title: Involvement of Werner syndrome protein in MUTYH-mediated repair of oxidative DNA damage.
abstract: Reactive oxygen species constantly generated as by-products of cellular metabolism readily attack genomic DNA creating mutagenic lesions such as 7,8-dihydro-8-oxo-guanine (8-oxo-G) that promote aging. 8-oxo-G:A mispairs arising during DNA replication are eliminated by base excision repair initiated by the MutY DNA glycosylase homologue (MUTYH). Here, by using formaldehyde crosslinking in mammalian cell extracts, we demonstrate that the WRN helicase/exonuclease defective in the premature aging disorder Werner syndrome (WS) is recruited to DNA duplex containing an 8-oxo-G:A mispair in a manner dependent on DNA polymerase λ (Polλ) that catalyzes accurate DNA synthesis over 8-oxo-G. Similarly, by immunofluorescence, we show that Polλ is required for accumulation of WRN at sites of 8-oxo-G lesions in human cells. Moreover, we show that nuclear focus formation of WRN and Polλ induced by oxidative stress is dependent on ongoing DNA replication and on the presence of MUTYH. Cell viability assays reveal that depletion of MUTYH suppresses the hypersensitivity of cells lacking WRN and/or Polλ to oxidative stress. Biochemical studies demonstrate that WRN binds to the catalytic domain of Polλ and specifically stimulates DNA gap filling by Polλ over 8-oxo-G followed by strand displacement synthesis. Our results suggest that WRN promotes long-patch DNA repair synthesis by Polλ during MUTYH-initiated repair of 8-oxo-G:A mispairs.
authors: Institute of Molecular Cancer Research, University of Zurich, Winterthurerstrasse 190, CH-8057 Zurich, Switzerland.
authors: Radhakrishnan
authors: R
authors: Kanagaraj
authors: Prasanna
authors: P
authors: Parasuraman
authors: Boris
authors: B
authors: Mihaljevic
authors: Barbara
authors: B
authors: van Loon
authors: Kamila
authors: K
authors: Burdova
authors: Christiane
authors: C
authors: König
authors: Antonia
authors: A
authors: Furrer
authors: Vilhelm A
authors: VA
authors: Bohr
authors: Ulrich
authors: U
authors: Hübscher
authors: Pavel
authors: P
authors: Janscak
mesh_terms: D000818:Animals; D020137:Base Pair Mismatch; D002460:Cell Line; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D045647:DNA Glycosylases / Q000378:metabolism*; D019951:DNA Polymerase beta / Q000378:metabolism; D004260:DNA Repair; D004261:DNA Replication; D005090:Exodeoxyribonucleases / Q000378:metabolism* / Q000502:physiology; D006147:Guanine / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D051379:Mice; D018384:Oxidative Stress; D053484:RecQ Helicases / Q000378:metabolism* / Q000502:physiology; D016196:S Phase / Q000235:genetics; D000071657:Werner Syndrome Helicase
chemical_list: C453560:7,8-dihydro-8-oxoguanine; D006147:Guanine; D004247:DNA; C415367:DNA polymerase beta2; D019951:DNA Polymerase beta; D005090:Exodeoxyribonucleases; D045647:DNA Glycosylases; C061738:mutY adenine glycosylase; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase