Role of Doa1 in the Saccharomyces cerevisiae DNA Damage Response

Abstract

The cellular response to DNA damage requires not only direct repair of the damage but also changes in the DNA replication machinery, chromatin, and transcription that facilitate survival. Here, we describe Saccharomyces cerevisiae Doa1, which helps to control the damage response by channeling ubiquitin from the proteosomal degradation pathway into pathways that mediate altered DNA replication and chromatin modification. DOA1 interacts with genes involved in PCNA ubiquitination, including RAD6, RAD18, RAD5, UBC13, and MMS2, as well as genes involved in histone H2B ubiquitination or deubiquitination, including RAD6, BRE1, LGE1, CDC73, UBP8, UBP10, and HTB2. In the absence of DOA1, damage-induced ubiquitination of PCNA does not occur. In addition, the level of ubiquitinated H2B is decreased under normal conditions and completely absent in the presence of DNA damage. In the case of PCNA, the defect associated with the doa1Δ mutant is alleviated by overexpression of ubiquitin, but in the case of H2B, it is not. The data suggest that Doa1 is the major source of ubiquitin for the DNA damage response and that Doa1 also plays an additional essential and more specific role in the monoubiquitination of histone H2B.

Supplemental material for this article may be found at http://mcb.asm.org/.

We gratefully acknowledge M. A. Osley for helpful discussions. Helpful suggestions of a referee are also acknowledged. We thank M. Hochstrasser, M. Ellison, E. Johnson, M. A. Osley, R. Rothstein, D. Finley, and S. Jentsch for providing strains and plasmids.

This work was funded by the NIH (GM068569).