A MODEL FOR KU HETERODIMER ASSEMBLY AND INTERACTION WITH DNA - IMPLICATIONS FOR THE FUNCTION OF KU ANTIGEN

Ku autoantigen, a heterodimer of 70- and 80-kDa subunits, is a DNA end -binding factor critical for DNA repair. Two domains of p70 mediate DN A binding, one on the C-terminal and one on the N-terminal portion. Th e latter must dimerize with p80 in order to bind DNA, whereas the form er is p80-independent. Both must be intact for end binding activity in gel shift assays. To evaluate the role of p80 in DNA binding, deletio n mutants were co-expressed with full-length p70 using recombinant bac uloviruses. We show by several criteria that amino acids 371-510 of p8 0 interact with p70. Both of the p70 dimerization domains bind to the same region of p80, but apparently to separate sites within that regio n. In DNA immunoprecipitation assays, amino acids 179-510 of p80 were required for p80-dependent DNA binding of p70, whereas in gel shift as says, amino acids 179-732 were necessary. Interestingly, both the p80- dependent and the p80-independent DNA binding sites preferentially bou nd to DNA ends, suggesting a model in which a single Ku heterodimer ma y juxtapose two broken DNA ends physically, facilitating their rejoini ng by DNA ligases.