MCM8/9 and FANCD2 interact within a shared pathway in response to replication stress caused by DNA crosslinks

Abstract

The Fanconi anemia (FA) protein FANCD2, and MCM8/9 heterohexameric helicase complex are critical for maintaining genomic integrity in response to replication stress. However, the nature of their relationship remains unclear. Here, we show that MCM8/9 physically interacts and functionally cooperates with FANCD2 during the repair of DNA interstrand crosslinks (ICLs). Using immunofluorescence and co-immunoprecipitation studies, we show that MCM8/9 interacts with FANCD2 through its core domain, independently of DNA. FANCD2 is essential for the recruitment of MCM9 to ICL damage induced nuclear foci and acts downstream the FANCD2I monoubiquitination. Although MCM8/9 foci formation requires its intact ATPase activity, BRCv motif and HROB, these are not required for FANCD2 binding, highlighting a distinction between physical interaction and functional activation. Interestingly, FANCD2 foci formation increase in MCM8 or MCM9 knockout cells, suggesting that MCM8/9 functions to mitigate replication associate stress. {gamma}H2AX DNA damage assays and cell survival assays show that combined loss of MCM9 and FANCD2 do not cause any additive DNA damage beyond individual knockouts, indicating an epistatic relationship and suggests they function in the same DNA repair pathway. Together, our findings identify MCM8/9 as a downstream effector of the FA pathway critical for resolving ICL induced DNA damage. HighlightsO_LIMCM8/9 interacts and colocalize with FANCD2 upon ICL DNA damage. C_LIO_LIFANCD2 is essential for recruitment of MCM9 to DNA damage site. C_LIO_LIMCM8/9 is epistatic to FANCD2 and within the same DNA damage response pathway. C_LI