The 3′→5′ Exonuclease of DNA Polymerase δ can Substitute for the 5′ Flap Endonuclease Rad27/Fen1 in Processing Okazaki Fragments and Preventing Genome Instability

• Published in: Proceedings of the National Academy of Sciences - PNAS 2001-04, Vol.98 (9), p.5122-5127
• Main Authors: Jin, Yong Hwan, Obert, Robyn, Peter M. J. Burgers, Kunkel, Thomas A., Resnick, Michael A., Gordenin, Dmitry A.
• Format: Article
• Language: English
• Subjects: Alleles; Amino Acid Transport Systems; Amino acids; Biological Sciences; Chromosomes, Fungal - genetics; DNA; DNA Polymerase III - genetics; DNA Polymerase III - metabolism; DNA Repair - genetics; DNA Replication - genetics; Endodeoxyribonucleases - deficiency; Endodeoxyribonucleases - genetics; Endodeoxyribonucleases - metabolism; Exodeoxyribonucleases - deficiency; Exodeoxyribonucleases - genetics; Exodeoxyribonucleases - metabolism; Flap Endonucleases; Fungal Proteins; Gene Deletion; Gene Duplication; Genes, Lethal - genetics; Genetic Complementation Test; Genetic mutation; Genome, Fungal; Genomes; Kinetics; Membrane Transport Proteins - genetics; Multienzyme Complexes - deficiency; Multienzyme Complexes - genetics; Multienzyme Complexes - metabolism; Mutagenesis - genetics; Phenotypes; Plasmids; Proofreading; Recombination, Genetic - genetics; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - growth & development; Saccharomyces cerevisiae Proteins; Viability; Yeasts
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.091095198

Many DNA polymerases (Pol) have an intrinsic 3′→5′ exonuclease (Exo) activity which corrects polymerase errors and prevents mutations. We describe a role of the 3′→5′ Exo of Pol δ as a supplement or backup for the Rad27/Fen1 5′ flap endonuclease. A yeast rad27 null allele was lethal in combination with Pol δ mutations in Exo I, Exo II, and Exo III motifs that inactivate its exonuclease, but it was viable with mutations in other parts of Pol δ. The rad27-p allele, which has little phenotypic effect by itself, was also lethal in combination with mutations in the Pol δ Exo I and Exo II motifs. However, rad27-p Pol δ Exo III double mutants were viable. They exhibited strong synergistic increases in CAN1 duplication mutations, intrachromosomal and interchromosomal recombination, and required the wild-type double-strand break repair genes RAD50, RAD51, and RAD52 for viability. Observed effects were similar to those of the rad27-null mutant deficient in the removal of 5′ flaps in the lagging strand. These results suggest that the 3′→5′ Exo activity of Pol δ is redundant with Rad27/Fen1 for creating ligatable nicks between adjacent Okazaki fragments, possibly by reducing the amount of strand-displacement in the lagging strand.