Human XPG nuclease structure, assembly, and activities with insights for neurodegeneration and cancer from pathogenic mutations

Susan E. Tsutakawa, Altaf H. Sarker, Clifford Ng, Andrew S. Arvai, David S. Shin, Brian Shih, Shuai Jiang, Aye C. Thwin, Miaw-Sheue Tsai, Alexandra Willcox, Mai Zong Her, Kelly S. Trego, Alan G. Raetz, Daniel Rosenberg, Albino Bacolla, Michal Hammel, Jack D. Griffith, Priscilla K. Cooper, John A. Tainer

Research output: Contribution to journalArticlepeer-review

32 Scopus citations

Abstract

Xeroderma pigmentosum group G (XPG) protein is both a functional partner in multiple DNA damage responses (DDR) and a pathway coordinator and structure-specific endonuclease in nucleotide excision repair (NER). Different mutations in the XPG gene ERCC5 lead to either of two distinct human diseases: Cancer-prone xeroderma pigmentosum (XP-G) or the fatal neurodevelopmental disorder Cockayne syndrome (XP-G/CS). To address the enigmatic structural mechanism for these differing disease phenotypes and for XPG’s role in multiple DDRs, here we determined the crystal structure of human XPG catalytic domain (XPGcat), revealing XPG-specific features for its activities and regulation. Furthermore, XPG DNA binding elements conserved with FEN1 superfamily members enable insights on DNA interactions. Notably, all but one of the known pathogenic point mutations map to XPGcat, and both XP-G and XP-G/CS mutations destabilize XPG and reduce its cellular protein levels. Mapping the distinct mutation classes provides structure-based predictions for disease phenotypes: Residues mutated in XP-G are positioned to reduce local stability and NER activity, whereas residues mutated in XP-G/CS have implied long-range structural defects that would likely disrupt stability of the whole protein, and thus interfere with its functional interactions. Combined data from crystallography, biochemistry, small angle X-ray scattering, and electron microscopy unveil an XPG homodimer that binds, unstacks, and sculpts duplex DNA at internal unpaired regions (bubbles) into strongly bent structures, and suggest how XPG complexes may bind both NER bubble junctions and replication forks. Collective results support XPG scaffolding and DNA sculpting functions in multiple DDR processes to maintain genome stability.
Original languageEnglish (US)
Pages (from-to)201921311
JournalProceedings of the National Academy of Sciences
DOIs
StatePublished - Jun 11 2020
Externally publishedYes

Bibliographical note

KAUST Repository Item: Exported on 2020-10-01
Acknowledgements: We thank the researchers, patients, and families who have critically contributed to defining the genetic basis for xeroderma pigmentosum group G (XPG) pathophysiology and its window into XPG biology; Steffen Emmert for providing primary patient cells XP72MA, XP165MA, and XP40GO; and Samir Hamdan and members of the Structural Biology of DNA Repair program, including Patrick Sung and Orlando Schärer, for comments. Work on XPG was supported by National Cancer Institute P01 CA092584 (to S.E.T., P.K.C., and J.A.T.), R35 CA220430 (to J.A.T.); NIH R01GM110387 (to S.E.T.), NIH P01 AG017242 and R01 ES019935 (to P.K.C.), NIH R01GM31819 and R01 ES013773 (to J.D.G.), NIH T32 AG000266 (to A.G.R.); and King Abdullah University of Science and Technology CRG3 (to J.A.T.). J.A.T. is a Cancer Prevention and Research Institute of Texas Scholar in Cancer Research and acknowledges support
by a Robert A. Welch Chemistry Chair. X-ray diffraction data were collected at the Stanford Synchrotron Radiation Laboratory, supported by the Department of Energy (DOE), Office of Biological and Environmental Research, NIH, National Center for Research Resources, Biomedical Technology Program, and the National Institute of General Medical Sciences, and at Advanced Light Source SIBYLS Beamline 12.3.1, supported by NIH project ALS-ENABLE (P30 GM124169) and the Integrated Diffraction Analysis Technologies program of the US DOE Office of Biological and Environmental Research.
This publication acknowledges KAUST support, but has no KAUST affiliated authors.

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