Background: More than half of human protein-coding genes have an alternative transcription start site (TSS). We aimed to investigate the contribution of alternative TSSs to the acute-stress-induced transcriptome response in human tissue (skeletal muscle) using the cap analysis of gene expression approach. TSSs were examined at baseline and during recovery after acute stress (a cycling exercise). Results: We identified 44,680 CAGE TSS clusters (including 3764 first defined) belonging to 12,268 genes and annotated for the first time 290 TSSs belonging to 163 genes. The transcriptome dynamically changes during the first hours after acute stress; the change in the expression of 10% of genes was associated with the activation of alternative TSSs, indicating differential TSSs usage. The majority of the alternative TSSs do not increase proteome complexity suggesting that the function of thousands of alternative TSSs is associated with the fine regulation of mRNA isoform expression from a gene due to the transcription factor-specific activation of various alternative TSSs. We identified individual muscle promoter regions for each TSS using muscle open chromatin data (ATAC-seq and DNase-seq). Then, using the positional weight matrix approach we predicted time course activation of “classic” transcription factors involved in response of skeletal muscle to contractile activity, as well as diversity of less/un-investigated factors. Conclusions: Transcriptome response induced by acute stress related to activation of the alternative TSSs indicates that differential TSSs usage is an essential mechanism of fine regulation of gene response to stress stimulus. A comprehensive resource of accurate TSSs and individual promoter regions for each TSS in muscle was created. This resource together with the positional weight matrix approach can be used to accurate prediction of TFs in any gene(s) of interest involved in the response to various stimuli, interventions or pathological conditions in human skeletal muscle.

Evaluation of the influence of primary and secondary aging on the manifestation of molecular and cellular hallmarks of aging is a challenging and currently unresolved issue. Our study represents the first demonstration of the distinct role of primary aging and chronic inflammation/physical inactivity – the most important drivers of secondary aging, in the regulation of transcriptomic and proteomic profiles in human skeletal muscle. To achieve this purpose, young healthy people ( n = 15), young ( n = 8) and older ( n = 37) patients with knee/hip osteoarthritis, a model to study the effect of long‐term inactivity and chronic inflammation on the vastus lateralis muscle, were included in the study. It was revealed that widespread and substantial age‐related changes in gene expression in older patients relative to young healthy people (~4000 genes regulating mitochondrial function, proteostasis, cell membrane, secretory and immune response) were related to the long‐term physical inactivity and chronic inflammation rather than primary aging. Primary aging contributed mainly to the regulation of genes (~200) encoding nuclear proteins (regulators of DNA repair, RNA processing, and transcription), mitochondrial proteins (genes encoding respiratory enzymes, mitochondrial complex assembly factors, regulators of cristae formation and mitochondrial reactive oxygen species production), as well as regulators of proteostasis. It was found that proteins associated with aging were regulated mainly at the post‐transcriptional level. The set of putative primary aging genes and their potential transcriptional regulators can be used as a resource for further targeted studies investigating the role of individual genes and related transcription factors in the emergence of a senescent cell phenotype.

Inactivity is associated with the development of numerous disorders. Regular aerobic exercise is broadly used as a key intervention to prevent and treat these pathological conditions. In our meta-analysis we aimed to identify and compare (i) the transcriptomic signatures related to disuse, regular and acute aerobic exercise in human skeletal muscle and (ii) the biological effects and transcription factors associated with these transcriptomic changes. A standardized workflow with robust cut-off criteria was used to analyze 27 transcriptomic datasets for the vastus lateralis muscle of healthy humans subjected to disuse, regular and acute aerobic exercise. We evaluated the role of transcriptional regulation in the phenotypic changes described in the literature. The responses to chronic interventions (disuse and regular training) partially correspond to the phenotypic effects. Acute exercise induces changes that are mainly related to the regulation of gene expression, including a strong enrichment of several transcription factors (most of which are related to the ATF/CREB/AP-1 superfamily) and a massive increase in the expression levels of genes encoding transcription factors and co-activators. Overall, the adaptation strategies of skeletal muscle to decreased and increased levels of physical activity differ in direction and demonstrate qualitative differences that are closely associated with the activation of different sets of transcription factors.

Drosophila melanogaster is one of the most extensively used genetic model organisms for studying LTR retrotransposons that are represented by various groups in its genome. However, the phenomenon of molecular domestication of LTR retrotransposons has been insufficiently studied in Drosophila, as well as in other invertebrates. The present work is devoted to studying the role of the domesticated gag gene, Gagr, in the Drosophila genome. The Gagr gene has been shown to be involved in the response to stress caused by exposure to ammonium persulfate, but not in the stress response to oligomycin A, zeomycin, and cadmium chloride. Ammonium persulfate tissue specifically activates the expression of Gagr in the tissues of the carcass, but not in the gut. We found that the Gagr gene promoter contains one binding motif for the transcription factor kayak, a component of the JNK signaling pathway, and two binding motifs for the transcription factor Stat92E, a component of the Jak-STAT signaling pathway. Remarkably, Gagr orthologs contain the second binding motif for Stat92E only in D. melanogaster, D. simulans and D. sechellia, whereas in D. yakuba and D. erecta, Gagr orthologs contain a single motif, and there are no binding sites for Stat92E in the promoters of Gagr orthologs in D. ananassae and in species outside the melanogaster group. The data obtained indicate the formation of the protective function of the Gagr gene during evolution.

(1) Background: The Gagr gene in Drosophila melanogaster’s genome originated from the molecular domestication of retrotransposons and retroviruses’ gag gene. In all Drosophila species, the Gagr protein homologs exhibit a conserved structure, indicative of a vital role. Previous studies have suggested a potential link between the Gagr gene function and stress responses. (2) Methods: We compared flies with Gagr gene knockdown in all tissues to control flies in physiological tests and RNA-sequencing experiments. (3) Results: Flies with the Gagr gene knockdown exhibited shorter lifespans compared to control flies. Transcriptome analysis revealed that Gagr knockdown flies showed elevated transcription levels of immune response genes. We used ammonium persulfate, a potent stress inducer, to elicit a stress response. In control flies, ammonium persulfate activated the Toll, JAK/STAT, and JNK/MAPK signaling pathways. In contrast, flies with the Gagr gene knockdown displayed reduced expression of stress response genes. Gene ontology enrichment analysis identified categories of genes upregulated under ammonium persulfate stress in control flies but not in Gagr knockdown flies. These genes are involved in developmental control, morphogenesis, and central nervous system function. (4) Conclusion: Our findings indicate the significance of the Gagr gene in maintaining immune response and homeostasis.

The prevention of muscle atrophy carries with it clinical significance for the control of increased morbidity and mortality following physical inactivity. While major transcriptional events associated with muscle atrophy-recovery processes are the subject of active research on the gene level, the contribution of non-coding regulatory elements and alternative promoter usage is a major source for both the production of alternative protein products and new insights into the activity of transcription factors. We used the cap-analysis of gene expression (CAGE) to create a genome-wide atlas of promoter-level transcription in fast (m. EDL) and slow (m. soleus) muscles in rats that were subjected to hindlimb unloading and subsequent recovery. We found that the genetic regulation of the atrophy-recovery cycle in two types of muscle is mediated by different pathways, including a unique set of non-coding transcribed regulatory elements. We showed that the activation of “shadow” enhancers is tightly linked to specific stages of atrophy and recovery dynamics, with the largest number of specific regulatory elements being transcriptionally active in the muscles on the first day of recovery after a week of disuse. The developed comprehensive database of transcription of regulatory elements will further stimulate research on the gene regulation of muscle homeostasis in mammals.