Regulation of nucleotide excision repair by UV-DDB: prioritization of damage recognition to internucleosomal DNA.
How tightly packed chromatin is thoroughly inspected for DNA damage is one of the fundamental unanswered questions in biology. In particular, the effective excision of carcinogenic lesions caused by the ultraviolet (UV) radiation of sunlight depends on UV-damaged DNA-binding protein (UV-DDB), but the mechanism by which this DDB1-DDB2 heterodimer stimulates DNA repair remained enigmatic. We hypothesized that a distinctive function of this unique sensor is to coordinate damage recognition in the nucleosome repeat landscape of chromatin. Therefore, the nucleosomes of human cells have been dissected by micrococcal nuclease, thus revealing, to our knowledge for the first time, that UV-DDB associates preferentially with lesions in hypersensitive, hence, highly accessible internucleosomal sites joining the core particles. Surprisingly, the accompanying CUL4A ubiquitin ligase activity is necessary to retain the xeroderma pigmentosum group C (XPC) partner at such internucleosomal repair hotspots that undergo very fast excision kinetics. This CUL4A complex thereby counteracts an unexpected affinity of XPC for core particles that are less permissive than hypersensitive sites to downstream repair subunits. That UV-DDB also adopts a ubiquitin-independent function is evidenced by domain mapping and in situ protein dynamics studies, revealing direct but transient interactions that promote a thermodynamically unfavorable β-hairpin insertion of XPC into substrate DNA. We conclude that the evolutionary advent of UV-DDB correlates with the need for a spatiotemporal organizer of XPC positioning in higher eukaryotic chromatin.
Institute of Pharmacology and Toxicology, University of Zürich-Vetsuisse, Zürich, Switzerland.
Jia
J
Fei
Nina
N
Kaczmarek
Andreas
A
Luch
Andreas
A
Glas
Thomas
T
Carell
Hanspeter
H
Naegeli
D000818:Animals; D001565:Benzoates / Q000494:pharmacology; D016466:CHO Cells; D006224:Cricetinae; D044842:Cullin Proteins / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000378:metabolism / Q000528:radiation effects; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005663:Furans / Q000494:pharmacology; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D009707:Nucleosomes / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D011720:Pyrazoles / Q000494:pharmacology; D011740:Pyrimidine Dimers / Q000378:metabolism; D034622:RNA Interference; D044764:Ubiquitin-Activating Enzymes / Q000037:antagonists & inhibitors; D054875:Ubiquitination; D014466:Ultraviolet Rays
C523934:4(4-(5-nitro-furan-2-ylmethylene)-3,5-dioxo-pyrazolidin-1-yl)-benzoic acid ethyl ester; D001565:Benzoates; C114234:CUL4A protein, human; D044842:Cullin Proteins; C117669:DDB2 protein, human; D004268:DNA-Binding Proteins; D005663:Furans; D009707:Nucleosomes; D011720:Pyrazoles; D011740:Pyrimidine Dimers; C086984:XPC protein, human; D004247:DNA; D044764:Ubiquitin-Activating Enzymes