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Sae2 antagonizes Rad9 accumulation at DNA double-strand breaks to attenuate checkpoint signaling and facilitate end

Tai-Yuan Yu1, Michael T Kimble1,2, Lorraine S Symington3,2

  • 1Department of Microbiology & Immunology, Columbia University Irving Medical Center, New York, NY 10032.

Proceedings of the National Academy of Sciences of the United States of America
|December 5, 2018
PubMed

Insights

The Sae2 protein has a DNA damage checkpoint role independent of its nuclease function. Sae2 modulates the DNA damage checkpoint, affecting genotoxin sensitivity and synthetic lethality with sgs1Δ mutants.

Area of Science:

  • Molecular Biology
  • DNA Repair
  • Cellular Biology

Background:

  • The Mre11-Rad50-Xrs2 (NBS1) complex is crucial for DNA double-strand break (DSB) repair.
  • Mre11 endonuclease, stimulated by Sae2 (CtIP), initiates 5'-3' resection at DSBs.
  • Mre11-nuclease deficient (mre11-nd) and sae2Δ mutants were expected to have similar DNA damage response phenotypes.

Purpose of the Study:

  • To investigate the distinct roles of Sae2 and Mre11 nuclease in DNA double-strand break repair.
  • To elucidate the resection-independent functions of Sae2 in the DNA damage response.
  • To understand how Sae2 modulates the DNA damage checkpoint pathway.

Main Methods:

  • Comparative analysis of mre11-nd and sae2Δ mutant phenotypes.
  • Assessment of genotoxin sensitivity in various mutant strains.
  • Investigation of synthetic lethality interactions, specifically with sgs1Δ.
  • Analysis of checkpoint protein (Tel1, Rad53, Rad9) localization and activity.
  • Biochemical assays to determine Sae2's interaction with Tel1 substrates.

Main Results:

  • sae2Δ cells showed greater genotoxin sensitivity than mre11-nd cells.
  • sae2Δ was synthetic lethal with sgs1Δ, while mre11-nd sgs1Δ was viable.
  • Sae2 attenuated the Tel1-Rad53 checkpoint and antagonized Rad9 accumulation independently of Mre11 nuclease.
  • Sae2 competes with other Tel1 substrates, reducing Rad9 binding to chromatin and Rad53.
  • Persistent Sae2 at DSBs in mre11-nd mutants counteracts checkpoint inhibition of resection.

Conclusions:

  • Sae2 plays a critical role in DNA double-strand break repair beyond its nuclease function.
  • Sae2's ability to modulate the DNA damage checkpoint is essential for cellular survival and proper DSB repair.
  • These findings reveal a complex interplay between Sae2, Mre11, checkpoint kinases, and nucleases in maintaining genome stability.
Keywords:
DNA damage checkpointDNA repairMre11Rad9Sae2

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