Lesion search mechanism of human NER protein

Abstract

DNA damage repair is critical for the genomic stability and integrity. In particular, searching for DNA lesions is important because it initiates the entire repair process. In nucleotide excision repair (NER) that is a conserved and versatile repair mechanism, Xeroderma pigmentosum complementation group C protein (XPC) finds DNA lesions and recruits downstream factors. structural studies revealed the molecular feature of damage identification by XPC, and single-molecule approach reported the diffusion of XPC on DNA. However, how XPC can recognize the defects on DNA while it diffuses along DNA and what factors influence the diffusive motions of XPC still remain elusive. To reveal the detailed mechanism behind damage search of XPC, we visualized the motion of human XPC-Rad23B (hXPC-Rad23B) on undamaged or lesion-containing DNA using a high-throughput single-molecule imaging technique, DNA curtain. We observed the heterogeneity in motions of hXPC-Rad23B, exhibiting diffusive, constrained, and immobile species. We found that the heterogeneity results from the interaction between hXPC-Rad23B and DNA breathing on consecutive AT-tracks. In diffusive motion, the diffusion coefficient dramatically increases according to ionic strength, suggesting that hXPC-Rad23B diffuse along DNA via hopping, which was further supported by our finding that hXPC-Rad23B can bypass protein obstacles upon collision. Furthermore, we found that hXPC-Rad23B recognized cyclobutane pyrimidine dimers (CPDs) with low efficiency, proposing that another factor is necessitated. Taken together, our results give an insight into how hXPC-Rad23B can rapidly finds DNA lesions in billions of base pairs of human genome.