title: APE2 Zf-GRF facilitates 3'-5' resection of DNA damage following oxidative stress.
abstract: The Xenopus laevis APE2 (apurinic/apyrimidinic endonuclease 2) nuclease participates in 3'-5' nucleolytic resection of oxidative DNA damage and activation of the ATR-Chk1 DNA damage response (DDR) pathway via ill-defined mechanisms. Here we report that APE2 resection activity is regulated by DNA interactions in its Zf-GRF domain, a region sharing high homology with DDR proteins Topoisomerase 3α (TOP3α) and NEIL3 (Nei-like DNA glycosylase 3), as well as transcription and RNA regulatory proteins, such as TTF2 (transcription termination factor 2), TFIIS, and RPB9. Biochemical and NMR results establish the nucleic acid-binding activity of the Zf-GRF domain. Moreover, an APE2 Zf-GRF X-ray structure and small-angle X-ray scattering analyses show that the Zf-GRF fold is typified by a crescent-shaped ssDNA binding claw that is flexibly appended to an APE2 endonuclease/exonuclease/phosphatase (EEP) catalytic core. Structure-guided Zf-GRF mutations impact APE2 DNA binding and 3'-5' exonuclease processing, and also prevent efficient APE2-dependent RPA recruitment to damaged chromatin and activation of the ATR-Chk1 DDR pathway in response to oxidative stress in Xenopus egg extracts. Collectively, our data unveil the APE2 Zf-GRF domain as a nucleic acid interaction module in the regulation of a key single-strand break resection function of APE2, and also reveal topologic similarity of the Zf-GRF to the zinc ribbon domains of TFIIS and RPB9.
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: Bret D
authors.initials: BD
authors.lastname: Wallace
authors.affiliation: Department of Biological Sciences, University of North Carolina at Charlotte, Charlotte, NC 28223.
authors.forename: Zachary
authors.initials: Z
authors.lastname: Berman
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: Geoffrey A
authors.initials: GA
authors.lastname: Mueller
authors.affiliation: Department of Biological Sciences, University of North Carolina at Charlotte, Charlotte, NC 28223.
authors.forename: Yunfeng
authors.initials: Y
authors.lastname: Lin
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: Timothy
authors.initials: T
authors.lastname: Chang
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: Sara N
authors.initials: SN
authors.lastname: Andres
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Wojtaszek
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: Eugene F
authors.initials: EF
authors.lastname: DeRose
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: C Denise
authors.initials: CD
authors.lastname: Appel
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709.
authors.forename: Robert E
authors.initials: RE
authors.lastname: London
authors.affiliation: Department of Biological Sciences, University of North Carolina at Charlotte, Charlotte, NC 28223 shan.yan@uncc.edu williamsrs@niehs.nih.gov.
authors.forename: Shan
authors.initials: S
authors.lastname: Yan
authors.affiliation: Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC 27709 shan.yan@uncc.edu williamsrs@niehs.nih.gov.
authors.forename: R Scott
authors.initials: RS
authors.lastname: Williams
mesh_terms: D000818:Animals; D004249:DNA Damage / Q000235:genetics*; D045647:DNA Glycosylases / Q000378:metabolism; D004260:DNA Repair / Q000235:genetics; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000378:metabolism*; D004720:Endonucleases / Q000378:metabolism; D018384:Oxidative Stress / Q000235:genetics*; D000072417:Protein Domains / Q000235:genetics; D014982:Xenopus laevis / Q000235:genetics / Q000378:metabolism
chemical_list: D004720:Endonucleases; D045647:DNA Glycosylases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D004264:DNA Topoisomerases, Type I
keywords: APE2; Xenopus laevis; Zf-GRF; crystallography; oxidative stress