Faculty, Staff and Student Publications
A Scanning-to-Incision Switch in Tfiih-Xpg Induced by DNA Damage Licenses Nucleotide Excision Repair
Publication Date
2-22-2023
Journal
Nucleic Acids Research
Abstract
Nucleotide excision repair (NER) is critical for removing bulky DNA base lesions and avoiding diseases. NER couples lesion recognition by XPC to strand separation by XPB and XPD ATPases, followed by lesion excision by XPF and XPG nucleases. Here, we describe key regulatory mechanisms and roles of XPG for and beyond its cleavage activity. Strikingly, by combing single-molecule imaging and bulk cleavage assays, we found that XPG binding to the 7-subunit TFIIH core (coreTFIIH) stimulates coreTFIIH-dependent double-strand (ds)DNA unwinding 10-fold, and XPG-dependent DNA cleavage by up to 700-fold. Simultaneous monitoring of rates for coreTFIIH single-stranded (ss)DNA translocation and dsDNA unwinding showed XPG acts by switching ssDNA translocation to dsDNA unwinding as a likely committed step. Pertinent to the NER pathway regulation, XPG incision activity is suppressed during coreTFIIH translocation on DNA but is licensed when coreTFIIH stalls at the lesion or when ATP hydrolysis is blocked. Moreover, ≥15 nucleotides of 5'-ssDNA is a prerequisite for efficient translocation and incision. Our results unveil a paired coordination mechanism in which key lesion scanning and DNA incision steps are sequentially coordinated, and damaged patch removal is only licensed after generation of ≥15 nucleotides of 5'-ssDNA, ensuring the correct ssDNA bubble size before cleavage.
Keywords
DNA, DNA Damage, DNA Repair, DNA, Single-Stranded, Endonucleases, Nucleotides, Transcription Factor TFIIH
DOI
10.1093/nar/gkac1095
PMID
36477609
PMCID
PMC9943652
PubMedCentral® Posted Date
12-8-2022
PubMedCentral® Full Text Version
Post-print
Published Open-Access
yes
Included in
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