The Effect of the Tau Protein on D. melanogaster Lifespan Depends on GSK3 Expression and Sex.

The microtubule-associated conserved protein tau has attracted significant attention because of its essential role in the formation of pathological changes in the nervous system, which can reduce longevity. The study of the effects caused by tau dysfunction and the molecular mechanisms underlying them is complicated because different forms of tau exist in humans and model organisms, and the changes in protein expression can be multidirectional. In this article, we show that an increase in the expression of the main isoform of the Drosophila melanogaster tau protein in the nervous system has differing effects on lifespan depending on the sex of individuals but has no effect on the properties of the nervous system, in particular, the synaptic activity and distribution of another microtubule-associated protein, Futsch, in neuromuscular junctions. Reduced expression of tau in the nervous system does not affect the lifespan of wild-type flies, but it does increase the lifespan dramatically shortened by overexpression of the shaggy gene encoding the GSK3 (Glycogen Synthase Kinase 3) protein kinase, which is one of the key regulators of tau phosphorylation levels. This effect is accompanied by the normalization of the Futsch protein distribution impaired by shaggy overexpression. The results presented in this article demonstrate that multidirectional changes in tau expression can lead to effects that depend on the sex of individuals and the expression level of GSK3.

Compound combinations targeting longevity: Challenges and perspectives.

Aging is one of the world's greatest concerns, requiring urgent, effective, large-scale interventions to decrease the number of late-life chronic diseases and improve human healthspan. Anti-aging drug therapy is one of the most promising strategies to combat the effects of aging. However, most geroprotective compounds are known to successfully affect only a few aging-related targets. Given this, there is a great biological rationale for the use of combinations of anti-aging interventions. In this review, we characterize the various types of compound combinations used to modulate lifespan, discuss the existing evidence on their role in life extension, and present some key points about current challenges and future prospects for the development of combination drug anti-aging therapy.

Shuttle craft Gene Affects Lifespan of Drosophila melanogaster by Controlling Early Development and Modifying Aging Program.

Fundamental mechanisms underlying genetic control of lifespan are intensively studied and discussed due to the increasing importance of extending healthy human life. The stc gene of the model organism Drosophila melanogaster encodes a transcription factor, homolog of the human transcription factor NF-X1, involved in regulation of neuronal development and other processes, as well as in control of lifespan. In this work, we demonstrate that the stc knockdown in embryonic and nerve cells leads to changes in lifespan, with the nature of changes depending on the cell type and sex of individuals. Based on our results, we suggest that stc gene is involved in transcription regulation throughout life, and, as a result, also affects a complex integral trait, lifespan. At the same time, we show that the reduction of stc expression in neurons can alleviate the negative effect of glutamate on longevity, possibly preventing development of glutamate excitotoxicity, thus modifying the cell death program and preventing death of individuals due to phenoptosis.

Epigenetic enzymes: A role in aging and prospects for pharmacological targeting.

The development of interventions aimed at improving healthspan is one of the priority tasks for the academic and public health authorities. It is also the main objective of a novel branch in biogerontological research, geroscience. According to the geroscience concept, targeting aging is an effective way to combat age-related disorders. Since aging is an exceptionally complex process, system-oriented integrated approaches seem most appropriate for such an interventional strategy. Given the high plasticity and adaptability of the epigenome, epigenome-targeted interventions appear highly promising in geroscience research. Pharmaceuticals targeted at mechanisms involved in epigenetic control of gene activity are actively developed and implemented to prevent and treat various aging-related conditions such as cardiometabolic, neurodegenerative, inflammatory disorders, and cancer. In this review, we describe the roles of epigenetic mechanisms in aging; characterize enzymes contributing to the regulation of epigenetic processes; particularly focus on epigenetic drugs, such as inhibitors of DNA methyltransferases and histone deacetylases that may potentially affect aging-associated diseases and longevity; and discuss possible caveats associated with the use of epigenetic drugs.

