Fork pausing complex engages topoisomerases at the replisome

Genes Dev. 2020 Jan 1;34(1-2):87-98. doi: 10.1101/gad.331868.119. Epub 2019 Dec 5.

Abstract

Replication forks temporarily or terminally pause at hundreds of hard-to-replicate regions around the genome. A conserved pair of budding yeast replisome components Tof1-Csm3 (fission yeast Swi1-Swi3 and human TIMELESS-TIPIN) act as a "molecular brake" and promote fork slowdown at proteinaceous replication fork barriers (RFBs), while the accessory helicase Rrm3 assists the replisome in removing protein obstacles. Here we show that the Tof1-Csm3 complex promotes fork pausing independently of Rrm3 helicase by recruiting topoisomerase I (Top1) to the replisome. Topoisomerase II (Top2) partially compensates for the pausing decrease in cells when Top1 is lost from the replisome. The C terminus of Tof1 is specifically required for Top1 recruitment to the replisome and fork pausing but not for DNA replication checkpoint (DRC) activation. We propose that forks pause at proteinaceous RFBs through a "sTOP" mechanism ("slowing down with topoisomerases I-II"), which we show also contributes to protecting cells from topoisomerase-blocking agents.

Keywords: Csm3; Mrc1; RFB; Tof1; Top1; Top2; replication fork pausing; replisome; topoisomerase.

MeSH terms

  • Cell Cycle Proteins / metabolism
  • DNA Helicases / metabolism
  • DNA Replication / genetics*
  • DNA Topoisomerases / metabolism*
  • DNA Topoisomerases, Type I / metabolism
  • DNA-Binding Proteins / metabolism
  • Mutation
  • Protein Transport
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism

Substances

  • Cell Cycle Proteins
  • Csm3p protein, S cerevisiae
  • DNA-Binding Proteins
  • Saccharomyces cerevisiae Proteins
  • TOF1 protein, S cerevisiae
  • Rrm3 protein, S cerevisiae
  • DNA Helicases
  • DNA Topoisomerases
  • TOP1 protein, S cerevisiae
  • DNA Topoisomerases, Type I