P95(VAV) ASSOCIATES WITH THE NUCLEAR-PROTEIN KU-70

The proto-oncogene vav is expressed solely in hematopoietic cells and plays an important role in cell signaling, although little is known ab out the proteins involved in these pathways. To gain further informati on, the Src homology 2 (SH2) and 3 (SH3) domains of Vav were used to s creen a lymphoid cell cDNA library by the yeast two-hybrid system. Amo ng the positive clones, we detected a nuclear protein, Ku-70, which is the DNA-binding element of the DNA-dependent protein kinase. In Jurka t and UT7 cells, Vav is partially localized in the nuclei, as judged f rom immunofluorescence and confocal microscopy studies. By using gluta thione S-transferase fusion proteins derived from Ku-70 and coimmunopr ecipitation experiments with lysates prepared from human thymocytes an d Jurkat and UT7 cells, we show that Vav associates with Ku-70. The in teraction of Vav with Ku-70 requires only the 150-residue carboxy-term inal portion of Ku-70, which binds to the 25 carboxy-terminal residues of the carboxy SH3 domain of Vav. A proline-to-leucine mutation in th e carboxy SH3 of Vav that blocks interaction with proline-rich sequenc es does not modify the binding of Ku-70, which lacks this motif. There fore, the interaction of Vav with Ku-70 may be a novel form of protein -protein interaction. The potential role of Vav/Ku-70 complexes is dis cussed.