Modulated Expression of the Protein Kinase GSK3 in Motor and Dopaminergic Neurons Increases Female Lifespan in Drosophila melanogaster.

Most eukaryotic genes express multiple transcripts and proteins, and a sophisticated gene expression strategy plays a crucial role in ensuring the cell-specificity of genetic information and the correctness of phenotypes. The Drosophila melanogaster gene shaggy encodes several isoforms of the conserved glycogen synthase kinase 3 (GSK3), which is vitally important for multiple biological processes. To characterize the phenotypic effects of differential shaggy expression, we explored how the multidirectional modulation of the expression of the main GSK3 isoform, Shaggy-PB, in different tissues and cells affects lifespan. To this end, we used lines with transgenic constructs that encode mutant variants of the protein. The effect of shaggy misexpression on lifespan depended on the direction of the presumed change in GSK3 activity and the type of tissue/cell. The modulation of GSK3 activity in motor and dopaminergic neurons improved female lifespan but caused seemingly negative changes in the structural (mitochondrial depletion; neuronal loss) and functional (perturbed locomotion) properties of the nervous system, indicating the importance of analyzing the relationship between lifespan and healthspan in invertebrate models. Our findings provide new insights into the molecular and cellular bases of lifespan extension, demonstrating that the fine-tuning of transcript-specific shaggy expression in individual groups of neurons is sufficient to provide a sex-specific increase in survival and slow aging.

Genomic Analysis of European Drosophila melanogaster Populations Reveals Longitudinal Structure, Continent-Wide Selection, and Previously Unknown DNA Viruses.

Genetic variation is the fuel of evolution, with standing genetic variation especially important for short-term evolution and local adaptation. To date, studies of spatiotemporal patterns of genetic variation in natural populations have been challenging, as comprehensive sampling is logistically difficult, and sequencing of entire populations costly. Here, we address these issues using a collaborative approach, sequencing 48 pooled population samples from 32 locations, and perform the first continent-wide genomic analysis of genetic variation in European Drosophila melanogaster. Our analyses uncover longitudinal population structure, provide evidence for continent-wide selective sweeps, identify candidate genes for local climate adaptation, and document clines in chromosomal inversion and transposable element frequencies. We also characterize variation among populations in the composition of the fly microbiome, and identify five new DNA viruses in our samples.

Knockdown of the neuronal gene Lim3 at the early stages of development affects mitochondrial function and lifespan in Drosophila.

Understanding the molecular mechanisms underlying variation in lifespan is central to ensure long life. Lim3 encoding a homolog of the vertebrate Lhx3/4 transcription factors plays a key role in Drosophila neuron development. Here, we demonstrated that Lim3 knockdown early in life decreased survival of adult flies. To study the mechanisms underlying this effect, we identified embryonic Lim3 targets using combined RNA-seq and RT-qPCR analyses complemented by in silico analysis of Lim3 binding sites. Though genes with neuronal functions were revealed as Lim3 targets, the characteristics of neurons were not affected by Lim3 depletion. Many of the direct and indirect Lim3 target genes were associated with mitochondrial function, ATP-related activity, redox processes and antioxidant defense. Consistent with the observed changes in the embryonic transcription of these genes, ROS levels were increased in embryos, which could cause changes in the transcription of indirect Lim3 targets known to affect lifespan. We hypothesize that altered mitochondrial activity is crucial for the decrease of adult lifespan caused by Lim3 knockdown early in life. In adults that encountered Lim3 depletion early in life, the transcription of several genes remained altered, and mitochondrial membrane potential, ATP level and locomotion were increased, confirming the existence of carry-over effects.

Disordered Expression of shaggy, the Drosophila Gene Encoding a Serine-Threonine Protein Kinase GSK3, Affects the Lifespan in a Transcript-, Stage-, and Tissue-Specific Manner.

