Abstract
Efficient pre-replication complex (pre-RC) formation on chromatin templates is crucial for the maintenance of genome integrity. However, the regulation of chromatin dynamics during this process has remained elusive. We found that a conserved protein, GRWD1 (glutamate-rich WD40 repeat containing 1), binds to two representative replication origins specifically during G1 phase in a CDC6- and Cdt1-dependent manner, and that depletion of GRWD1 reduces loading of MCM but not CDC6 and Cdt1. Furthermore, chromatin immunoprecipitation coupled with high-throughput sequencing (Seq) revealed significant genome-wide co-localization of GRWD1 with CDC6. We found that GRWD1 has histone-binding activity. To investigate the effect of GRWD1 on chromatin architecture, we used formaldehyde-assisted isolation of regulatory elements (FAIRE)-seq or FAIRE-quantitative PCR analyses, and the results suggest that GRWD1 regulates chromatin openness at specific chromatin locations. Taken together, these findings suggest that GRWD1 may be a novel histone-binding protein that regulates chromatin dynamics and MCM loading at replication origins.
Highlights
DNA replication is of fundamental importance in cells and is strictly regulated to occur precisely once per cell cycle
A similar acidic domain has been found in many proteins involved in histone binding, and the WD40 repeat domain of GRWD1 has homology with the p48 subunit of CAF1, which can interact with histones [17]
Cdt1 and ORC1 protein levels decreased during S phases, whereas MCM7 dissociated from chromatin in late S/G2/M phase, as expected [1]
Summary
DNA replication is of fundamental importance in cells and is strictly regulated to occur precisely once per cell cycle. An essential step in DNA replication is the formation of a pre-replication complex (pre-RC) by the ORC, CDC6, Cdt and MCM helicase complexes at a replication origin during the low Cdk period. Cells with depleted MCM replicate at normal rates, they are hypersensitive to replicative stress and defective in Rad17-dependent ATR-mediated replication checkpoint activation [7,8,9]. Mice with reduced expression of MCM2 develop normally but their life spans are greatly reduced [11]. These findings suggest that efficient MCM loading is critical for tolerance of replication stress and activation of the checkpoint
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
