title: The analysis of S. cerevisiae cells deleted for mitotic cyclin Clb2 reveals a novel requirement of Sgs1 DNA helicase and Exonuclease 1 when replication forks break in the presence of alkylation damage.
abstract: In this study, we report the effects of deleting the principal mitotic cyclin, Clb2, in different repair deficient contexts on sensitivity to the alkylating DNA damaging agent, methyl methanesulphonate (MMS). A yeast clb2 mutant is sensitive to MMS and displays synergistic effect when combined with inactivation of numerous genes involved in DNA recombination and replication. In contrast, clb2 has basically no additional effect with deletion of the RecQ helicase SGS1, the exonuclease EXO1 and the protein kinase RAD53 suggesting that Clb2 functions in these pathways. In addition, clb2 increases the viability of the mec1 kinase deficient mutant, suggesting Mec1 inhibits a deleterious Clb2 activity. Interestingly, we found that the rescue by EXO1 deletion of rad53K227 mutant, deficient in checkpoint activation, requires Sgs1, suggesting a role for Rad53, independent of its checkpoint function, in regulating an ordered recruitment of Sgs1 and Exo1 to fork structure. Overall, our data suggest that Clb2 affects recombinant structure of replication fork blocked by alkylating DNA damage at numerous steps and could regulate Sgs1 and Exo1 activity. In addition, we found novel requirement of Sgs1 DNA helicase and Exonuclease 1 when replication forks breaks in the presence of alkylation damage. Models for the functional interactions of mitotic cyclin Clb2, Sgs1 and Exo1 with replication fork stabilization are proposed.
authors.affiliation: Laboratoire d'Ingenierie des Systèmes Macromoléculaires CNRS UPR9027, Aix-Marseille University, 13402 Marseille Cedex 20, France; Université Paris-Sud, CNRS UMR8621, Institut de Génétique et Microbiologie, Bâtiment 400, 91405 Orsay Cedex, France. Electronic address: laurence.signon@igmors.u-psud.fr.
authors.forename: Laurence
authors.initials: L
authors.lastname: Signon
authors.affiliation: Laboratoire d'Ingenierie des Systèmes Macromoléculaires CNRS UPR9027, Aix-Marseille University, 13402 Marseille Cedex 20, France.
authors.forename: Marie Noelle
authors.initials: MN
authors.lastname: Simon
mesh_terms: D000477:Alkylating Agents / Q000494:pharmacology*; D000478:Alkylation; D019926:Cyclin B / Q000235:genetics*; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects*; D004249:DNA Damage; D004261:DNA Replication / Q000187:drug effects / Q000235:genetics; D004843:Epistasis, Genetic / Q000187:drug effects; D005090:Exodeoxyribonucleases / Q000502:physiology*; D017353:Gene Deletion; D008741:Methyl Methanesulfonate / Q000494:pharmacology*; D008938:Mitosis / Q000187:drug effects / Q000235:genetics; D030781:Organisms, Genetically Modified; D053484:RecQ Helicases / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000502:physiology*
chemical_list: D000477:Alkylating Agents; C093749:CLB2 protein, S cerevisiae; D019926:Cyclin B; D029701:Saccharomyces cerevisiae Proteins; D008741:Methyl Methanesulfonate; D005090:Exodeoxyribonucleases; C028450:exodeoxyribonuclease I; C090472:SGS1 protein, S cerevisiae; D053484:RecQ Helicases
keywords: Checkpoint; DNA damage; Mitotic cyclin; Recombination; Replication