GSK3 (glycogen synthase kinase 3) is a conserved protein kinase governing numerous regulatory pathways. In Drosophila melanogaster, GSK3 is encoded by shaggy (sgg), which forms 17 annotated transcripts corresponding to 10 protein isoforms. Our goal was to demonstrate how differential sgg transcription affects lifespan, which GSK3 isoforms are important for the nervous system, and which changes in the nervous system accompany accelerated aging. Overexpression of three sgg transcripts affected the lifespan in a stage- and tissue-specific way: sgg-RA and sgg-RO affected the lifespan only when overexpressed in muscles and in embryos, respectively; the essential sgg-RB transcript affected lifespan when overexpressed in all tissues tested. In the nervous system, only sgg-RB overexpression affected lifespan, causing accelerated aging in a neuron-specific way, with the strongest effects in dopaminergic neurons and the weakest effects in GABAergic neurons. Pan-neuronal sgg-RB overexpression violated the properties of the nervous system, including the integrity of neuron bodies; the number, distribution, and structure of mitochondria; cytoskeletal characteristics; and synaptic activity. Such changes observed in young individuals indicated premature aging of their nervous system, which paralleled a decline in survival. Our findings demonstrated the key role of GSK3 in ensuring the link between the pathology of neurons and lifespan.

Reduced Neuronal Transcription of Escargot, the Drosophila Gene Encoding a Snail-Type Transcription Factor, Promotes Longevity.

In recent years, several genes involved in complex neuron specification networks have been shown to control life span. However, information on these genes is scattered, and studies to discover new neuronal genes and gene cascades contributing to life span control are needed, especially because of the recognized role of the nervous system in governing homeostasis, aging, and longevity. Previously, we demonstrated that several genes that encode RNA polymerase II transcription factors and that are involved in the development of the nervous system affect life span in Drosophila melanogaster. Among other genes, escargot (esg) was demonstrated to be causally associated with an increase in the life span of male flies. Here, we present new data on the role of esg in life span control. We show that esg affects the life spans of both mated and unmated males and females to varying degrees. By analyzing the survival and locomotion of the esg mutants, we demonstrate that esg is involved in the control of aging. We show that increased longevity is caused by decreased esg transcription. In particular, we demonstrate that esg knockdown in the nervous system increased life span, directly establishing the involvement of the neuronal esg function in life span control. Our data invite attention to the mechanisms regulating the esg transcription rate, which is changed by insertions of DNA fragments of different sizes downstream of the structural part of the gene, indicating the direction of further research. Our data agree with the previously made suggestion that alterations in gene expression during development might affect adult lifespan, due to epigenetic patterns inherited in cell lineages or predetermined during the development of the structural and functional properties of the nervous system.

Polycomb/Trithorax group-dependent regulation of the neuronal gene Lim3 involved in Drosophila lifespan control.

Molecular mechanisms governing gene expression and defining complex phenotypes are central to understanding the basics of development and aging. Here, we demonstrate that naturally occurring polymorphisms of the Lim3 regulatory region that are associated with variation in gene expression and Drosophila lifespan control are located exclusively in the Polycomb response element (PRE). We find that the Polycomb group (PcG) protein Polycomb (PC) is bound to the PRE only in embryos where Lim3 is present in both repressed and active states. In contrast, the Trithorax group (TrxG) protein absent, small, or homeotic discs 1 (ASH1) is bound downstream of the PRE, to a region adjacent to the Lim3 transcription start site in embryos and adult flies, in which Lim3 is in an active state. Furthermore, mutations in Pc and ash1 genes affect Lim3 expression depending on the structural integrity of the Lim3 PRE, thus confirming functional interactions between these proteins and Lim3 regulatory region. In addition, we demonstrate that the evolutionary conserved Lim3 core promoter provides basic Lim3 expression, whereas structural changes in the Lim3 PRE of distal promoter provide stage-, and tissue-specific Lim3 expression. Therefore, we hypothesize that PcG/TrxG proteins, which are directly involved in Lim3 transcription regulation, participate in lifespan control.

HDAC inhibitors: A new promising drug class in anti-aging research.

