A four-subunit DNA polymerase ζ complex containing Pol δ accessory subunits is essential for PCNA-mediated mutagenesis

Alena V. Makarova, Joseph L. Stodola, Peter M. Burgers

Research output: Contribution to journalArticle

111 Scopus citations

Abstract

DNA polymerase ζ (Pol ζ) plays a key role in DNA translesion synthesis (TLS) and mutagenesis in eukaryotes. Previously, a two-subunit Rev3-Rev7 complex had been identified as the minimal assembly required for catalytic activity in vitro. Herein, we show that Saccharomyces cerevisiae Pol ζ binds to the Pol31 and Pol32 subunits of Pol δ, forming a four-subunit Pol ζ4 complex (Rev3-Rev7-Pol31-Pol32). A [4Fe-4S] cluster in Rev3 is essential for the formation of Pol ζ4 and damage-induced mutagenesis. Pol32 is indispensible for complex formation, providing an explanation for the long-standing observation that pol32Δ strains are defective for mutagenesis. The Pol31 and Pol32 subunits are also required for proliferating cell nuclear antigen (PCNA)-dependent TLS by Pol ζ as Pol ζ2 lacks functional interactions with PCNA. Mutation of the C-terminal PCNA-interaction motif in Pol32 attenuates PCNA-dependent TLS in vitro and mutagenesis in vivo. Furthermore, a mutant form of PCNA, encoded by the mutagenesis-defective pol30-113 mutant, fails to stimulate Pol ζ4 activity, providing an explanation for the observed mutagenesis phenotype. A stable Pol ζ4 complex can be identified in all phases of the cell cycle suggesting that this complex is not regulated at the level of protein interactions between Rev3-Rev7 and Pol31-Pol32.

Original languageEnglish
Pages (from-to)11618-11626
Number of pages9
JournalNucleic acids research
Volume40
Issue number22
DOIs
StatePublished - Dec 1 2012

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