ATRX promotes heterochromatin formation to protect cells from G-quadruplex DNA-mediated stress

Vincent U Gant,Sara Martire,Amrita Basu,Aishwarya Sundaresan,Laura A Banaszynski,Ryan O’Hara,Jane N Warshaw,Minhua Li,Yu-Ching Teng

doi:10.1038/s41467-021-24206-5

Abstract

ATRX is a tumor suppressor that has been associated with protection from DNA replication stress, purportedly through resolution of difficult-to-replicate G-quadruplex (G4) DNA structures. While several studies demonstrate that loss of ATRX sensitizes cells to chemical stabilizers of G4 structures, the molecular function of ATRX at G4 regions during replication remains unknown. Here, we demonstrate that ATRX associates with a number of the MCM replication complex subunits and that loss of ATRX leads to G4 structure accumulation at newly synthesized DNA. We show that both the helicase domain of ATRX and its H3.3 chaperone function are required to protect cells from G4-induced replicative stress. Furthermore, these activities are upstream of heterochromatin formation mediated by the histone methyltransferase, ESET, which is the critical molecular event that protects cells from G4-mediated stress. In support, tumors carrying mutations in either ATRX or ESET show increased mutation burden at G4-enriched DNA sequences. Overall, our study provides new insights into mechanisms by which ATRX promotes genome stability with important implications for understanding impacts of its loss on human disease.

Highlights

ATRX is a tumor suppressor that has been associated with protection from DNA replication stress, purportedly through resolution of difficult-to-replicate G-quadruplex (G4) DNA structures
This effect could be rescued by expression of wild-type ATRX in ATRX knockout (ATRX KO) ESCs, whereas we find that both the helicase activity of ATRX and its ability to interact with DAXX are important to reduce G4-EdU persistence (Supplementary Fig. 7)
We found that H3K9me[3] enrichment at ATRX-bound G4 regions was reduced in ATRX KO, DAXX knockout (DAXX KO), and H3.3 KO cells (Fig. 6a–d)

Summary

Introduction

ATRX is a tumor suppressor that has been associated with protection from DNA replication stress, purportedly through resolution of difficult-to-replicate G-quadruplex (G4) DNA structures. We demonstrate that ATRX associates with a number of the MCM replication complex subunits and that loss of ATRX leads to G4 structure accumulation at newly synthesized DNA. We show that both the helicase domain of ATRX and its H3.3 chaperone function are required to protect cells from G4-induced replicative stress. We find that ATRX maintains a closed chromatin state at G4-containing regions and that this function is mediated by DAXX-dependent deposition of H3.3 These activities are upstream of heterochromatin formation, which is required for protection from G4-mediated stress. Our findings support the conclusion that ATRX protects genomic integrity at G4-containing regions by maintaining these regions in a closed heterochromatic state

Methods

Results

Conclusion