Critical Function of γH2A in S-Phase

Area of Science:

• Molecular Biology
• Cellular Biology
• Genetics

Background:

• Phosphorylation of histone H2AX (γH2AX) by ATM and ATR is a key event in DNA damage response.
• While extensively studied for DNA double-strand breaks, γH2AX's role in replication stress is emerging.
• Replication stress also triggers γH2A formation in Schizosaccharomyces pombe.

Purpose of the Study:

• Investigate the role of γH2A in DNA replication stress.
• Identify factors essential for γH2A function when Replication Factor C (RFC) is defective.
• Elucidate the mechanism by which γH2A stabilizes stalled replication forks.

Main Methods:

• Focused genetic screen in fission yeast.
• Immunoblot analysis.
• Localization studies of proteins at replication forks.

Main Results:

• γH2A is critical in RFC-defective cells, independent of canonical checkpoint proteins like Chk1, Cds1/Chk2, and Rad9-Hus1-Rad1.
• Rad9-Hus1-Rad1 is not required for Rad3/ATR-mediated γH2A formation during S-phase.
• Defects in DNA polymerase epsilon also necessitate γH2A.
• γH2A interacts with Brc1, which localizes to stalled forks and prevents aberrant Replication Protein A (RPA) foci formation.
• Brc1-bound chromatin stabilizes replisomes under limited PCNA or DNA polymerase availability.

Conclusions:

• γH2A plays a vital, non-canonical role in maintaining genome stability during replication stress, particularly when RFC function is compromised.
• The γH2A-Brc1 interaction is crucial for stabilizing stalled replication forks and preventing DNA damage.
• This study highlights a novel pathway for replication fork stabilization mediated by γH2A and Brc1.