title: The Ctf18RFC clamp loader is essential for telomere stability in telomerase-negative and mre11 mutant alleles.
abstract: The function of the replication clamp loaders in the semi-conservative telomere replication and their relationship to telomerase- and recombination mechanisms of telomere addition remains ambiguous. We have investigated the variant clamp loader Ctf18 RFC (Replication Factor C). To understand the role of Ctf18 at the telomere, we first investigated genetic interactions after loss of Ctf18 and TLC1 (the yeast telomerase RNA). We find that the tlc1Δ ctf18Δ double mutant confers a rapid >1000-fold decrease in viability. The rate of loss was similar to the kinetics of cell death in rad52Δ tlc1Δ cells. However, the Ctf18 pathway is distinct from Rad52, required for the repair of DSBs, as demonstrated by the synthetic lethality of rad52▵ tlc1Δ ctf18Δ triple mutants. These data suggest that each mutant elicits non-redundant defects acting on the same substrate. Second, interactions of the yeast hyper-recombinational mutant, mre11A470T, with ctf18▵ confer a synergistic cold sensitivity. The phenotype of these double mutants ultimately results in telomere loss and the generation of recombinational survivors. We observed a similar synergism between single mutants that led to hypersensitivity to the DNA alkylating agent, methane methyl sulphonate (MMS), the replication fork inhibitor hydroxyurea (HU), and to a failure to separate telomeres of sister chromatids. Hence, ctf18Δ and mre11A470T act in different pathways on telomere substrates for multiple phenotypes. The mre11A470T cells also displayed a DNA damage response (DDR) at 15°C but not at 30°C while ctf18Δ mutants conferred a constitutive DDR activity. Both the 15°C DDR pattern and growth rate were reversible at 30°C and displayed telomerase activity in vivo. We hypothesize that Ctf18 confers protection against stalling and/or breaks at the replication fork in cells that either lack, or are compromised for, telomerase activity. This Ctf18-based function is likely to contribute another level to telomere size homeostasis.
authors.affiliation: Department of Biochemistry and Molecular Biology, Tulane University Medical Center, New Orleans, Louisiana, United States of America.
authors.forename: Honghai
authors.initials: H
authors.lastname: Gao
authors.affiliation: Department of Biochemistry and Molecular Biology, Tulane University Medical Center, New Orleans, Louisiana, United States of America.
authors.forename: Daniel L
authors.initials: DL
authors.lastname: Moss
authors.affiliation: Department of Biochemistry and Molecular Biology, Tulane University Medical Center, New Orleans, Louisiana, United States of America.
authors.forename: Courtney
authors.initials: C
authors.lastname: Parke
authors.affiliation: Department of Biochemistry and Molecular Biology, Tulane University Medical Center, New Orleans, Louisiana, United States of America.
authors.forename: Danielle
authors.initials: D
authors.lastname: Tatum
authors.affiliation: Department of Biochemistry and Molecular Biology, Tulane University Medical Center, New Orleans, Louisiana, United States of America.
authors.forename: Arthur J
authors.initials: AJ
authors.lastname: Lustig
mesh_terms: D000483:Alleles; D018797:Cell Cycle Proteins / Q000378:metabolism; D064447:Checkpoint Kinase 2 / Q000378:metabolism; D002842:Chromatids / Q000737:chemistry; D004260:DNA Repair; D004706:Endodeoxyribonucleases / Q000235:genetics* / Q000378:metabolism; D005090:Exodeoxyribonucleases / Q000235:genetics* / Q000378:metabolism; D006918:Hydroxyurea / Q000737:chemistry; D007700:Kinetics; D008741:Methyl Methanesulfonate / Q000737:chemistry; D009154:Mutation; D010641:Phenotype; D051818:Replication Protein C / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism*; D019098:Telomerase / Q000378:metabolism*; D016615:Telomere / Q000648:ultrastructure*; D013696:Temperature
chemical_list: C092693:CTF18 protein, S cerevisiae; D018797:Cell Cycle Proteins; D029701:Saccharomyces cerevisiae Proteins; D008741:Methyl Methanesulfonate; D064447:Checkpoint Kinase 2; C067022:RAD53 protein, S cerevisiae; D019098:Telomerase; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C094191:MRE11 protein, S cerevisiae; D051818:Replication Protein C; D006918:Hydroxyurea