Swi1 and Swi3 Are Components of a Replication Fork Protection Complex in Fission Yeast

Abstract

Swi1 is required for programmed pausing of replication forks near the mat1 locus in the fission yeast Schizosaccharomyces pombe. This fork pausing is required to initiate a recombination event that switches mating type. Swi1 is also needed for the replication checkpoint that arrests division in response to fork arrest. How Swi1 accomplishes these tasks is unknown. Here we report that Swi1 copurifies with a 181-amino-acid protein encoded by swi3⁺. The Swi1-Swi3 complex is required for survival of fork arrest and for activation of the replication checkpoint kinase Cds1. Association of Swi1 and Swi3 with chromatin during DNA replication correlated with movement of the replication fork. swi1Δ and swi3Δ mutants accumulated Rad22 (Rad52 homolog) DNA repair foci during replication. These foci correlated with the Rad22-dependent appearance of Holliday junction (HJ)-like structures in cells lacking Mus81-Eme1 HJ resolvase. Rhp51 and Rhp54 homologous recombination proteins were not required for viability in swi1Δ or swi3Δ cells, indicating that the HJ-like structures arise from single-strand DNA gaps or rearranged forks instead of broken forks. We propose that Swi1 and Swi3 define a fork protection complex that coordinates leading- and lagging-strand synthesis and stabilizes stalled replication forks.

We thank V. Martin for providing the Rad11-3FLAG strain. Members of the Scripps Cell Cycle groups are thanked for their support and encouragement. We thank Joel Huberman for his helpful discussion.

E.N. was a fellow of the Human Frontier Science Program. E.N. was also supported by the UEHARA Memorial Foundation. This work was funded by MERK-MGRI-241 (W.H.M. and J.R.Y.), NIH EY1328801 (J.R.Y.), and NIH grant GM59447 awarded to P.R.