Human XPG nuclease structure, assembly, and activities with insights for neurodegeneration and cancer from pathogenic mutations
A partir de données issues d'analyses cristallographiques, biochimiques et microscopiques, cette étude examine la structure du domaine catalytique de la nucléase humaine XPG, une protéine impliquée dans plusieurs voies de réparation de l'ADN, identifie les caractéristiques structurelles liées à ses activités enzymatiques et à sa régulation, puis explique comment des mutations au niveau du gène codant pour XPG peuvent induire une neurodégénérescence ou le développement d'un xeroderma pigmentosum, une maladie se caractérisant notamment par un risque accru de cancer de la peau
DNA repair is essential to life and to avoidance of genome instability and cancer. Xeroderma pigmentosum group G (XPG) protein acts in multiple DNA repair pathways, both as an active enzyme and as a scaffold for coordinating with other repair proteins. We present here the structure of the catalytic domain responsible for its DNA binding and nuclease activity. Our analysis provides structure-based hypotheses for how XPG recognizes its bubble DNA substrate and predictions of the structural impacts of XPG disease mutations associated with two phenotypically distinct diseases: xeroderma pigmentosum (XP, skin cancer prone) or Cockayne syndrome (XP/CS, severe progressive developmental defects).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.