Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 114, Issue 2, Pages 304-309Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1610011114
Keywords
oxidative stress; APE2; Zf-GRF; crystallography; Xenopus laevis
Categories
Funding
- University of North Carolina at Charlotte
- National Institute of General Medical Sciences of the National Institutes of Health (NIH) [R15 GM101571, R15 GM114713]
- DOE's Office of Biological and Environmental Research
- NIH Project MINOS [R01 GM105404]
- DOE, Office of Science, Office of Basic Energy Sciences [W-31-109-Eng-38]
- NIH Intramural Program
- NIEHS [1Z01ES102765, 1Z01ES050111]
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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 3 alpha(TOP3 alpha) 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.
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