Androgen signaling stabilizes genomes to counteract senescence by promoting XRCC4 transcription
Ontology highlight
ABSTRACT: Aging is accompanied by a decreased DNA repair capacity, which might contribute to age-associated functional decline in multiple tissues. Disruption in hormone signaling, associated with reproductive organ dysfunction, is an early event of age-related tissue degeneration, but whether it impacts DNA repair in non-reproductive organs remains elusive. Using skin fibroblasts derived from healthy donors with a broad age range, we show here that the downregulation of expression of XRCC4, a factor involved in nonhomologous end-joining (NHEJ) repair, which is the dominant pathway to repair somatic double-strand breaks, is mediated through transcriptional mechanisms. We show that the androgen receptor (AR), whose expression is also reduced during aging, directly binds to and enhances the activity of the XRCC4 promoter, facilitating XRCC4 transcription and thus stabilizing the genome. We also demonstrate that dihydrotestosterone (DHT), a powerful AR agonist, restores XRCC4 expression and stabilizes the genome in different models of cellular aging. Moreover, DHT treatment reverses senescence-associated phenotypes, opening a potential avenue to aging interventions in the future.
Project description:Cellular senescence is a physiological response by which an organism halts the proliferation of potentially harmful and damaged cells. However, the accumulation of senescent cells over time can become deleterious leading to diseases and physiological decline. Our data reveal a novel interplay between senescence and the stress response that affects both the progression of senescence and the behavior of senescent cells. We show that constitutive exposure to stress induces the formation of stress granules (SGs) in proliferative and presenescent cells, but not in fully senescent cells. Stress granule assembly alone is sufficient to decrease the number of senescent cells without affecting the expression of bona fide senescence markers. SG-mediated inhibition of senescence is associated with the recruitment of the plasminogen activator inhibitor-1 (PAI-1), a known promoter of senescence, to these entities. PAI-1 localization to SGs increases the translocation of cyclin D1 to the nucleus, promotes RB phosphorylation, and maintains a proliferative, non-senescent state. Together, our data indicate that SGs may be targets of intervention to modulate senescence in order to impair or prevent its deleterious effects.
Project description:XRCC4-like factor (XLF) is involved in non-homologous end joining-mediated repair of DNA double-strand breaks (DSBs). Mutations in the WRN gene results in the development of Werner syndrome (WS), a rare autosomal recessive disorder characterized by premature ageing and genome instability. In the present study, it was identified that XLF protein levels were lower in WRN-deficient fibroblasts, compared with normal fibroblasts. Depletion of WRN in HeLa cells led to a decrease of XLF mRNA and its promoter activity. Chromatin immunoprecipitation assays demonstrated that WRN was associated with the XLF promoter. Depletion of XLF in normal human fibroblasts increased the percentage of β-galactosidase (β-gal) staining-positive cells, indicating acceleration in cellular senescence. Taken together, the results suggest that XLF is a transcriptional target of WRN and may be involved in the regulation of cellular senescence.
Project description:SIRT1, the closest mammalian homolog of yeast Sir2, is an NAD(+)-dependent deacetylase with relevant functions in cancer, aging, and metabolism among other processes. SIRT1 has a diffuse nuclear localization but is recruited to the PML nuclear bodies (PML-NBs) after PML upregulation. However, the functions of SIRT1 in the PML-NBs are unknown. In this study we show that primary mouse embryo fibroblasts lacking SIRT1 contain reduced PML protein levels that are increased after reintroduction of SIRT1. In addition, overexpression of SIRT1 in HEK-293 cells increases the amount of PML protein whereas knockdown of SIRT1 reduces the size and number of PML-NBs and the levels of PML protein in HeLa cells. SIRT1 stimulates PML sumoylation in vitro and in vivo in a deacetylase-independent manner. Importantly, the absence of SIRT1 reduces the apoptotic response of vesicular stomatitis virus-infected cells and favors the extent of this PML-sensitive virus replication. These results show a novel function of SIRT1 in the control of PML and PML-NBs.
