A mechanism for Rad53 to couple leading- And lagging-strand DNA synthesis under replication stress in budding yeast

Albert Serra-Cardona, Chuanhe Yu, Xinmin Zhang, Xu Hua, Yuan Yao, Jiaqi Zhou, Haiyun Gan, Zhiguo Zhang

Research output: Contribution to journalArticlepeer-review

Abstract

In response to DNA replication stress, DNA replication checkpoint kinase Mec1 phosphorylates Mrc1, which in turn activates Rad53 to prevent the generation of deleterious single-stranded DNA, a process that remains poorly understood. We previously reported that lagging-strand DNA synthesis proceeds farther than leading strand in rad53-1 mutant cells defective in replication checkpoint under replication stress, resulting in the exposure of long stretches of the leading-strand templates. Here, we show that asymmetric DNA synthesis is also observed in mec1-100 and mrc1-AQ cells defective in replication checkpoint but, surprisingly, not in mrc1Δ cells in which both DNA replication and checkpoint functions of Mrc1 are missing. Furthermore, depletion of either Mrc1 or its partner, Tof1, suppresses the asymmetric DNA synthesis in rad53-1 mutant cells. Thus, the DNA replication checkpoint pathway couples leading- and lagging-strand DNA synthesis by attenuating the replication function of Mrc1-Tof1 under replication stress.

Original languageEnglish (US)
Article numbere2109334118
JournalProceedings of the National Academy of Sciences of the United States of America
Volume118
Issue number38
DOIs
StatePublished - Sep 21 2021

Bibliographical note

Funding Information:
ACKNOWLEDGMENTS. We thank Dr. Bruce Stillman for anti-Mcm2 antibodies and Dr. Steven Brill for the antibody against yeast RPA. This work is supported by the NIH Grant R35 GM115018 (to Z.Z.), the Major Program of National Natural Science Foundation of China (Grant 32090031) and the Guangdong Provincial Key Laboratory of Synthetic Genomics (2019B030301006) (to H.G.), and the NIH Grant K99GM134180 (to A.S.-C.). The NIH/National Cancer Institute Cancer Center Support Grant P30CA013696 to the Herbert Irving Comprehensive Cancer Center supports the sequencing core facilities, which were critical for the completion of this study.

Publisher Copyright:
© 2021 National Academy of Sciences. All rights reserved.

Keywords

  • Asymmetric DNA synthesis
  • Deleterious ssDNA
  • Mrc1
  • Rad53
  • Replication stress

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