BMI1 reduces ATR activation and signalling caused by hydroxyurea
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Xiaozeng Lin1,2,3, Fengxiang Wei1,2,3,4, Peter Whyte5 and Damu Tang1,2,3
1Division of Nephrology, Department of Medicine, McMaster University, Hamilton, Ontario, Canada
2Father Sean O’Sullivan Research Institute, Hamilton, Ontario, Canada
3The Hamilton Center for Kidney Research, St. Joseph’s Hospital, Hamilton, Ontario, Canada
4The Genetics Laboratory, Longgang District Maternity and Child Healthcare Hospital, Longgang District, Shenzhen, Guangdong, P.R. China
5Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada
Damu Tang, email: [email protected]
Keywords: BMI1, DNA damage response, ATR, CHK1, S checkpoint activation
Received: February 14, 2017 Accepted: September 03, 2017 Published: September 20, 2017
BMI1 facilitates DNA damage response (DDR) induced by double strand DNA breaks; however, it remains unknown whether BMI1 functions in single strand DNA (ssDNA) lesions-initiated DDR. We report here that BMI1 reduces hydroxyurea-elicited ATR activation, thereby reducing the S-phase checkpoints. Hydroxyurea induces ssDNA lesions, which activate ATR through binding TOPBP1 as evidenced by phosphorylation of ATR at threonine 1989 (ATRpT1989). ATR subsequently phosphorylates H2AX at serine 139 (γH2AX) and CHK1 at serine 345 (CHK1pS345), leading to phosphorylation of CDK1 at tyrosine 15 (CDK1pY15) and S-phase arrest. BMI1 overexpression reduced γH2AX, CHK1pS345, CDK1pY15, S-phase arrest, and ATR activation in HU-treated MCF7 and DU145 cells, whereas BMI1 knockdown enhanced these events. BMI1 contains a ring finger, helix-turn, proline/serine domain and two nuclear localization signals (NLS). Individual deletion of these domains did not abolish BMI1-derived reductions of CHK1pS345 in MCF7 cells following HU exposure, suggesting that these structural features are not essential for BMI1 to attenuate ATR-mediated CHK1pS345. BMI1 interacts with both TOPBP1 and ATR. Furthermore, all of our BMI1 mutants associate with endogenous TOPBP1. It has previously been established that association of TOPBP1 and ATR is required for ATR activation. Thus, our results suggest that BMI1 decreases ATR activation through a mechanism that involves binding to TOPBP1 and/or ATR.
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