MCM8IP activates the MCM8-9 helicase to promote DNA synthesis and homologous recombination upon DNA damage

Jen-Wei Huang,Samuel B Hayward,Roopesh Anand,Tarun S Nambiar,Alberto Ciccia,Giuseppe Leuzzi,Gregory J Brunette,Ananya Acharya,Sarah A Joseph,Rajesh K Soni,Raquel Cuella-Martin,Nathan L Clark,Kara A Bernstein,Angelo Taglialatela,Petr Cejka

doi:10.1038/s41467-020-16718-3

Abstract

Homologous recombination (HR) mediates the error-free repair of DNA double-strand breaks to maintain genomic stability. Here we characterize C17orf53/MCM8IP, an OB-fold containing protein that binds ssDNA, as a DNA repair factor involved in HR. MCM8IP-deficient cells exhibit HR defects, especially in long-tract gene conversion, occurring downstream of RAD51 loading, consistent with a role for MCM8IP in HR-dependent DNA synthesis. Moreover, loss of MCM8IP confers cellular sensitivity to crosslinking agents and PARP inhibition. Importantly, we report that MCM8IP directly associates with MCM8-9, a helicase complex mutated in primary ovarian insufficiency, and RPA1. We additionally show that the interactions of MCM8IP with MCM8-9 and RPA facilitate HR and promote replication fork progression and cellular viability in response to treatment with crosslinking agents. Mechanistically, MCM8IP stimulates the helicase activity of MCM8-9. Collectively, our work identifies MCM8IP as a key regulator of MCM8-9-dependent DNA synthesis during DNA recombination and replication.

Highlights

Homologous recombination (HR) mediates the error-free repair of DNA double-strand breaks to maintain genomic stability
We focused our attention on replication protein A (RPA), a central component of the DNA damage response that interacts with a multitude of DNA replication, recombination, and repair proteins[19,21]
Following UV laser microirradiation, BirA*–RPA1 readily accumulated along the damaged tracts marked by γH2AX staining, indicating that fusion with BirA* did not impair the ability of RPA1 to localize to sites of DNA damage (Supplementary Fig. 1a)

Summary

Introduction

Homologous recombination (HR) mediates the error-free repair of DNA double-strand breaks to maintain genomic stability. MCM8-9 has been implicated in HR steps downstream of RAD51 loading[10,17], including DNA damage-induced DNA synthesis at acutely stalled replication forks, presumably the sites of one-ended DSBs, and at I-SceI-generated two-ended DSBs17. These observations raise the possibility that MCM8-9 can facilitate D-loop extension during recombination-associated DNA synthesis[17,18]. MCM8IP-deficient cells exhibit HR defects downstream of RAD51 loading, especially in long-tract gene conversion, and loss of MCM8IP is associated with cellular sensitivity to cisplatin and PARP inhibition. Our findings suggest a role for the MCM8IP–MCM8-9 complex in promoting DNA damageassociated DNA synthesis

Methods

Results

Conclusion