Histone H2A phosphorylation controls Crb2 recruitment at DNA breaks, maintains checkpoint arrest, and influences DNA repair in fission yeast

Mol Cell Biol. 2004 Jul;24(14):6215-30. doi: 10.1128/MCB.24.14.6215-6230.2004.

Abstract

Mammalian ATR and ATM checkpoint kinases modulate chromatin structures near DNA breaks by phosphorylating a serine residue in the carboxy-terminal tail SQE motif of histone H2AX. Histone H2A is similarly regulated in Saccharomyces cerevisiae. The phosphorylated forms of H2AX and H2A, known as gamma-H2AX and gamma-H2A, are thought to be important for DNA repair, although their evolutionarily conserved roles are unknown. Here, we investigate gamma-H2A in the fission yeast Schizosaccharomyces pombe. We show that formation of gamma-H2A redundantly requires the ATR/ATM-related kinases Rad3 and Tel1. Mutation of the SQE motif to AQE (H2A-AQE) in the two histone H2A genes caused sensitivity to a wide range of genotoxic agents, increased spontaneous DNA damage, and impaired checkpoint maintenance. The H2A-AQE mutations displayed a striking synergistic interaction with rad22Delta (Rad52 homolog) in ionizing radiation (IR) survival. These phenotypes correlated with defective phosphorylation of the checkpoint proteins Crb2 and Chk1 and a failure to recruit large amounts of Crb2 to damaged DNA. Surprisingly, the H2A-AQE mutations substantially suppressed the IR hypersensitivity of crb2Delta cells by a mechanism that required the RecQ-like DNA helicase Rqh1. We propose that gamma-H2A modulates checkpoint and DNA repair through large-scale recruitment of Crb2 to damaged DNA. This function correlates with evidence that gamma-H2AX regulates recruitment of several BRCA1 carboxyl terminus domain-containing proteins (NBS1, 53BP1, MDC1/NFBD1, and BRCA1) in mammals.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Cycle / physiology*
  • Cell Cycle Proteins / metabolism*
  • Checkpoint Kinase 2
  • DNA Damage*
  • DNA Helicases / metabolism
  • DNA Repair*
  • Epistasis, Genetic
  • Fungal Proteins / metabolism
  • Genomic Instability
  • Histones / genetics
  • Histones / metabolism*
  • Humans
  • Intracellular Signaling Peptides and Proteins
  • Molecular Sequence Data
  • Mutation
  • Nuclear Proteins / metabolism*
  • Phosphorylation
  • Protein Kinases / metabolism
  • Protein Serine-Threonine Kinases
  • Saccharomyces cerevisiae Proteins
  • Schizosaccharomyces / cytology
  • Schizosaccharomyces / genetics*
  • Schizosaccharomyces / metabolism
  • Schizosaccharomyces / radiation effects
  • Schizosaccharomyces pombe Proteins / metabolism*
  • Sequence Alignment

Substances

  • Cell Cycle Proteins
  • Crb2 protein, S pombe
  • Fungal Proteins
  • Histones
  • Intracellular Signaling Peptides and Proteins
  • Nuclear Proteins
  • Saccharomyces cerevisiae Proteins
  • Schizosaccharomyces pombe Proteins
  • Protein Kinases
  • Checkpoint Kinase 2
  • Protein Serine-Threonine Kinases
  • TEL1 protein, S cerevisiae
  • rad3 protein, S pombe
  • tel1 protein, S pombe
  • DNA Helicases
  • Rqh1 protein, S pombe