title: Prereplicative repair of oxidized bases in the human genome is mediated by NEIL1 DNA glycosylase together with replication proteins.
abstract: Base oxidation by endogenous and environmentally induced reactive oxygen species preferentially occurs in replicating single-stranded templates in mammalian genomes, warranting prereplicative repair of the mutagenic base lesions. It is not clear how such lesions (which, unlike bulky adducts, do not block replication) are recognized for repair. Furthermore, strand breaks caused by base excision from ssDNA by DNA glycosylases, including Nei-like (NEIL) 1, would generate double-strand breaks during replication, which are not experimentally observed. NEIL1, whose deficiency causes a mutator phenotype and is activated during the S phase, is present in the DNA replication complex isolated from human cells, with enhanced association with DNA in S-phase cells and colocalization with replication foci containing DNA replication proteins. Furthermore, NEIL1 binds to 5-hydroxyuracil, the oxidative deamination product of C, in replication protein A-coated ssDNA template and inhibits DNA synthesis by DNA polymerase δ. We postulate that, upon encountering an oxidized base during replication, NEIL1 initiates prereplicative repair by acting as a "cowcatcher" and preventing nascent chain growth. Regression of the stalled replication fork, possibly mediated by annealing helicases, then allows lesion repair in the reannealed duplex. This model is supported by our observations that NEIL1, whose deficiency slows nascent chain growth in oxidatively stressed cells, is stimulated by replication proteins in vitro. Furthermore, deficiency of the closely related NEIL2 alone does not affect chain elongation, but combined NEIL1/2 deficiency further inhibits DNA replication. These results support a mechanism of NEIL1-mediated prereplicative repair of oxidized bases in the replicating strand, with NEIL2 providing a backup function.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555, USA. mlhegde@utmb.edu
authors.forename: Muralidhar L
authors.initials: ML
authors.lastname: Hegde
authors.forename: Pavana M
authors.initials: PM
authors.lastname: Hegde
authors.forename: Larry J
authors.initials: LJ
authors.lastname: Bellot
authors.forename: Santi M
authors.initials: SM
authors.lastname: Mandal
authors.forename: Tapas K
authors.initials: TK
authors.lastname: Hazra
authors.forename: Guo-Min
authors.initials: GM
authors.lastname: Li
authors.forename: Istvan
authors.initials: I
authors.lastname: Boldogh
authors.forename: Alan E
authors.initials: AE
authors.lastname: Tomkinson
authors.forename: Sankar
authors.initials: S
authors.lastname: Mitra
mesh_terms: D015153:Blotting, Western; D001973:Bromodeoxyuridine; D047369:Chromatin Immunoprecipitation; D045647:DNA Glycosylases / Q000378:metabolism*; D004258:DNA Polymerase III / Q000378:metabolism; D004260:DNA Repair / Q000235:genetics*; D004261:DNA Replication / Q000502:physiology*; D015894:Genome, Human / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D008856:Microscopy, Fluorescence; D018384:Oxidative Stress / Q000235:genetics / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics
chemical_list: D034741:RNA, Small Interfering; C435720:POLD3 protein, human; D004258:DNA Polymerase III; D045647:DNA Glycosylases; C473634:NEIL1 protein, human; D001973:Bromodeoxyuridine
keywords: genome damage repair; oxidized base repair at DNA replication fork; replication fork stalling
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