Relative levels of the two mammalian Rad23 homologs determine composition and stability of the xeroderma pigmentosum group C protein complex.
Mammalian cells express two Rad23 homologs, HR23A and HR23B, which have been implicated in regulation of proteolysis via the ubiquitin/proteasome pathway. Recently, the proteins have been shown to stabilize xeroderma pigmentosum group C (XPC) protein that is involved in DNA damage recognition for nucleotide excision repair (NER). Because the vast majority of XPC forms a complex with HR23B rather than HR23A, we investigated possible differences between the two Rad23 homologs in terms of their effects on the XPC protein. In wild-type mouse embryonic fibroblasts (MEFs), endogenous XPC was found to be relatively stable, while its steady-state level and stability appeared significantly reduced by targeted disruption of the mHR23B gene, but not by that of mHR23A. Loss of both mHR23 genes caused a strong further reduction of the XPC protein level. Quantification of the two mHR23 proteins revealed that in normal cells mHR23B is actually approximately 10 times more abundant than mHR23A. In addition, overexpression of mHR23A in the mHR23A/B double knock out cells restored not only the steady-state level and stability of the XPC protein, but also cellular NER activity to near wild-type levels. These results indicate that the two Rad23 homologs are largely functionally equivalent in NER, and that the difference in expression levels explains for a major part the difference in complex formation with as well as stabilization effects on XPC.
Okuda
Yuki
Y
Cellular Physiology Laboratory, RIKEN Discovery Research Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Nishi
Ryotaro
R
Ng
Jessica M Y
JM
Vermeulen
Wim
W
van der Horst
Gijsbertus T J
GT
Mori
Toshio
T
Hoeijmakers
Jan H J
JH
Hanaoka
Fumio
F
Sugasawa
Kaoru
K
D000818:Animals; D003433:Crosses, Genetic; D004249:DNA Damage; D004260:DNA Repair; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D014983:Xeroderma Pigmentosum
D004268:DNA-Binding Proteins; C513287:RAD23B protein, human; C475578:Rad23a protein, mouse; C499250:Rad23b protein, mouse; C501002:Xpc protein, mouse; C086985:RAD23A protein, human; D045643:DNA Repair Enzymes