Brc1 Promotes the Focal Accumulation and SUMO Ligase Activity of Smc5-Smc6 during Replication Stress

ABSTRACT

As genetic instability drives disease or loss of cell fitness, cellular safeguards have evolved to protect the genome, especially during sensitive cell cycle phases, such as DNA replication. Fission yeast Brc1 has emerged as a key factor in promoting cell survival when replication forks are stalled or collapsed. Brc1 is a multi-BRCT protein that is structurally related to the budding yeast Rtt107 and human PTIP DNA damage response factors, but functional similarities appear limited. Brc1 is a dosage suppressor of a mutation in the essential Smc5-Smc6 genome stability complex and is thought to act in a bypass pathway. In this study, we reveal an unexpectedly intimate connection between Brc1 and Smc5-Smc6 function. Brc1 is required for the accumulation of the Smc5-Smc6 genome stability complex in foci during replication stress and for activation of the intrinsic SUMO ligase activity of the complex by collapsed replication forks. Moreover, we show that the chromatin association and SUMO ligase activity of Smc5-Smc6 require the Nse5-Nse6 heterodimer, explaining how this nonessential cofactor critically supports the DNA repair roles of Smc5-Smc6. We also found that Brc1 interacts with Nse5-Nse6, as well as gamma-H2A, so it can tether Smc5-Smc6 at replicative DNA lesions to promote survival.

KEYWORDS:

• Brc1
• Smc5-Smc6
• SUMO
• Rad18
• replication stress

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ACKNOWLEDGMENTS

We thank Jan Palecek, Matthew O’Connell, and Felicity Watts for strains. We also thank members of the Cell Cycle Groups at TSRI for their support.

This study was funded by NIH grants GM068608 and GM081840 awarded to M.N.B. Work in the laboratory of P.R. was supported by grants CA077325, CA117638, and GM059447.

M.O., M.C.G., M.N., M.C.R., O.L., P.R., and M.N.B. were all involved in study design, experiment execution, and writing of the manuscript.

None of us has a conflict of interest with regard to the paper.