Project description:Heterochromatin, a key component of the eukaryotic nucleus, is fundamental to the regulation of genome stability, gene expression and cellular functions. However, the factors and mechanisms involved in heterochromatin formation and maintenance still remain largely unknown. Here, we show that insulin receptor tyrosine kinase substrate (IRTKS), an I-BAR domain protein, is indispensable for constitutive heterochromatin formation via liquid‒liquid phase separation (LLPS). In particular, IRTKS droplets can infiltrate heterochromatin condensates composed of HP1 and diverse DNA-bound nucleosomes. IRKTS can stabilize HP1 by recruiting the E2 ligase Ubc9 to SUMOylate HP1 which enables it to form larger phase-separated droplets than unmodified HP1. Furthermore, IRTKS deficiency leads to loss of heterochromatin, resulting in genome-wide changes in chromatin accessibility and aberrant transcription of repetitive DNA elements. This leads to activation of cGAS-STING pathway and type-I interferon (IFN-I) signaling, as well as to the induction of cellular senescence and senescence-associated secretory phenotype (SASP) responses. Collectively, our findings establish a mechanism by which IRTKS condensates consolidate constitutive heterochromatin, revealing an unexpected role of IRTKS as an epigenetic mediator of cellular senescence.
Project description:Continuous translation elongation, irrespective of amino acid sequences, is a prerequisite for living organisms to produce their proteomes. However, nascent polypeptide products bear an inherent risk of elongation abortion. For example, negatively charged sequences with occasional intermittent prolines, termed intrinsic ribosome destabilization (IRD) sequences, weaken the translating ribosomal complex, causing certain nascent chain sequences to prematurely terminate translation. Here, we show that most potential IRD sequences in the middle of open reading frames remain cryptic and do not interrupt translation, due to two features of the nascent polypeptide. Firstly, the nascent polypeptide itself spans the exit tunnel, and secondly, its bulky amino acid residues occupy the tunnel entrance region, thereby serving as a bridge and protecting the large and small ribosomal subunits from dissociation. Thus, nascent polypeptide products have an inbuilt ability to ensure elongation continuity.
Project description:The accepted androgen receptor (AR) role is to promote proliferation and survival of prostate epithelium and thus prostate cancer progression. While growth-inhibitory, tumor-suppressive AR effects have also been documented, the underlying mechanisms are poorly understood. Here, we for the first time link AR anti-cancer action with cell senescence in vitro and in vivo. First, AR-driven senescence was p53-independent. Instead, AR induced p21, which subsequently reduced ΔN isoform of p63. Second, AR activation increased reactive oxygen species (ROS) and thereby suppressed Rb phosphorylation. Both pathways were critical for senescence as was proven by p21 and Rb knock-down and by quenching ROS with N-Acetyl cysteine and p63 silencing also mimicked AR-induced senescence. The two pathways engaged in a cross-talk, likely via PML tumor suppressor, whose localization to senescence-associated chromatin foci was increased by AR activation. All these pathways contributed to growth arrest, which resolved in senescence due to concomitant lack of p53 and high mTOR activity. This is the first demonstration of senescence response caused by a nuclear hormone receptor.
Project description:The bipolar androgen therapy (BAT) includes the treatment of prostate cancer (PCa) patients with supraphysiological androgen level (SAL). Interestingly, SAL induces cell senescence in PCa cell lines as well as ex vivo in tumor samples of patients. The SAL-mediated cell senescence was shown to be androgen receptor (AR)-dependent and mediated in part by non-genomic AKT signaling. RNA-seq analyses compared with and without SAL treatment as well as by AKT inhibition (AKTi) revealed a specific transcriptome landscape. Comparing the top 100 genes similarly regulated by SAL in two human PCa cell lines that undergo cell senescence and being counteracted by AKTi revealed 33 commonly regulated genes. One gene, ERBB receptor feedback inhibitor 1 (ERRFI1), encodes the mitogen-inducible gene 6 (MIG6) that is potently upregulated by SAL, whereas the combinatory treatment of SAL with AKTi reverses the SAL-mediated upregulation. Functionally, knockdown of ERRFI1 enhances the pro-survival AKT pathway by enhancing phosphorylation of AKT and the downstream AKT target S6, whereas the phospho-retinoblastoma (pRb) protein levels were decreased. Further, the expression of the cell cycle inhibitor p15INK4b is enhanced by SAL and ERRFI1 knockdown. In line with this, cell senescence is induced by ERRFI1 knockdown and is enhanced slightly further by SAL. Treatment of SAL in the ERRFI1 knockdown background enhances phosphorylation of both AKT and S6 whereas pRb becomes hypophosphorylated. Interestingly, the ERRFI1 knockdown does not reduce AR protein levels or AR target gene expression, suggesting that MIG6 does not interfere with genomic signaling of AR but represses androgen-induced cell senescence and might therefore counteract SAL-induced signaling. The findings indicate that SAL treatment, used in BAT, upregulates MIG6, which inactivates both pRb and the pro-survival AKT signaling. This indicates a novel negative feedback loop integrating genomic and non-genomic AR signaling.
