Abstract
The BRE gene, alias BRCC45, produces a 44 kDa protein that is normally distributed in both cytoplasm and nucleus. In this study, we used adult fibroblasts isolated from wild-type (WT) and BRE knockout (BRE−/−) mice to investigate the functional role of BRE in DNA repair and cellular senescence. We compared WT with BRE−/− fibroblasts at different cell passages and observed that the mutant fibroblasts entered replicative senescence earlier than the WT fibroblasts. With the use of gamma irradiation to induce DNA damage in fibroblasts, the percentage of SA-β-Gal+ cells was significantly higher in BRE−/− fibroblasts compared with WT cells, suggesting that BRE is also associated with DNA damage-induced premature senescence. We also demonstrated that the gamma irradiation induced γ-H2AX foci, a DNA damage marker, persisted significantly longer in BRE−/− fibroblasts than in WT fibroblasts, confirming that the DNA repair process is impaired in the absence of BRE. In addition, the BRCA1-A complex recruitment and homologous recombination (HR)-dependent DNA repair process upon DNA damage were impaired in BRE−/− fibroblasts. Taken together, our results demonstrate a role for BRE in both replicative senescence and DNA damage-induced premature senescence. This can be attributed to BRE being required for BRCA1-A complex-driven HR DNA repair.
Highlights
The result of real-time (RT) qPCR, which amplified a transcript region spanning the boundary of exons 7 and 8, confirmed that productive BRE transcript expression was undetectable in BRE−/− fibroblasts (Fig. 1b)
It has been reported that BRE is required for the integrity of BRCA1-A complex in the nucleus and BRISC (BRCC36 isopeptidase complex) in the cytoplasm[14,22]
We found that the adult fibroblasts derived from BRE−/− mice undergo cellular senescence earlier than the WT fibroblasts
Summary
Our results demonstrate a role for BRE in both replicative senescence and DNA damage-induced premature senescence This can be attributed to BRE being required for BRCA1-A complex-driven HR DNA repair. Cellular senescence is an irreversible physiological process that accompanies abnormal and reduced metabolic activities It is characterized by the loss of proliferation ability of somatic cells such as fibroblasts with some ageing-associated phenotypes[1]. The γ -H2AX (the phosphorylated form of H2AX) is localized around the DSBs, where it is K63-polyubiquitinated by RNF8 and Ubc[13] This event leads to the recruitment of a DNA-repair complex, BRCA1-A, to these sites through the interaction between the K63 polyubiquitin chains and RAP80, which is the ubiquitin-binding subunit of the www.nature.com/scientificreports/. It can bind to the cytoplasmic region of Fas[18], TNF-R119, and death-inducing signaling complex (DISC)[20] to protect cells from apoptosis
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