by 6.9%) lifespan and locomotor activity. Increasing of both CSE and CBS expression levels resulted in thermotolerance, but the resistance to combination of arid and food-free conditions decreased. The resistance to oxidative stress (paraquat) was not affected in flies with overexpression of CBS and cytoplasmic CSE, but decreased in flies overexpressing mitochondrial form of CSE. Thus, transgene overexpression of the CSE and CBS in Drosophila induce similar effects on stress-resistance and locomotor activity, however lifespan extending effect was revealed for CBS overexpression only.

The Evaluation of Geroprotective Effects of Selected Flavonoids in Drosophila melanogaster and Caenorhabditis elegans.

Flavonoids is an intensively studied group of natural compounds with antioxidant, antineoplastic, antihyperglycemic, cardioprotective, and neuroprotective properties. The present study intends to investigate the geroprotective action of three selected flavonoids (naringin, luteolin, chrysin) in two model organisms, Caenorhabditis elegans and Drosophila melanogaster. Luteolin and chrysin were shown to improve lifespan parameters when administered to both model organisms. The observed positive effects of these flavonoids in D. melanogaster were limited to females and were not associated with reduced fecundity or locomotor impairment. The life-extending effects of flavonoids were observed in N2 wild-type worms but absent in aak-2(gt33) mutants implying that these effects can be associated with AMP-activated protein kinase activity. Naringin improved lifespan parameters of C. elegans, but had no effect on D. melanogaster females; in some cases, naringin was found to decrease the lifespan of males. Compared to chrysin and luteolin, however, naringin more effectively activates Nrf2 target genes (particularly, GstD1) under oxidative stress. Then we compared molecular mechanisms of studied compounds and a well-known geroprotector rapamycin, using software tool GeroScope. There are no transcriptomic data on luteolin or chrysin provided by LINCS Project database. The bioinformatics comparison of transcriptomics data for A549 and MCF7 human cell lines treated with rapamycin or naringin revealed that these compounds share just a few common signaling pathways and quite distinct in their geroprotective action. Thus, based on C. elegans effects of naringin, luteolin, chrysin on lifespan we have revealed new potential geroprotectors.

Geroprotective and Radioprotective Activity of Quercetin, (-)-Epicatechin, and Ibuprofen in Drosophila melanogaster.

The modulation of longevity genes and aging-associated signaling pathways using pharmacological agents is one of the potential ways to prolong the lifespan and increase the vitality of an organism. Phytochemicals flavonoids and non-steroidal anti-inflammatory drugs have a large potential as geroprotectors. The goal of the present study was to investigate the effects of long-term and short-term consumption of quercetin, (-)-epicatechin, and ibuprofen on the lifespan, resistance to stress factors (paraquat, hyperthermia, γ-radiation, and starvation), as well as age-dependent physiological parameters (locomotor activity and fecundity) of Drosophila melanogaster. The long-term treatment with quercetin and (-)-epicatechin didn't change or decreased the lifespan of males and females. In contrast, the short-term treatment with flavonoids had a beneficial effect and stimulated the resistance to paraquat and acute γ-irradiation. The short-term ibuprofen consumption had a positive effect on the lifespan of females when it was carried out at the middle age (30-40 days), and to the survival of flies under conditions of oxidative and genotoxic stresses. However, it didn't change the lifespan of males and females after the treatment during first 10 days of an imago life. Additionally, quercetin, (-)-epicatechin, and ibuprofen decreased the spontaneous locomotor activity of males, but had no effect of stimulated the physical activity and fecundity of females. Revealed quercetin, (-)-epicatechin, and ibuprofen activity can be associated with the stimulation of stress response mechanisms through the activation of pro-longevity pathways, or the induction of hormesis.

Lifespan and Stress Resistance in Drosophila with Overexpressed DNA Repair Genes.

DNA repair declines with age and correlates with longevity in many animal species. In this study, we investigated the effects of GAL4-induced overexpression of genes implicated in DNA repair on lifespan and resistance to stress factors in Drosophila melanogaster. Stress factors included hyperthermia, oxidative stress, and starvation. Overexpression was either constitutive or conditional and either ubiquitous or tissue-specific (nervous system). Overexpressed genes included those involved in recognition of DNA damage (homologs of HUS1, CHK2), nucleotide and base excision repair (homologs of XPF, XPC and AP-endonuclease-1), and repair of double-stranded DNA breaks (homologs of BRCA2, XRCC3, KU80 and WRNexo). The overexpression of different DNA repair genes led to both positive and negative effects on lifespan and stress resistance. Effects were dependent on GAL4 driver, stage of induction, sex, and role of the gene in the DNA repair process. While the constitutive/neuron-specific and conditional/ubiquitous overexpression of DNA repair genes negatively impacted lifespan and stress resistance, the constitutive/ubiquitous and conditional/neuron-specific overexpression of Hus1, mnk, mei-9, mus210, and WRNexo had beneficial effects. This study demonstrates for the first time the effects of overexpression of these DNA repair genes on both lifespan and stress resistance in D. melanogaster.

Basic mechanisms of longevity: A case study of Drosophila pro-longevity genes.

Drosophila is one of the most convenient model organisms in the genetics of aging and longevity. Unlike the nematodes, which allow for the detection of new pro-aging genes by knockout and RNAi-mediated knock-down, Drosophila also provides an opportunity to find new pro-longevity genes by driver-induced overexpression. Similar studies on other models are extremely rare. In this review, we focused on genes whose overexpression prolongs the life of fruit flies. The majority of longevity-associated genes regulates metabolism and stress resistance, and belongs to the IGF-1R, PI3K, PKB, AMPK and TOR metabolic regulation cluster and the FOXO, HDAC, p53 stress response cluster.