DNA repair factor XPC is modified by SUMO-1 and ubiquitin following UV irradiation.
Nucleotide excision repair (NER) is the major DNA repair process that removes diverse DNA lesions including UV-induced photoproducts. There are more than 20 proteins involved in NER. Among them, XPC is thought to be one of the first proteins to recognize DNA damage during global genomic repair (GGR), a sub-pathway of NER. In order to study the mechanism through which XPC participates in GGR, we investigated the possible modifications of XPC protein upon UV irradiation in mammalian cells. Western blot analysis of cell lysates from UV-irradiated normal human fibroblast, prepared by direct boiling in an SDS lysis buffer, showed several anti-XPC antibody-reactive bands with molecular weight higher than the original XPC protein. The reciprocal immunoprecipitation and siRNA transfection analysis demonstrated that XPC protein is modified by SUMO-1 and ubiquitin. By using several NER-deficient cell lines, we found that DDB2 and XPA are required for UV-induced XPC modifications. Interestingly, both the inactivation of ubiquitylation and the treatment of proteasome inhibitors quantitatively inhibited the UV-induced XPC modifications. Furthermore, XPC protein is degraded significantly following UV irradiation in XP-A cells in which sumoylation of XPC does not occur. Taken together, we conclude that XPC protein is modified by SUMO-1 and ubiquitin following UV irradiation and these modifications require the functions of DDB2 and XPA, as well as the ubiquitin-proteasome system. Our results also suggest that at least one function of UV-induced XPC sumoylation is related to the stabilization of XPC protein.
Wang
Qi-En
QE
Department of Radiology, The Ohio State University 103 Wiseman Hall, 400 W. 12th Avenue, Columbus, OH 43210, USA.
Zhu
Qianzheng
Q
Wani
Gulzar
G
El-Mahdy
Mohamed A
MA
Li
Jinyou
J
Wani
Altaf A
AA
D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism* / Q000502:physiology; D005347:Fibroblasts / Q000378:metabolism / Q000528:radiation effects; D006801:Humans; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D025842:SUMO-1 Protein / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*; D014466:Ultraviolet Rays; D051760:Xeroderma Pigmentosum Group A Protein
C117669:DDB2 protein, human; D004268:DNA-Binding Proteins; D025842:SUMO-1 Protein; D025801:Ubiquitin; C497314:XPA protein, human; D051760:Xeroderma Pigmentosum Group A Protein; C086984:XPC protein, human; D046988:Proteasome Endopeptidase Complex