Histone H3K79 demethylation by KDM2B facilitates proper DNA replication through PCNA dissociation from chromatin.

Abstract

ObjectivesThe level of histone H3 lysine 79 methylation is regulated by the cell cycle and involved in cell proliferation. KDM2B is an H3K79 demethylase. Proliferating cell nuclear antigen (PCNA) is a component of the DNA replication machinery. This study aimed at elucidating a molecular link between H3K79me recognition of PCNA and cell cycle control.Materials and methodsWe generated KDM2B‐depleted 293T cells and histone H3‐K79R mutant‐expressing 293T cells. Western blots were primarily utilized to examine the H3K79me level and its effect on subsequent PCNA dissociation from chromatin. We applied IP, peptide pull‐down, isothermal titration calorimetry (ITC) and ChIP experiments to show the PCNA binding towards methylated H3K79 and DNA replication origins. Flow cytometry, MTT, iPOND and DNA fibre assays were used to assess the necessity of KDM2B for DNA replication and cell proliferation.ResultsWe revealed that KDM2B‐mediated H3K79 demethylation regulated cell cycle progression. We found that PCNA bound chromatin in an H3K79me‐dependent manner during S phase. KDM2B was responsible for the timely dissociation of PCNA from chromatin, allowing to efficient DNA replication. Depletion of KDM2B aberrantly enriched chromatin with PCNA and caused slow dissociation of residual PCNA, leading to a negative effect on cell proliferation.ConclusionsWe suggested a novel interaction between PCNA and H3K79me. Thus, our findings provide a new mechanism of KDM2B in regulation of DNA replication and cell proliferation.