RNF20-mediated H2B monoubiquitination protects stalled forks and promotes fork restart

Chromatin modifications play an important role in transcription, DNA replication and repair. Nonetheless, whether histone modifications regulate replication stress responses remains obscure. Here, we show that RNF20 localizes to and promotes H2B monoubiquitination (H2Bub) at replicating sites. Knockdown of RNF20 leads to degradation of stalled forks by MRE11 nuclease, which can be rescued by inhibition of MRE11 and co-depletion of SMARCAL1/HLTF/ZRANB3 fork remodelers. RNF20 facilitates the loading of RAD51 and RAD51C at the stalled fork sites and participates in the same pathway of RAD51/RAD51C-mediated fork protection and restart. Analyses with the RING domain and phosphorylation-deficient mutants of RNF20 showed that its catalytic activity and ATR/ATM-mediated phosphorylation are essential for its role in replication stress responses. Notably, treatment of RNF20-depleted cells with chromatin relaxing agents rescue the fork protection and restart defects. Collectively, our studies uncover the role of RNF20-mediated H2Bub in regulating the chromatin dynamics to safeguard the replicating genomes.