GSE211192
Asymmetric distribution of H3K9me3 at replicating DNA strands silences L1 retrotransposons during S phase
In higher eukaryotes, repetitive DNA sequences are transcriptionally silenced through histone H3 lysine 9 tri-methylation (H3K9me3). A loss of silencing of these elements leads to genome instability and human diseases, including cancer and aging1-3. While the role of H3K9me3 in the establishment and maintenance of heterochromatin and repression of DNA repeats has been extensively studied4-6, the pattern and mechanism underlying the partitioning of H3K9me3 at replicating DNA strands were unknown. Here, we report that H3K9me3 is preferentially transferred onto the leading strands of replication forks, which occurs predominantly at Long Interspersed Nuclear Element (LINE) retrotransposons that are theoretically transcribed at the head-on direction with replication fork movement. In contrast, all other modified forms of histones tested are distributed in a nearly symmetric manner at the leading and lagging strands. Mechanistically, the Human Silencing Hub (HUSH) complex interacts with the leading strand DNA polymerase Pol e, and contributes to the asymmetric segregation of H3K9me3 during chromatin replication. Cells deficient for Pol e (POLE3 and POLE4), the HUSH complex (MPP8 and TASOR), or cells expressing a MPP8 mutant defective of H3K9me3 binding, or TASOR mutants with reduced interaction with Pol e, show compromised H3K9me3 asymmetry and increased LINE expression. These results reveal an unexpected mechanism whereby the HUSH complex functions with Pol e to promote asymmetric H3K9me3 distribution at LINEs to suppress their expression during S phase. Overall design: The CUT&TAG, CUT&RUN, and eSPAN experiments were conducted in WT, MCM2-2A, POLE3KO, POLE4KO,MPP8KO,TASORKO,POLE3MPP8KO,SETDB1KD, G9aKO, GLPKO, SUV39h1KO, etc mES cells, activated mouse B cells and HeLa cells. The histone markers performed by CUT&TAG and eSPAN methods in mES cells , activated mouse B cells and HeLa cells included H3K9me3, H3K9me2, H3K27me3 ,H4K20me3, H4K20me2 etc. For the regulation of gene expression, we performed GRO-seq experiments in WT, MCM2-2A mutant, POLE3 KO and POLE4 KO mES cells.