MDC1 PST-repeat region promotes histone H2AX-independent chromatin association and DNA damage tolerance

Israel Salguero,Matylda Sczaniecka-Clift,Mukerrem Demir,Marcus D Wilson,Stephen P Jackson,Julia Coates,Rimma Belotserkovskaya,Satpal Jhujh

doi:10.1038/s41467-019-12929-5

Abstract

Histone H2AX and MDC1 are key DNA repair and DNA-damage signalling proteins. When DNA double-strand breaks (DSBs) occur, H2AX is phosphorylated and then recruits MDC1, which in turn serves as a docking platform to promote the localization of other factors, including 53BP1, to DSB sites. Here, by using CRISPR-Cas9 engineered human cell lines, we identify a hitherto unknown, H2AX-independent, function of MDC1 mediated by its PST-repeat region. We show that the PST-repeat region directly interacts with chromatin via the nucleosome acidic patch and mediates DNA damage-independent association of MDC1 with chromatin. We find that this region is largely functionally dispensable when the canonical γH2AX-MDC1 pathway is operative but becomes critical for 53BP1 recruitment to DNA-damage sites and cell survival following DSB induction when H2AX is not available. Consequently, our results suggest a role for MDC1 in activating the DDR in areas of the genome lacking or depleted of H2AX.

Highlights

Cells are constantly subjected to a plethora of exogeneous and endogenously-derived DNA damaging agents
Our results imply that MDC1 promotes survival to DNA damage in the absence of H2AX via its ability to help recruit repair factors to double-strand breaks (DSBs) regions
We have established that DNA-damage independent MDC1 association with chromatin is largely mediated by its proline-serine-threonine rich (PST)-repeat region, a region that does not have any discernible impact on ionising radiation (IR) survival in a H2AX+/+ background but becomes important for IR survival in H2AX−/− cells

Summary

Introduction

Cells are constantly subjected to a plethora of exogeneous and endogenously-derived DNA damaging agents. To protect the genome from DSBs, cells have evolved various proteins that are recruited to damaged chromatin regions to engage DNA repair processes and to trigger a signalling cascade that, amongst other things, can induce cell death or temporary or permanent delays in cell cycle progression. Once RNF168 is localised to the DSB region, it ubiquitylates histone H2A, leading to recruitment and retention of DNA repair factors such as 53BP114–17 and its downstream effectors including RIF1, PTIP and the recently identified Shieldin complex[18] It seems that the same or similar mechanisms leading to 53BP1 accrual at ionising radiation (IR) induced foci (IRIF) mediate 53BP1 recruitment to assemblies called nuclear bodies (NBs) in G1 phase cells. We show that this PST-repeat region binds to nucleosomes, and thereby mediates constitutive association of MDC1 with chromatin, a function that becomes critical for IR resistance in the absence of H2AX

Methods

Results

Conclusion