MCB Accepts, published online ahead of print on 26 October 2009

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Google Scholar Articles by Kile, A. C. Articles by Koepp, D. M. PubMed PubMed Citation Articles by Kile, A. C. Articles by Koepp, D. M.

 Previous Article  |  Next Article 

Mol. Cell. Biol. doi:10.1128/MCB.00612-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Activation of the S-phase checkpoint inhibits degradation of the F-box protein Dia2

Andrew C. Kile and Deanna M. Koepp^*

Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455

^* To whom correspondence should be addressed. Email: koepp015{at}umn.edu.

A stable genome is critical to cell viability and proliferation. During DNA replication, the S-phase checkpoint pathway responds to replication stress. In budding yeast, the chromatin-bound F-box protein Dia2 is required to maintain genomic stability and may help replication complexes overcome sites of damaged DNA and natural fragile regions. SCF (Skp1/Cul1/F-box protein) complexes are modular ubiquitin ligases. We show here that Dia2 is itself targeted for ubiquitin-mediated proteolysis and activation of the S-phase checkpoint pathway inhibits Dia2 protein degradation. S-phase checkpoint mutants fail to stabilize Dia2 in response to replication stress. Deletion of DIA2 from these checkpoint mutants exacerbates their sensitivity to hydroxyurea and MMS, suggesting that stabilization of Dia2 contributes to the replication stress response. Unlike other F-box proteins, deletion of the F-box domain in Dia2 does not stabilize the protein. Rather, an N-terminal domain that is also required for nuclear localization is necessary for degradation. When a strong NLS is added to dia2 mutants lacking this domain, the Dia2 protein is both stable and nuclear. Together, our results suggest that Dia2 protein turnover does not involve an autocatalytic mechanism and that Dia2 proteolysis is inhibited by activation of the replication stress response.