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Genetics, Vol. 151, 499-509, February 1999, Copyright © 1999

Characterization of the Repeat-Tract Instability and Mutator Phenotypes Conferred by a Tn3 Insertion in RFC1, the Large Subunit of the Yeast Clamp Loader

Yali Xiea, Chris Counterb, and Eric Alania
a Section of Genetics and Development, Cornell University, Ithaca, New York 14853-2703
b Department of Pharmacology and Cancer Biology, Department of Radiation Oncology, Duke University Medical Center, Durham, North Carolina 27710

Corresponding author: Eric Alani, Section of Genetics and Development, Cornell University, 459 Biotechnology Bldg., Ithaca, NY 14853-2703., eea3{at}cornell.edu (E-mail)

Communicating editor: P. L. FOSTER

The RFC1 gene encodes the large subunit of the yeast clamp loader (RFC) that is a component of eukaryotic DNA polymerase holoenzymes. We identified a mutant allele of RFC1 (rfc1::Tn3) from a large collection of Saccharomyces cerevisiae mutants that were inviable when present in a rad52 null mutation background. Analysis of rfc1::Tn3 strains indicated that they displayed both a mutator and repeat-tract instability phenotype. Strains bearing this allele were characterized in combination with mismatch repair (msh2{Delta}, pms1{Delta}), double-strand break repair (rad52), and DNA replication (pol3-01, pol30-52, rth1{Delta}/rad27{Delta}) mutations in both forward mutation and repeat-tract instability assays. This analysis indicated that the rfc1::Tn3 allele displays synthetic lethality with pol30, pol3, and rad27 mutations. Measurement of forward mutation frequencies in msh2{Delta} rfc1:Tn3 and pms1{Delta} rfc1:Tn3 strains indicated that the rfc1::Tn3 mutant displayed a mutation frequency that appeared nearly multiplicative with the mutation frequency exhibited by mismatch-repair mutants. In repeat-tract instability assays, however, the rfc1::Tn3 mutant displayed a tract instability phenotype that appeared epistatic to the phenotype displayed by mismatch-repair mutants. From these data we propose that defects in clamp loader function result in DNA replication errors, a subset of which are acted upon by the mismatch-repair system.




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