title: UV light-induced degradation of RNA polymerase II is dependent on the Cockayne's syndrome A and B proteins but not p53 or MLH1.
abstract: It has been hypothesized that the degradation of the largest subunit of RNA polymerase II (polIILS) is required for transcription-coupled repair (TCR) of UV light-induced transcription-blocking lesions. In this study we further investigated the mechanism of UV-induced degradation of polIILS using cell lines with specific defects in TCR or in the recovery of RNA synthesis. It was found that the hypophosphorylated IIa form of polIILS rapidly decreased following UV-irradiation in all cell lines tested. Inhibition of proteasome activity resulted in an increase of the hyperphosphorylated IIo form of polIILS in UV-irradiated cells, while inhibition of CTD-kinases resulted in the retention of the IIa form. In UV-irradiated Cockayne's syndrome cells, which are defective in TCR, the levels of the IIo form increased in a similar manner as when proteasome inhibitors were added to UV-irradiated normal cells. In contrast, TCR-deficient HCT116 cells, which lack the mismatch repair protein MLH1, showed proficient degradation of polIILS as did cells with deficiencies in the recovery of RNA synthesis following UV-irradiation due to defective p53. Furthermore, we found that proteasome function was important for the recovery of mRNA synthesis even in TCR-deficient HCT116 cells. Our results suggest that proteasome-mediated degradation of polIILS is preceded by phosphorylation of the C-terminal domain of polIILS and requires the CS-A and CS-B but not MLH1 or p53 proteins. Furthermore, our results suggest that following UV-irradiation, the degradation of polIILS is required for the efficient recovery of mRNA synthesis but not for TCR per se.
authors.affiliation: Department of Radiology, Centre for Cancer Therapeutics, Ottawa Regional Cancer Centre, University of Ottawa, Ont., K1H 8L6, Ottawa, Canada.
authors.forename: B C
authors.initials: BC
authors.lastname: McKay
authors.forename: F
authors.initials: F
authors.lastname: Chen
authors.forename: S T
authors.initials: ST
authors.lastname: Clarke
authors.forename: H E
authors.initials: HE
authors.lastname: Wiggin
authors.forename: L M
authors.initials: LM
authors.lastname: Harley
authors.forename: M
authors.initials: M
authors.lastname: Ljungman
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002352:Carrier Proteins; D002460:Cell Line; D006224:Cricetinae; D003546:Cysteine Endopeptidases; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair / Q000528:radiation effects; D045643:DNA Repair Enzymes; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000528:radiation effects; D005455:Fluorescent Antibody Technique; D006801:Humans; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors; D000070957:MutL Protein Homolog 1; D009363:Neoplasm Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins; D010447:Peptide Hydrolases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects / Q000528:radiation effects; D000075223:Poly-ADP-Ribose Binding Proteins; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D012313:RNA / Q000096:biosynthesis; D012319:RNA Polymerase II / Q000378:metabolism / Q000528:radiation effects*; D014157:Transcription Factors; D014158:Transcription, Genetic / Q000528:radiation effects; D016159:Tumor Suppressor Protein p53 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D014466:Ultraviolet Rays
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C095472:ERCC8 protein, human; D004791:Enzyme Inhibitors; C501486:MLH1 protein, human; D009097:Multienzyme Complexes; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D000075223:Poly-ADP-Ribose Binding Proteins; D011506:Proteins; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D012313:RNA; D012319:RNA Polymerase II; C410949:RNA polymerase II largest subunit; D010447:Peptide Hydrolases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; D000070957:MutL Protein Homolog 1; D004265:DNA Helicases; C078673:ERCC6 protein, human; D045643:DNA Repair Enzymes