UV damage mapping reveals different impacts of XPD mutations on global genomic and transcription-coupled DNA repair

ABSTRACT: Nucleotide Excision Repair (NER) removes bulky DNA lesions, such as ultraviolet (UV) light-induced cyclobutane pyrimidine dimers (CPDs) by utilizing two subpathways: Transcription-Coupled NER (TC-NER) and Global Genomic NER (GG-NER). XPD is a DNA helicase in the transcription factor IIH (TFIIH) complex that is important for DNA unwinding and damage verification during NER. Germline mutations in XPD can lead to human genetic disorders, including Xeroderma Pigmentosum (XP) or XP in combination with Cockayne Syndrome (XP/CS). While XP and XP/CS mutations in XPD generally disrupt NER, it remains uncharacterized whether they impair TC-NER, GG-NER or both NER subpathways. Many XPD mutation sites are conserved from archaeal to eukaryotic species. To understand how XPD mutations affect DNA repair, we introduced two pairs of XP and XP/CS mutations into the chromosomal locations of the yeast RAD3 gene, a homolog of human XPD. Interestingly, we found that the two XP/CS point mutations rendered yeast cells extremely sensitive to UV light, while the two XP mutants exhibited mild UV sensitivity. Consistently, repair of CPDs in the genomic DNA was slower in the two XP/CS relative to the two XP mutants. Using CPD sequencing (CPD-seq 2.0), a genome-wide UV damage sequencing method, our data indicates that XP/CS mutations abrogated repair by both GG-NER and TC-NER in the yeast genome, while XP mutations impaired GG-NER but retained significant TC-NER activity. In human XP-D cell lines, we also found that XP/CS cells showed greater sensitivity to Illudin S, a DNA damaging agent triggering TC-NER repair. Our study suggests that the XP/CS mutations in RAD3 or XPD gene have more severe DNA repair defects, particularly in the TC-NER subpathway, which may contribute to the Cockayne Syndrome phenotypes of some XP-D patients.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE334098 | GEO | 2026/08/31

REPOSITORIES: GEO