In the last years, epigenetic regulation of gene expression is regarded as an important factor involved in a broad spectrum of aging-associated processes including loss of physical activity, frailty and genomic instability, and in development of various pathological conditions such as atherosclerosis, type 2 diabetes, cancer, immune deficits and neurodegenerative diseases. Accordingly, inhibitors of the members of superfamilies of histone deacetylases (HDACs) have been proposed, among other drugs targeting epigenetic pathways, as a promising type of therapeutics that is able to combat aging and its manifestations. The main focus of this article is a review of the literature describing the healthspan-promoting and life-extending effects of inhibitors of HDAC activity in both animal and clinical studies.

Tissue-specific transcription of the neuronal gene Lim3 affects Drosophila melanogaster lifespan and locomotion.

The identity of neuronal cell types is established and maintained by the expression of neuronal genes coding for ion channels, neurotransmitters, and neuropeptides, among others. Some of these genes have been shown to affect lifespan; however, their role in lifespan control remains largely unclear. The Drosophila melanogaster gene Lim3 encodes a transcription factor involved in complicated motor neuron specification networks. We previously identified Lim3 as a candidate gene affecting lifespan. To obtain direct evidence of the involvement of Lim3 in lifespan control, Lim3 overexpression and RNAi knockdown were induced in the nervous system and muscles of Drosophila using the GAL4-UAS binary system. We demonstrated that Lim3 knockdown in the nervous system increased survival at an early age and that Lim3 knockdown in muscles both increased survival at an early age and extended median lifespan, directly establishing the involvement of Lim3 in lifespan control. Lim3 overexpression in nerves and muscles was deleterious and led to lethality and decreased lifespan, respectively. Lim3 misexpression in both nerves and muscles increased locomotion regardless of changes in lifespan, which indicated that the effects of Lim3 on lifespan and locomotion can be uncoupled. Decreased synaptic activity was observed in the neuromuscular junctions of individuals with Lim3 overexpression in muscles, in association with decreased lifespan. However, no changes in NMJ activity were associated with the positive shift in locomotion observed in all misexpression genotypes. Our data suggested that modifications in the microtubule network may be induced by Lim3 misexpression in muscles and cause an increase in locomotion.

Embryonic expression of shuttle craft, a Drosophila gene involved in neuron development, is associated with adult lifespan.

Despite the progress in aging research that highlights the role of the nervous system in longevity, whether genes that control development and consequently structure of the nervous system affect lifespan is unclear. We demonstrated that a mutation inshuttle craft, a gene involved in the nervous system development, increased the lifespan of unmated females and decreased the lifespan of mated females, without affecting males. Precise reversions of the mutation lead to the restoration of the lifespan specific to control females. In mutant unmated females, increased lifespan was associated with elevated locomotion at older ages, indicating slowed aging. In mutant mated females, reproduction was decreased compared to controls, indicating a lack of tradeoff between this trait and lifespan. No differences in shuttle craft transcription were observed between whole bodies, ovaries, and brains of mutant and control females of different ages, either unmated or mated. The amount of shuttle craft transcript appeared to be substantially decreased in mutant embryos. Our results demonstrated that a gene that regulates development of the nervous system might also influence longevity, and thus expanded the spectrum of genes involved in lifespan control. We hypothesize that this "carry-over" effect might be the result of transcription regulation in embryos.

Neuronal inputs and outputs of aging and longevity.

An animal's survival strongly depends on its ability to maintain homeostasis in response to the changing quality of its external and internal environment. This is achieved through intracellular and intercellular communication within and among different tissues. One of the organ systems that plays a major role in this communication and the maintenance of homeostasis is the nervous system. Here we highlight different aspects of the neuronal inputs and outputs of pathways that affect aging and longevity. Accordingly, we discuss how sensory inputs influence homeostasis and lifespan through the modulation of different types of neuronal signals, which reflects the complexity of the environmental cues that affect physiology. We also describe feedback, compensatory, and feed-forward mechanisms in these longevity-modulating pathways that are necessary for homeostasis. Finally, we consider the temporal requirements for these neuronal processes and the potential role of natural genetic variation in shaping the neurobiology of aging.