Project description:To contribute to further understanding the cellular and molecular complexities of inflammatory-immune responses in allergic disorders, we have tested the pro-homeostatic elovanoids (ELV) in human nasal epithelial cells (HNEpC) in culture challenged by several allergens. ELV are novel bioactive lipid mediators synthesized from the omega-3 very-long-chain polyunsaturated fatty acids (VLC-PUFA,n-3). We ask if: (a) several critical signaling events that sustain the integrity of the human nasal epithelium and other organ barriers are perturbed by house dust mites (HDM) and other allergens, and (b) if ELV would participate in beneficially modulating these events. HDM is a prevalent indoor allergen that frequently causes allergic respiratory diseases, including allergic rhinitis and allergic asthma, in HDM-sensitized individuals. Our study used HNEpC as an in vitro model to study the effects of ELV in counteracting HDM sensitization resulting in inflammation, endoplasmic reticulum (ER) stress, autophagy, and senescence. HNEpC were challenged with the following allergy inducers: LPS, poly(I:C), or Dermatophagoides farinae plus Dermatophagoides pteronyssinus extract (HDM) (30 µg/mL), with either phosphate-buffered saline (PBS) (vehicle) or ELVN-34 (500 nM). Results show that ELVN-34 promotes cell viability and reduces cytotoxicity upon HDM sensitization of HNEpC. This lipid mediator remarkably reduces the abundance of pro-inflammatory cytokines and chemokines IL-1β, IL-8, VEGF, IL-6, CXCL1, CCL2, and cell adhesion molecule ICAM1 and restores the levels of the pleiotropic anti-inflammatory IL-10. ELVN-34 also lessens the expression of senescence gene programming as well as of gene transcription engaged in pro-inflammatory responses. Our data also uncovered that HDM triggered the expression of key genes that drive autophagy, unfolded protein response (UPR), and matrix metalloproteinases (MMP). ELVN-34 has been shown to counteract these effects effectively. Together, our data reveal a novel, pro-homeostatic, cell-protective lipid-signaling mechanism in HNEpC as potential therapeutic targets for allergies.
Project description:The HIV-1 accessory protein Vif hijacks a cellular Cullin-RING ubiquitin ligase, CRL5, to promote degradation of the APOBEC3 (A3) family of restriction factors. Recently, the cellular transcription cofactor CBF? was shown to form a complex with CRL5-Vif and to be essential for A3 degradation and viral infectivity. We now demonstrate that CBF? is required for assembling a well-ordered CRL5-Vif complex by inhibiting Vif oligomerization and by activating CRL5-Vif via direct interaction. The CRL5-Vif-CBF? holoenzyme forms a well-defined heterohexamer, indicating that Vif simultaneously hijacks CRL5 and CBF?. Heterodimers of CBF? and RUNX transcription factors contribute toward the regulation of genes, including those with immune system functions. We show that binding of Vif to CBF? is mutually exclusive with RUNX heterodimerization and impacts the expression of genes whose regulatory domains are associated with RUNX1. Our results provide a mechanism by which a pathogen with limited coding capacity uses one factor to hijack multiple host pathways.