Dissociation of CAK from core TFIIH reveals a functional link between XP-G/CS and the TFIIH disassembly state

• Published in: PloS one Vol. 5; no. 6; p. e11007
• Main Authors: Arab, Hany H, Wani, Gulzar, Ray, Alo, Shah, Zubair I, Zhu, Qianzheng, Wani, Altaf A
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 08-06-2010; Public Library of Science (PLoS)
• Subjects: Biochemistry/ and Repair; Cancer; Cell cycle; Cell Line; Chromatin; Chromatin Immunoprecipitation; Cyclin-dependent kinases; Cyclin-Dependent Kinases - metabolism; Damage; Degradation; Deoxyribonucleic acid; Dismantling; Dissociation; DNA; DNA Damage; DNA repair; DNA-Binding Proteins - metabolism; DNA-directed RNA polymerase; Endonucleases - metabolism; Genomes; Hypotheses; Immunoassay; Immunoprecipitation; Inhibition; Kinases; Molecular Biology/DNA Repair; Molecular Biology/Transcription Initiation and Activation; Mutation; Nuclear Proteins - metabolism; Nucleotide excision repair; p53 Protein; Phosphorylation; Proteins; Repair; Ribonucleic acid; RNA; RNA polymerase; RNA polymerase II; Transcription (Genetics); Transcription Factor TFIIH - metabolism; Transcription Factors - metabolism; Tumor proteins; Ultraviolet radiation; Ultraviolet Rays; XPD protein; United States > US; Ohio
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0011007

Transcription factor II H (TFIIH) is comprised of core TFIIH and Cdk-activating kinase (CAK) complexes. Here, we investigated the molecular and cellular manifestation of the TFIIH compositional changes by XPG truncation mutations. We showed that both core TFIIH and CAK are rapidly recruited to damage sites in repair-proficient cells. Chromatin immunoprecipitation against TFIIH and CAK components revealed a physical engagement of CAK in nucleotide excision repair (NER). While XPD recruitment to DNA damage was normal, CAK was not recruited in severe XP-G and XP-G/CS cells, indicating that the associations of CAK and XPD to core TFIIH are differentially affected. A CAK inhibition approach showed that CAK activity is not required for assembling pre-incision machinery in vivo or for removing genomic photolesions. Instead, CAK is involved in Ser5-phosphorylation and UV-induced degradation of RNA polymerase II. The CAK inhibition impaired transcription from undamaged and UV-damaged reporter, and partially decreased transcription of p53-dependent genes. The overall results demonstrated that a) XP-G/CS mutations affect the disassembly state of TFIIH resulting in the dissociation of CAK, but not XPD from core TFIIH, and b) CAK activity is not essential for global genomic repair but involved in general transcription and damage-induced RNA polymerase II degradation.