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.

Transient induction of p53 can cause reversible quiescence and irreversible senescence. Using nutlin-3a (a small molecule that activates p53 without causing DNA damage), we have previously identified cell lines in which nutlin-3a caused quiescence. Importantly, nutlin-3a caused quiescence by actively suppressing the senescence program (while still causing cell cycle arrest). Noteworthy, in these cells nutlin-3a inhibited the mTOR (mammalian Target of Rapamycin) pathway, which is known to be involved in the senescence program. Here we showed that shRNA-mediated knockdown of TSC2, a negative regulator of mTOR, partially converted quiescence into senescence in these nutlin-arrested cells. In accord, in melanoma cell lines and mouse embryo fibroblasts, which easily undergo senescence in response to p53 activation, nutlin-3a failed to inhibit mTOR. In these senescence-prone cells, the mTOR inhibitor rapamycin converted nutlin-3a-induced senescence into quiescence. We conclude that status of the mTOR pathway can determine, at least in part, the choice between senescence and quiescence in p53-arrested cells.

SignificanceThe accumulation of senescent cells over a lifetime causes age-related pathologies; however, the inability to reliably identify senescent cells in vivo has hindered clinical efforts to employ this knowledge as a means to ameliorate or reverse aging. Here, we describe a reporter allele, p16tdTom, enabling the in vivo identification and isolation of cells featuring high-level activation of the p16INK4a promoter. Our findings provide an insight into the functional and molecular characteristics of p16INK4a-activated cells in vitro and in vivo. We show that such cells accumulate with aging or other models of injury, and that they exhibit clinically targetable features of cellular senescence.

p53 tumor suppressor gene controls cell response to a variety of stresses inducing growth arrest or apoptosis in damaged cells. It largely determines the sensitivity of tumor and normal cells to radiation and chemotherapy, and, therefore, de®nes both the ecacy and limitations of anti-cancer treatment. To determine molecular mechanisms of p53-dependent stress response in normal tissues we identi®ed and compared the spectra of radiation-responsive genes in cells of dierent origin and p53 status using a cDNA array hybridization technique. The majority of genes identi®ed were p53-dependent and cell type speci®c. Several of the new p53 responders encode known secreted growth inhibitory factors. This suggests that p53, in addition to its intrinsic antiproliferation activity, can cause`bystander eect' by inducing export of growth suppressive stimuli from damaged cells to neighboring cells. Consistently, a p53-dependent accumulation of factors, which causes growth inhibitory eects in a variety of cell lines, was found after gamma irradiation in the media from established and primary cell cultures and in the urine of irradiated mice. Moreover, p53-dependent factors released by normal human ®broblasts potentiated the cytotoxic eect of a chemotherapeutic drug on co-cultivated tumor cells. This suggests a previously unknown role for normal cells in chemo-and radiation therapy of cancer.

G 13 protein, one of the heterotrimeric guanine nucleotide-binding proteins (G proteins), regulates diverse and complex cellular responses by transducing signals from the cell surface presumably involving more than one pathway. Yeast two-hybrid screening of a mouse brain cDNA library identified radixin, a member of the ERM family of three closely related proteins (ezrin, radixin, and moesin), as a protein that interacted with G␣ 13 . Interaction between radixin and G␣ 13 was confirmed by in vitro binding assay and by co-immunoprecipitation technique. Activated G␣ 13 induced conformational activation of radixin, as determined by binding of radixin to polymerized F-actin and by immunofluorescence in intact cells. Finally, two dominant negative mutants of radixin inhibited G␣ 13 -induced focus formation of Rat-1 fibroblasts but did not affect Ras-induced focus formation. Our results identifying a new signaling pathway for G␣ 13 indicate that ERM proteins can be activated by and serve as effectors of heterotrimeric G proteins.