DIFFERENTIAL ETOPOSIDE SENSITIVITY OF CELLS DEFICIENT IN THE KU AND DNA-PKCS COMPONENTS OF THE DNA-DEPENDENT PROTEIN-KINASE

Etoposides block cell division by interfering with the action of topoi somerase II, leaving enzyme-DNA double-strand breaks. We found that ce rtain components of the trimeric DNA-dependent protein kinase influenc e cell survival following etoposide damage. Interestingly, either Ku70 - or Ku80-deficient cell lines, but not mutant cell lines of the DNA-P K catalytic sub-unit (DNA-PKcs), were found to be hypersensitive to th e effects of etoposide VP16, Ku70- and Ku80-deficient cells can be com plemented to an etoposide resistant phenotype by introducing wildtype Ku70 or Ku80 cDNAs, Mutational analysis of introduced Ku70 cDNAs into murine embryonic stem cells deleted for Ku70 (-/-) showed that mutants where heterodimerization and DNA binding functions of Ku were disrupt ed, also blocked the restoration of etoposide resistance. In contrast with the differential etoposide sensitivity of DNA-PK mutants, both Ku - and DNA-PKcs-deficient cell lines showed G(2) ionizing radiation-ind uced delays, a cell cycle phase where topoisomerase II function is cri tical. Thus, the topoisomerase II cleaved complexes may be an example of DNA lesions requiring the Ku heterodimer, but not DNA-PK for DNA re pair.