RESTORATION OF X-RAY AND ETOPOSIDE RESISTANCE, KU-END BINDING-ACTIVITY AND V(D)J RECOMBINATION TO THE CHINESE-HAMSTER SXI-3 MUTANT BY A HAMSTER KU86 CDNA

Ku is a heterodimeric protein composed of 86 and 70 kDa subunits that binds preferentially to the double-stranded ends of DNA. Recent molecu lar characterization of ionizing-radiation sensitive (IR(s)) mutants b elonging to the XRCC5 complementation group demonstrated the involveme nt of Ku in DNA double-strand break (DSB) repair and lymphoid V(D)J re combination. Here, we describe the isolation of a full-length hamster cDNA encoding the large subunit of the Ku heterodimer and demonstrate that the stable expression of this cDNA can functionally restore IR(r) . Ku DNA end-binding activity and V(D)J recombination proficiency in t he Chinese hamster IR(s) sxi-3 mutant. Moreover, we also demonstrate t hat sxi-3 cells are hypersensitive to etoposide, a DNA topoisomerase I I inhibitor. and that resistance to this drug was restored by the Ku86 cDNA. These experiments suggest that a defect in the large subunit of the heterodimeric Ku protein is the sole factor responsible for the k nown defects of sxi-3 cells and our data further support the role of K u in DNA DSB repair and V(D)J recombination.