UV sensitivity and impaired nucleotide excision repair in DNA-dependent protein kinase mutant cells

Abstract

DNA-dependent protein kinase (DNA-PK), a member of the phosphatidyl-inositol (PI) 3-kinase family, is involved in the repair of DNA double-strand breaks. Its regulatory subunit, Ku, binds to DNA and recruits the kinase catalytic subunit (DNA-PK(CS)). We show here a new role of DNA-PK in the modulation of the process of nucleotide excision repair (NER) in vivo since, as compared with their respective parental cell lines, DNA-PK mutants (scid, V-3 and xrs 6 cells) exhibit sensitivity to UV-C irradiation (2.0- to 2.5-fold) and cisplatin (~ 3- to 4-fold) associated with a decreased activity (40-55%) of unscheduled DNA synthesis after UV-C irradiation. Moreover, we observed that wortmannin sensitized parental cells in vivo when combined with either cisplatin or UV-C light, but had no effect on the DNA-PK(CS) deficient scid cells. Despite a lower repair synthesis activity (~ 2-fold) measured in vitro with nuclear cell extracts from DNA-PK mutants, a direct involvement of DNA-PK in the NER reaction in vitro has not been observed. This study establishes a regulatory function of DNA-PK in the NER process in vivo but rules out a physical role of the complex in the repair machinery at the site of the DNA lesion.