Checkpoint Kinase Rad53 Couples Leading- and Lagging-Strand DNA Synthesis under Replication Stress

Haiyun Gan, Chuanhe Yu, Sujan Devbhandari, Sushma Sharma, Junhong Han, Andrei Chabes, Dirk Remus, Zhiguo Zhang

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

The checkpoint kinase Rad53 is activated during replication stress to prevent fork collapse, an essential but poorly understood process. Here we show that Rad53 couples leading- and lagging-strand synthesis under replication stress. In rad53-1 cells stressed by dNTP depletion, the replicative DNA helicase, MCM, and the leading-strand DNA polymerase, Pol ε, move beyond the site of DNA synthesis, likely unwinding template DNA. Remarkably, DNA synthesis progresses further along the lagging strand than the leading strand, resulting in the exposure of long stretches of single-stranded leading-strand template. The asymmetric DNA synthesis in rad53-1 cells is suppressed by elevated levels of dNTPs in vivo, and the activity of Pol ε is compromised more than lagging-strand polymerase Pol δ at low dNTP concentrations in vitro. Therefore, we propose that Rad53 prevents the generation of excessive ssDNA under replication stress by coordinating DNA unwinding with synthesis of both strands. Gan et al. find that Rad53 prevents fork collapse via coordinating the unwinding of double-stranded DNA by the replicative helicase with synthesis of both leading and lagging strand. They observed that long stretches of single-stranded leading-strand template coated with RPA are produced in rad53-1 mutant cells under replication stress.

Original languageEnglish (US)
Pages (from-to)446-455.e3
JournalMolecular Cell
Volume68
Issue number2
DOIs
StatePublished - Oct 19 2017
Externally publishedYes

Keywords

  • ChIP-ssSeq
  • DNA replication checkpoint
  • Rad53
  • dNTP pools
  • eSPAN
  • fork collapse
  • lagging strand DNA synthesis
  • leading strand DNA synthesis

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    Gan, H., Yu, C., Devbhandari, S., Sharma, S., Han, J., Chabes, A., Remus, D., & Zhang, Z. (2017). Checkpoint Kinase Rad53 Couples Leading- and Lagging-Strand DNA Synthesis under Replication Stress. Molecular Cell, 68(2), 446-455.e3. https://doi.org/10.1016/j.molcel.2017.09.018