CHARACTERIZATION OF THE RNA-BINDING PROPERTIES OF KU PROTEIN

Ku protein, a heterodimer of 70 and 83 kDa polypeptides, is the regula tory component of the DNA-dependent protein kinase (DNA-PK). Ku protei n binds to DNA ends and is essential for DNA double-strand break repai r and V(D)J recombination. Although there is some evidence that Ku pro tein also binds RNA, its RNA binding properties have not been systemat ically explored. In the present study, Ku-binding RNAs were identified using systematic evolution of ligands by exponential enrichment (SELE X) technology. These RNAs were assigned to three classes based on comm on sequence motifs. Most of the selected RNAs bound to Ku protein with a K-d less than or equal to 2 nM, comparable to the affinity of DNA f ragments for Ku protein under similar conditions. Many of the RNAs inh ibited DNA-PK activity by competing with DNA for a common binding site in Ku protein. None of several RNAs that were tested activated DNA-PK in the absence of DNA. The identification of diverse RNAs that bind a vidly to Ku protein is consistent with the idea that natural RNAs may serve as modulators of DNA-PK activity. Moreover, the RNAs identified in this study may have utility as tools for experimental manipulation of DNA double-strand break repair activity in cells and cell extracts.