Journal Article

Functional Analysis of Human FEN1 in <i>Saccharomyces Cerevisiae</i> and Its Role in Genome Stability

Amy L. Greene, Joyce R. Snipe, Dmitry A. Gordenin and Michael A. Resnick

in Human Molecular Genetics

Volume 8, issue 12, pages 2263-2273
Published in print November 1999 | ISSN: 0964-6906
Published online November 1999 | e-ISSN: 1460-2083 | DOI: http://dx.doi.org/10.1093/hmg/8.12.2263
Functional Analysis of Human FEN1 in Saccharomyces Cerevisiae and Its Role in Genome Stability

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The flap endonuclease, FEN1, is an evolutionarily conserved component of DNA replication from archaebacteria to humans. Based on in vitro results, it processes Okazaki fragments during replication and is involved in base excision repair. FEN1 removes the last primer ribonucleotide on the lagging strand and it cleaves a 5′ flap that may result from strand displacement during replication or during base excision repair. Its biological importance has been revealed largely through studies in the yeast Saccharomyces cerevisiae where deletion of the homologous gene RAD27 results in genome instability and mutagen sensitivity. While the invivo function of Rad27 has been well characterized through genetic and biochemical approaches, little is understood about the in vivo functions of human FEN1. Guided by our recent results with yeast RAD27, we explored the function of human FEN1 in yeast. We found that the human FEN1 protein complements a yeast rad27 null mutant for a variety of defects including mutagen sensitivity, genetic instability and the synthetic lethal interactions of a rad27 rad51 and a rad27 pol3-01 mutant. Furthermore, a mutant form of FEN1 lacking nuclease function exhibits dominant-negative effects on cell growth and genome instability similar to those seen with the homologous yeast rad27 mutation. This genetic impact is stronger when the human and yeast PCNA-binding domains are exchanged. These data indicate that the human FEN1 and yeast Rad27 proteins act on the same substrate in vivo. Our study defines a sensitive yeast system for the identification and characterization of mutations in FEN1.

Journal Article.  8540 words.  Illustrated.

Subjects: Genetics and Genomics

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