Impact of DNA sequences on DNA ‘opening’ by the Rad4/XPC nucleotide excision repair complex

Debamita Paul, Hong Mu, Amirrasoul Tavakoli, Qing Dai, Sagnik Chakraborty, Chuan He, Anjum Ansari, Suse Broyde, Jung Hyun Min

Research output: Contribution to journalArticlepeer-review

Abstract

Rad4/XPC recognizes diverse DNA lesions to initiate nucleotide excision repair (NER). However, NER propensities among lesions vary widely and repair-resistant lesions are persistent and thus highly mutagenic. Rad4 recognizes repair-proficient lesions by unwinding (‘opening’) the damaged DNA site. Such ‘opening’ is also observed on a normal DNA sequence containing consecutive C/G's (CCC/GGG) when tethered to Rad4 to prevent protein diffusion. However, it was unknown if such tethering-facilitated DNA ‘opening’ could occur on any DNA or if certain structures/sequences would resist being ‘opened’. Here, we report that DNA containing alternating C/G's (CGC/GCG) failed to be opened even when tethered; instead, Rad4 bound in a 180°-reversed manner, capping the DNA end. Fluorescence lifetime studies of DNA conformations in solution showed that CCC/GGG exhibits local pre-melting that is absent in CGC/GCG. In MD simulations, CGC/GCG failed to engage Rad4 to promote ‘opening’ contrary to CCC/GGG. Altogether, our study illustrates how local sequences can impact DNA recognition by Rad4/XPC and how certain DNA sites resist being ‘opened’ even with Rad4 held at that site indefinitely. The contrast between CCC/GGG and CGC/GCG sequences in Rad4-DNA recognition may help decipher a lesion's mutagenicity in various genomic sequence contexts to explain lesion-determined mutational hot and cold spots.

Original languageEnglish (US)
Article number103194
JournalDNA Repair
Volume107
DOIs
StatePublished - Nov 2021

Keywords

  • DNA damage recognition
  • Fluorescence lifetime
  • Förster resonance energy transfer
  • Molecular dynamics simulation
  • Nucleotide excision repair
  • Protein-DNA interaction
  • Rad4
  • Sequence impact
  • Time-resolved fluorescence
  • x-ray crystallography
  • Xeroderma pigmentosum
  • XPC

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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