Single-molecule visualization reveals the damage search mechanism for the human NER protein XPC-RAD23B

被引:44
作者
Cheon, Na Young [1 ]
Kim, Hyun-Suk [2 ]
Yeo, Jung-Eun [2 ]
Scharer, Orlando D. [1 ,2 ]
Lee, Ja Yil [1 ,2 ]
机构
[1] Ulsan Natl Inst Sci & Technol, Sch Life Sci, Ulsan 44919, South Korea
[2] Inst Basic Sci, Ctr Genom Integr, Ulsan 44919, South Korea
基金
新加坡国家研究基金会;
关键词
NUCLEOTIDE EXCISION-REPAIR; DIFFUSION-DRIVEN MECHANISMS; DNA-DAMAGE; IMAGING REVEALS; NUCLEIC-ACIDS; DUAL INCISION; RECOGNITION; TRANSLOCATION; COMPLEX; XPC;
D O I
10.1093/nar/gkz629
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
DNA repair is critical for maintaining genomic integrity. Finding DNA lesions initiates the entire repair process. In human nucleotide excision repair (NER), XPC-RAD23B recognizes DNA lesions and recruits downstream factors. Although previous studies revealed the molecular features of damage identification by the yeast orthologs Rad4-Rad23, the dynamic mechanisms by which human XPC-RAD23B recognizes DNA defects have remained elusive. Here, we directly visualized the motion of XPC-RAD23B on undamaged and lesion-containing DNA using high-throughput single-molecule imaging. We observed three types of one-dimensional motion of XPC-RAD23B along DNA: diffusive, immobile and constrained. We found that consecutive AT-tracks led to increase in proteins with constrained motion. The diffusion coefficient dramatically increased according to ionic strength, suggesting that XPC-RAD23B diffuses along DNA via hopping, allowing XPC-RAD23B to bypass protein obstacles during the search for DNA damage. We also examined how XPC-RAD23B identifies cyclobutane pyrimidine dimers (CPDs) during diffusion. XPC-RAD23B makes futile attempts to bind to CPDs, consistent with low CPD recognition efficiency. Moreover, XPC-RAD23B binds CPDs in biphasic states, stable for lesion recognition and transient for lesion interrogation. Taken together, our results provide new insight into how XPC-RAD23B searches for DNA lesions in billions of base pairs in human genome.
引用
收藏
页码:8337 / 8347
页数:11
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