title: In vivo evidence for translesion synthesis by the replicative DNA polymerase δ.
abstract: The intolerance of DNA polymerase δ (Polδ) to incorrect base pairing contributes to its extremely high accuracy during replication, but is believed to inhibit translesion synthesis (TLS). However, chicken DT40 cells lacking the POLD3 subunit of Polδ are deficient in TLS. Previous genetic and biochemical analysis showed that POLD3 may promote lesion bypass by Polδ itself independently of the translesion polymerase Polζ of which POLD3 is also a subunit. To test this hypothesis, we have inactivated Polδ proofreading in pold3 cells. This significantly restored TLS in pold3 mutants, enhancing dA incorporation opposite abasic sites. Purified proofreading-deficient human Polδ holoenzyme performs TLS of abasic sites in vitro much more efficiently than the wild type enzyme, with over 90% of TLS events resulting in dA incorporation. Furthermore, proofreading deficiency enhances the capability of Polδ to continue DNA synthesis over UV lesions both in vivo and in vitro These data support Polδ contributing to TLS in vivo and suggest that the mutagenesis resulting from loss of Polδ proofreading activity may in part be explained by enhanced lesion bypass.
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Sakyo-ku, Kyoto 606-8501, Japan Department of Chemistry, Graduate School of Science and Engineering, Tokyo Metropolitan University, Minami-Osawa 1-1, Hachioji- shi, Tokyo 192-0397, Japan.
authors.forename: Kouji
authors.initials: K
authors.lastname: Hirota
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Masataka
authors.initials: M
authors.lastname: Tsuda
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.lastname: Mohiuddin
authors.affiliation: Department of Biology, School of Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
authors.forename: Toshiki
authors.initials: T
authors.lastname: Tsurimoto
authors.affiliation: Weizmann Institute of Science, Department of Biological Chemistry, Rehovot 76100, Israel.
authors.forename: Isadora S
authors.initials: IS
authors.lastname: Cohen
authors.affiliation: Weizmann Institute of Science, Department of Biological Chemistry, Rehovot 76100, Israel.
authors.forename: Zvi
authors.initials: Z
authors.lastname: Livneh
authors.affiliation: Department of Chemistry, Graduate School of Science and Engineering, Tokyo Metropolitan University, Minami-Osawa 1-1, Hachioji- shi, Tokyo 192-0397, Japan.
authors.forename: Kaori
authors.initials: K
authors.lastname: Kobayashi
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Takeo
authors.initials: T
authors.lastname: Narita
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Kana
authors.initials: K
authors.lastname: Nishihara
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Junko
authors.initials: J
authors.lastname: Murai
authors.affiliation: Division of Chemistry, Graduate School of Engineering Science, Osaka University, 1-3 Machikaneyama, Toyonaka, Osaka 560-8531, Japan.
authors.forename: Shigenori
authors.initials: S
authors.lastname: Iwai
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH, UK.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Guilbaud
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH, UK jes@mrc-lmb.cam.ac.uk.
authors.forename: Julian E
authors.initials: JE
authors.lastname: Sale
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshidakonoe, Sakyo-ku, Kyoto 606-8501, Japan stakeda@rg.med.kyoto-u.ac.jp.
authors.forename: Shunichi
authors.initials: S
authors.lastname: Takeda
mesh_terms: D000483:Alleles; D002460:Cell Line; D004247:DNA / Q000096:biosynthesis* / Q000737:chemistry*; D004249:DNA Damage; D004258:DNA Polymerase III / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004261:DNA Replication; D020035:Holoenzymes / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D007136:Immunoglobulins / Q000235:genetics; D014466:Ultraviolet Rays
chemical_list: D020035:Holoenzymes; D007136:Immunoglobulins; D004247:DNA; D004258:DNA Polymerase III