Efficient PCNA complex formation is dependent upon both transcription coupled repair and genome overall repair.
The protein proliferating cell nuclear antigen (PCNA) is an auxiliary factor for DNA polymerase delta and is involved in the resynthesis step of nucleotide excision repair (NER). After UV irradiation of quiescent cells, PCNA forms an insoluble complex with nuclear substructures. We have investigated associations between NER and its subcomponent pathway, transcription coupled repair (TCR) on PCNA complex formation using genetically related hamster cell lines with different repair characteristics. In DNA repair proficient cells, the PCNA complex was readily detectable within 30 min after UV irradiation by both immunofluorescence and western blot analyses. This complex formation after UV occurs efficiently in quiescent cells. In UV5 (human XP-D homolog) and UV 24 (human XP-B homolog) cells, which are totally deficient in NER, the PCNA complex was not detectable at 30 min after UV. The PCNA complex formation is restored to normal levels in UV5 cells after transfection with the human XPD gene, encoding a subunit of the basal transcription factor, TFIIH. In UV61 (Human CS-B homolog) cells, that are defective only in transcription coupled repair (TCR) of cyclobutane pyrimidine dimers (CPDs), the rate of PCNA complex formation was 2-fold slower than in repair proficient cells. This defect was complemented by transfection of the CSB gene into the UV61 cells. We thus conclude that efficient PCNA complex formation after UV is dependent upon both the NER and TCR pathways in hamster cells. The association of several other DNA repair proteins including XPA, RPA, TFIIH and p53 with the insoluble PCNA complex in UV treated cells suggests a central role for PCNA in different steps of NER.
Balajee
A S
AS
Laboratory of Molecular Genetics, National Institute on Aging, NIH, Baltimore, MD 21224, USA.
May
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A
Dianova
I
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Bohr
V A
VA
D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D006224:Cricetinae; D004260:DNA Repair / Q000235:genetics; D019084:Fluorescent Antibody Technique, Indirect; D016678:Genome; D006801:Humans; D015530:Nuclear Matrix / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D011485:Protein Binding; D011740:Pyrimidine Dimers / Q000378:metabolism; D014158:Transcription, Genetic; D014466:Ultraviolet Rays
D018809:Proliferating Cell Nuclear Antigen; D011740:Pyrimidine Dimers