Cell-Cycle-Regulated Interaction between Mcm10 and Double Hexameric Mcm2-7 Is Required for Helicase Splitting and Activation during S Phase
Yun Quan,
Yisui Xia,
Lu Liu,
Jiamin Cui,
Zhen Li,
Qinhong Cao,
Xiaojiang S. Chen,
Judith L. Campbell,
Huiqiang Lou
Affiliations
Yun Quan
State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, 2 Yuan-Ming-Yuan West Road, Beijing 100193, China
Yisui Xia
State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, 2 Yuan-Ming-Yuan West Road, Beijing 100193, China
Lu Liu
State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, 2 Yuan-Ming-Yuan West Road, Beijing 100193, China
Jiamin Cui
State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, 2 Yuan-Ming-Yuan West Road, Beijing 100193, China
Zhen Li
State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, 2 Yuan-Ming-Yuan West Road, Beijing 100193, China
Qinhong Cao
State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, 2 Yuan-Ming-Yuan West Road, Beijing 100193, China
Xiaojiang S. Chen
Molecular and Computational Biology, USC Norris Cancer Center, and Chemistry Department, University of Southern California, Los Angeles, CA 90089, USA
Judith L. Campbell
Braun Laboratories, California Institute of Technology, Pasadena, CA 91125, USA
Huiqiang Lou
State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, 2 Yuan-Ming-Yuan West Road, Beijing 100193, China
Mcm2-7 helicase is loaded onto double-stranded origin DNA as an inactive double hexamer (DH) in G1 phase. The mechanisms of Mcm2-7 remodeling that trigger helicase activation in S phase remain unknown. Here, we develop an approach to detect and purify the endogenous DHs directly. Through cellular fractionation, we provide in vivo evidence that DHs are assembled on chromatin in G1 phase and separated during S phase. Interestingly, Mcm10, a robust MCM interactor, co-purifies exclusively with the DHs in the context of chromatin. Deletion of the main interaction domain, Mcm10 C terminus, causes growth and S phase defects, which can be suppressed through Mcm10-MCM fusions. By monitoring the dynamics of MCM DHs, we show a significant delay in DH dissolution during S phase in the Mcm10-MCM interaction-deficient mutants. Therefore, we propose an essential role for Mcm10 in Mcm2-7 remodeling through formation of a cell-cycle-regulated supercomplex with DHs.