Endonuclease domain of the Drosophila melanogaster R2 non-LTR retrotransposon and related retroelements: a new model for transposition.

The molecular mechanisms of the transposition of non-long terminal repeat (non-LTR) retrotransposons are not well understood; the key questions of how the 3'-ends of cDNA copies integrate and how site-specific integration occurs remain unresolved. Integration depends on properties of the endonuclease (EN) domain of retrotransposons. Using the EN domain of the Drosophila R2 retrotransposon as a model for other, closely related non-LTR retrotransposons, we investigated the EN domain and found that it resembles archaeal Holliday-junction resolvases. We suggest that these non-LTR retrotransposons are co-transcribed with the host transcript. Combined with the proposed resolvase activity of the EN domain, this model yields a novel mechanism for site-specific retrotransposition within this class of retrotransposons, with resolution proceeding via a Holliday junction intermediate.

Amplification of "defective" retrotransposon gtwin in D. melanogaster strain carrying large complex chromosomal aberration.

Transposable elements (TE) are found in all eukaryotic genomes and play a significant role in their structure and functioning. The majority of mobile elements are silent in the genomes indicating the existence of cell control mechanisms of their activity. Establishment of immunity to TE is of great interest, but it cannot be studied directly and there are only few examples of present or recent active transpositions of mobile elements. G32, a Drosophila melanogaster strain, is characterized by the presence of large complex chromosomal aberration in the 3rd chromosome, active transpositions of gtwin in the past, and its stability at present. To address the question as to what had happened to the element while the cell took it under the control, we performed the detailed cytological and molecular analyses of gtwin's structure and its distribution in G32. Two variants of gtwin were found, one of which is amplified in G32 despite the alteration of tRNA-primer binding site. This element is accumulated in the aberrant chromosome and associated with the inversions breakpoints. Gtwin copies are predominantly localized in euchromatic regions and at least three of them are situated in heterochromatin. One copy was found in the piRNA cluster that might have caused silencing of the element.

A naturally occurring polymorphism at Drosophila melanogaster Lim3 Locus, a homolog of human LHX3/4, affects Lim3 transcription and fly lifespan.

Lim3 encodes an RNA polymerase II transcription factor with a key role in neuron specification. It was also identified as a candidate gene that affects lifespan. These pleiotropic effects indicate the fundamental significance of the potential interplay between neural development and lifespan control. The goal of this study was to analyze the causal relationships between Lim3 structural variations, and gene expression and lifespan changes, and to provide insights into regulatory pathways controlling lifespan. Fifty substitution lines containing second chromosomes from a Drosophila natural population were used to analyze the association between lifespan and sequence variation in the 5'-regulatory region, and first exon and intron of Lim3A, in which we discovered multiple transcription start sites (TSS). The core and proximal promoter organization for Lim3A and a previously unknown mRNA named Lim3C were described. A haplotype of two markers in the Lim3A regulatory region was significantly associated with variation in lifespan. We propose that polymorphisms in the regulatory region affect gene transcription, and consequently lifespan. Indeed, five polymorphic markers located within 380 to 680 bp of the Lim3A major TSS, including two markers associated with lifespan variation, were significantly associated with the level of Lim3A transcript, as evaluated by real time RT-PCR in embryos, adult heads, and testes. A naturally occurring polymorphism caused a six-fold change in gene transcription and a 25% change in lifespan. Markers associated with long lifespan and intermediate Lim3A transcription were present in the population at high frequencies. We hypothesize that polymorphic markers associated with Lim3A expression are located within the binding sites for proteins that regulate gene function, and provide general rather than tissue-specific regulation of transcription, and that intermediate levels of Lim3A expression confer a selective advantage and longer lifespan.