WW Domain‐containing Oxidoreductase Induces Cellular Senescence in Response to Replication Stress

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The induction of DNA damage response (DDR) to ensure accurate duplication of genetic information is crucial for the maintenance of cellular genome integrity during DNA replication. Under replication stress, cellular senescence is a DDR mechanism that prevents the proliferation of cells with DNA damage to avoid mitotic anomalies and inheritance of the damage over cell generations. A reduction in the ability of cells to trigger senescence has been suggested to be associated with cancer initiation and progression. Human WWOX gene resides within a common chromosomal fragile site FRA16D and encodes a tumor suppressor WW domain‐containing oxidoreductase. Downregulation of WWOX protein expression has been frequently found in various human cancers. Increased genome instability has been shown in the cells null for WWOX. However, the role of WWOX in DDR remains largely unclear. In this study, we demonstrated that Wwox gene knockout (Wwox−/−) mouse embryonic fibroblasts (MEFs) failed to undergo replication‐induced cellular senescence after multiple passages in culture. Strikingly, accelerated cell cycle progression and increased apoptosis were detected in late‐passage Wwox−/− MEFs. Increased γH2AX‐positive cells were observed in late‐passage Wwox−/− MEFs, indicating massive accumulation of DNA damages in these cells. The p16INK4A/Rb‐ and p53/p21cip1/waf1‐mediated signaling pathways have been suggested to control cellular senescence. We determined significantly reduced expression of p16INK4A and p21cip1/waf1 proteins in late‐passage Wwox−/− MEFs. Treatment of Wwox−/− MEFs with DNA methylation inhibitor 5‐aza‐2′‐deoxycytidine reversed p16INK4A and p21cip1/waf1 expression, and ectopic expression of p16INK4A and p21cip1/waf1 increased cellular senescence in late‐passage Wwox−/− MEFs. These results suggest that hypermethylation of p16INK4A and p21cip1/waf1 promoters may lead to the failure of senescence induction in Wwox−/− MEFs. Moreover, inhibition of ROS production by N‐acetyl‐L‐cysteine during cell replication in vitro increased protein expression of p16INK4A and p21cip1/waf1 and the induction of cellular senescence in Wwox−/− MEFs. In conclusion, our results support a crucial role for WWOX in DDR for maintaining genome integrity via the induction of cellular senescence in response to replication stress.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.