Graphical Abstract Highlights d SIRT6 KO mice accumulate L1 cDNA, triggering interferon response via cGAS pathway d Wild-type aged mice accumulate L1 cDNA and display type I interferon response d Reverse-transcriptase inhibitors rescue type I interferon response and DNA damage d Reverse-transcriptase inhibitors extend lifespan and improve health of SIRT6 KO mice SUMMARYMice deficient for SIRT6 exhibit a severely shortened lifespan, growth retardation, and highly elevated LINE1 (L1) activity. Here we report that SIRT6-deficient cells and tissues accumulate abundant cytoplasmic L1 cDNA, which triggers strong type I interferon response via activation of cGAS.Remarkably, nucleoside reverse-transcriptase inhibitors (NRTIs), which inhibit L1 retrotransposition, significantly improved health and lifespan of SIRT6 knockout mice and completely rescued type I interferon response. In tissue culture, inhibition of L1 with siRNA or NRTIs abrogated type I interferon response, in addition to a significant reduction of DNA damage markers. These results indicate that L1 activation contributes to the pathologies of SIRT6 knockout mice. Similarly, L1 transcription, cytoplasmic cDNA copy number, and type I interferons were elevated in the wild-type aged mice. As sterile inflammation is a hallmark of aging, we propose that modulating L1 activity may be an important strategy for attenuating age-related pathologies. Context and SignificanceMammalian aging is complex and likely reflects accumulated damage to our genes/DNA. Retrotransposons are a special class of parasitic genetic elements that can replicate their DNA within our genes, at times amounting to up to 20% of human DNA. Retrotransposons, such as the commonly occurring L1, have been associated with aging, neurodegeneration, and cancer. University of Rochester scientists uncovered L1 retrotransposons as the culprit in many aspects of accelerated aging in mice, a model for human aging. They also linked these special gene elements to inflammation. Experimentally blocking retrotransposon amplification improved the health and lifespan of mice. Although there is a long road ahead, inhibiting retrotransposon activity, and the related inflammation, could eventually be a therapy for age-related diseases.
Genomic instability is a hallmark of aging tissues. Genomic instability may arise from the inefficient or aberrant function of DNA double-stranded break (DSB) repair. DSBs are repaired by homologous recombination (HR) and nonhomologous DNA end joining (NHEJ). HR is a precise pathway, whereas NHEJ frequently leads to deletions or insertions at the repair site. Here, we used normal human fibroblasts with a chromosomally integrated HR reporter cassette to examine the changes in HR efficiency as cells progress to replicative senescence. We show that HR declines sharply with increasing replicative age, with an up to 38-fold decrease in efficiency in presenescent cells relative to young cells. This decline is not explained by a reduction of the number of cells in S/G 2 /M stage as presenescent cells are actively dividing. Expression of proteins involved in HR such as Rad51, Rad51C, Rad52, NBS1, and Sirtuin 6 (SIRT6) diminished with cellular senescence. Supplementation of Rad51, Rad51C, Rad52, and NBS1 proteins, either individually or in combination, did not rescue the senescence-related decline of HR. However, overexpression of SIRT6 in "middle-aged" and presenescent cells strongly stimulated HR repair, and this effect was dependent on mono-ADP ribosylation activity of poly(ADP-ribose) polymerase (PARP1). These results suggest that in aging cells, the precise HR pathway becomes repressed giving way to a more error-prone NHEJ pathway. These changes in the processing of DSBs may contribute to age-related genomic instability and a higher incidence of cancer with age. SIRT6 activation provides a potential therapeutic strategy to prevent the decline in genome maintenance.A ging is associated with an increased mutation rate (1) and the appearance of genomic rearrangements (2). The accumulation of mutations and rearrangements is a contributing cause of aging and leads to a decline of tissue functionality and an increased incidence of tumors. These mutations and genomic rearrangements arise from aberrant repair of DNA doublestranded breaks (DSBs).DSBs are dangerous DNA lesions. If left unrepaired or repaired incorrectly, DSBs result in a massive loss of genetic information, chromosomal aberrations, or cell death. DSBs are repaired by two major pathways: nonhomologous end joining (NHEJ) and homologous recombination (HR) (3). NHEJ modifies the broken DNA ends and ligates them together with no requirement for homology, often generating deletions or insertions (4). In contrast, HR uses an undamaged DNA template to repair the break, leading to the reconstitution of the original sequence (5). HR repair is responsible for approximately one quarter of DNA repair events and has much slower repair kinetics than NHEJ (6). HR repair begins with the MRE11, NBS1, and Rad50 complex binding to DNA ends and mediating end resection. The RPA protein is then recruited to DNA ends, in a process regulated by CtIP (7). Once the ends are resected, Rad51 forms nucleoprotein filaments and mediates strand invasion of the filament into duplex DNA, usual...
