Mutational analysis of a function of xeroderma pigmentosum group A (XPA) protein in strand-specific DNA repair.
To analyze the function of the xeroderma pigmentosum group A (XPA) protein in strand-specific DNA repair, we examined repair of UV-induced cyclobutane pyrimidine dimer (CPD) in transcribed and non-transcribed strands of the dihydrofolate reductase gene of xeroderma pigmentosum group A (XP-A) cell line (XP12ROSV) which was transfected with various types of mutant XPA cDNA. The transfectant overexpressing mutant XPA with a defect in the interaction with either ERCC1, replication protein A (RPA), or general transcription factor TFIIH, showed more or less decreased repair of CPD in each strand in parallel, while in the transfectant overexpressing R207G (Arg207to Gly) mutant XPA derived from XP129, a UV-resistant XP12ROSV revertant, the rate of CPD repair was almost normal in each strand. We also examined the dose responses of the XPA protein on CPD repair in each strand by the modulation of the expression levels of wild-type or R207G mutant XPA using an inducible expression system, LacSwitchtrade mark promoter. There were good correlations between the rate of CPD repair in each strand and the amount of XPA protein produced in these Lac cells. Our results indicate that the XPA protein is equally important for the CPD repair in both transcribed and non-transcribed strands and that the R207G mutation found in XP129 may not be responsible for a selective defect in CPD repair in the non-transcribed strand in XP129.
Kobayashi
T
T
Institute for Molecular and Cellular Biology, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
Takeuchi
S
S
Saijo
M
M
Nakatsu
Y
Y
Morioka
H
H
Otsuka
E
E
Wakasugi
M
M
Nikaido
O
O
Tanaka
K
K
D002460:Cell Line; D002470:Cell Survival; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D011740:Pyrimidine Dimers / Q000378:metabolism; D013379:Substrate Specificity; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D014162:Transfection; D014466:Ultraviolet Rays; D014983:Xeroderma Pigmentosum / Q000235:genetics / Q000378:metabolism*; D051760:Xeroderma Pigmentosum Group A Protein
D004268:DNA-Binding Proteins; D011740:Pyrimidine Dimers; C497314:XPA protein, human; D051760:Xeroderma Pigmentosum Group A Protein; D013762:Tetrahydrofolate Dehydrogenase