title: A structure-function analysis of the yeast Elg1 protein reveals the importance of PCNA unloading in genome stability maintenance.
abstract: The sliding clamp, PCNA, plays a central role in DNA replication and repair. In the moving replication fork, PCNA is present at the leading strand and at each of the Okazaki fragments that are formed on the lagging strand. PCNA enhances the processivity of the replicative polymerases and provides a landing platform for other proteins and enzymes. The loading of the clamp onto DNA is performed by the Replication Factor C (RFC) complex, whereas its unloading can be carried out by an RFC-like complex containing Elg1. Mutations in ELG1 lead to DNA damage sensitivity and genome instability. To characterize the role of Elg1 in maintaining genomic integrity, we used homology modeling to generate a number of site-specific mutations in ELG1 that exhibit different PCNA unloading capabilities. We show that the sensitivity to DNA damaging agents and hyper-recombination of these alleles correlate with their ability to unload PCNA from the chromatin. Our results indicate that retention of modified and unmodified PCNA on the chromatin causes genomic instability. We also show, using purified proteins, that the Elg1 complex inhibits DNA synthesis by unloading SUMOylated PCNA from the DNA. Additionally, we find that mutations in ELG1 suppress the sensitivity of rad5Δ mutants to DNA damage by allowing trans-lesion synthesis to take place. Taken together, the data indicate that the Elg1-RLC complex plays an important role in the maintenance of genomic stability by unloading PCNA from the chromatin.
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Ramat Aviv 69978, Israel.
authors.forename: Keren
authors.initials: K
authors.lastname: Shemesh
authors.affiliation: Department of Biology, Masaryk University, CZ-625 00 Brno, Czech Republic.
authors.forename: Marek
authors.initials: M
authors.lastname: Sebesta
authors.affiliation: Department of Biology, Masaryk University, CZ-625 00 Brno, Czech Republic.
authors.forename: Martin
authors.initials: M
authors.lastname: Pacesa
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Ramat Aviv 69978, Israel.
authors.forename: Soumitra
authors.initials: S
authors.lastname: Sau
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Ramat Aviv 69978, Israel.
authors.forename: Alex
authors.initials: A
authors.lastname: Bronstein
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Ramat Aviv 69978, Israel.
authors.forename: Oren
authors.initials: O
authors.lastname: Parnas
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Ramat Aviv 69978, Israel.
authors.forename: Batia
authors.initials: B
authors.lastname: Liefshitz
authors.affiliation: Institute of Biotechnology, Vilnius University, Graiciuno 8, Vilnius LT-02241, Lithuania.
authors.forename: Ceslovas
authors.initials: C
authors.lastname: Venclovas
authors.affiliation: Department of Biology, Masaryk University, CZ-625 00 Brno, Czech Republic.|National Center for Biomolecular Research, Masaryk University, CZ-625 00 Brno, Czech Republic.|International Clinical Research Center, St. Anne's University Hospital in Brno, CZ- 656 91 Brno, Czech Republic.
authors.forename: Lumir
authors.initials: L
authors.lastname: Krejci
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Ramat Aviv 69978, Israel.
authors.forename: Martin
authors.initials: M
authors.lastname: Kupiec
mesh_terms: D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis; D004249:DNA Damage; D004265:DNA Helicases / Q000235:genetics; D042822:Genomic Instability; D008741:Methyl Methanesulfonate / Q000633:toxicity; D009154:Mutation; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D011995:Recombination, Genetic; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D040681:Structural Homology, Protein; D013329:Structure-Activity Relationship; D013489:Suppression, Genetic
chemical_list: D002352:Carrier Proteins; D002843:Chromatin; C477609:Elg1 protein, S cerevisiae; C546594:POL30 protein, S cerevisiae; D018809:Proliferating Cell Nuclear Antigen; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D008741:Methyl Methanesulfonate; C076225:RAD5 protein, S cerevisiae; D004265:DNA Helicases