The naked mole-rat (Heterocephalus glaber) is a subterranean eusocial rodent with a markedly long lifespan and resistance to tumorigenesis. Multiple data implicate modulation of protein translation in longevity. Here we report that 28S ribosomal RNA (rRNA) of the naked mole-rat is processed into two smaller fragments of unequal size. The two breakpoints are located in the 28S rRNA divergent region 6 and excise a fragment of 263 nt. The excised fragment is unique to the naked mole-rat rRNA and does not show homology to other genomic regions. Because this hidden break site could alter ribosome structure, we investigated whether translation rate and amino acid incorporation fidelity were altered. We report that naked mole-rat fibroblasts have significantly increased translational fidelity despite having comparable translation rates with mouse fibroblasts. Although we cannot directly test whether the unique 28S rRNA structure contributes to the increased fidelity of translation, we speculate that it may change the folding or dynamics of the large ribosomal subunit, altering the rate of GTP hydrolysis and/or interaction of the large subunit with tRNA during accommodation, thus affecting the fidelity of protein synthesis. In summary, our results show that naked mole-rat cells produce fewer aberrant proteins, supporting the hypothesis that the more stable proteome of the naked mole-rat contributes to its longevity.aging | NMR
The naked mole rat (Heterocephalus glaber) is a long-lived and tumor-resistant rodent. Tumor resistance in the naked mole rat is mediated by the extracellular matrix component hyaluronan of very high molecular weight (HMW-HA). HMW-HA triggers hypersensitivity of naked mole rat cells to contact inhibition, which is associated with induction of the INK4 (inhibitors of cyclin dependent kinase 4) locus leading to cell-cycle arrest. The INK4a/b locus is among the most frequently mutated in human cancer. This locus encodes three distinct tumor suppressors: p15 INK4b , p16 INK4a , and ARF (alternate reading frame). Although p15 INK4b has its own ORF, p16 INK4a and ARF share common second and third exons with alternative reading frames. Here, we show that, in the naked mole rat, the INK4a/b locus encodes an additional product that consists of p15 INK4b exon 1 joined to p16 INK4a exons 2 and 3. We have named this isoform pALT INK4a/b (for alternative splicing). We show that pALT INK4a/b is present in both cultured cells and naked mole rat tissues but is absent in human and mouse cells. Additionally, we demonstrate that pALT INK4a/b expression is induced during early contact inhibition and upon a variety of stresses such as UV, gamma irradiation-induced senescence, loss of substrate attachment, and expression of oncogenes. When overexpressed in naked mole rat or human cells, pALT INK4a/b has stronger ability to induce cell-cycle arrest than either p15 INK4b or p16 INK4a . We hypothesize that the presence of the fourth product, pALT INK4a/b of the INK4a/b locus in the naked mole rat, contributes to the increased resistance to tumorigenesis of this species.naked mole rat | INK4 | p16 | p15
Summary Assembly of DNA into chromatin requires a delicate balancing act, as both dearth and excess of histones severely disrupts chromatin function [1–3]. In particular, cells need to carefully control histone stoichiometry: if different types of histones are incorporated into chromatin in an imbalanced manner, it can lead to altered gene expression, mitotic errors, and death [4–6]. Both the balance between individual core histones and between core histones and histone variants is critical [5, 7]. Here, we find that in early Drosophila embryos histone balance in the nuclei is regulated by lipid droplets, cytoplasmic fat storage organelles . Lipid droplets were previously known to function in long-term histone storage: newly laid embryos contain large amounts of excess histones generated during oogenesis , and the maternal supplies of core histone H2A and the histone variant H2Av are anchored to lipid droplets via the novel protein Jabba . We find that in these embryos synthesis of new H2A and H2Av is imbalanced and that newly produced H2Av can be recruited to lipid droplets. When droplet sequestration is disrupted by mutating Jabba, embryos display an elevated H2Av/H2A ratio in nuclei as well as mitotic defects, reduced viability, and hypersensitivity to H2Av overexpression. We propose that in Drosophila embryos lipid droplets serve as a histone buffer, not only storing maternal histones to support the early cell cycles, but also transiently sequestering H2Av produced in excess and thus ensuring proper histone balance in the nucleus.
Most human somatic cells do not divide indefinitely but enter a terminal growth arrest termed replicative senescence. Replicatively senescent cells are generally believed to arrest in G1 or G0 stage of the cell cycle. While doing cell cycle analysis on three different lines of normal human fibroblasts we observed that 36-60% of the replicatively senescent cells had 4N DNA content. Only up to 5% of senescent cells had more than one nucleus ruling out the possibility that the 4N cell population were G1-arrested bi-nucleated cells. Furthermore, it is unlikely that the 4N cells are tetraploids, because actively dividing pre-senescent cultures lacked the 8N tetraploid G2 population. Collectively these results suggest that the 4N population consists of G2 arrested cells. The notion that a large fraction of senescent cell population is arrested in G2 is important for understanding the biology of replicative senescence.
SummaryNaked mole rat (NMR) is a valuable model for aging and cancer research due to its exceptional longevity and cancer resistance. We observed that the reprogramming efficiency of NMR fibroblasts in response to OSKM was drastically lower than that of mouse fibroblasts. Expression of SV40 LargeT antigen (LT) dramatically improved reprogramming of NMR fibroblasts. Inactivation of Rb alone, but not p53, was sufficient to improve reprogramming efficiency, suggesting that NMR chromatin may be refractory to reprogramming. Analysis of the global histone landscape revealed that NMR had higher levels of repressive H3K27 methylation marks and lower levels of activating H3K27 acetylation marks than mouse. ATAC-seq revealed that in NMR, promoters of reprogramming genes were more closed than mouse promoters, while expression of LT led to massive opening of the NMR promoters. These results suggest that NMR displays a more stable epigenome that resists de-differentiation, contributing to the cancer resistance and longevity of this species.
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