Characterization of Grb4, an adapter protein interacting with Bcr-Abl

We report here the characterization of an adapter protein Identified in a y east a-hybrid screen with the use of Bcr-Abl as the bait. Grb4 bound to Bcr -Abl in a variety of systems, both in vitro and in vivo, and is an excellen t substrate of the Bcr-Abl tyrosine kinase, The association of Grb4 and Bcr -Abl in intact cells was mediated by an are homology (SH)2-mediated phospho tyrosine-dependent interaction as well as an SHE-mediated phosphotyrosine-i ndependent interaction. Grb4 has 68% homology to the adapter protein Nck an d has similar but distinct binding specificities in K562 lysates. Subcellul ar localization studies indicate that Grb4 localizes to both the nucleus an d the cytoplasm. Coexpression of kinase-active Bcr-Abl with Grb4 resulted i n the translocation of Grb4 from the cytoplasm and the nucleus to the cytos keleton to colocalize with Bcr-Abl. In addition, expression of Grb4 with ki nase-active Bcr-Abl resulted in a redistribution of actin-associated Bcr-Ab l, Finally, coexpression of Grb4 and oncogenic v-Abl strongly inhibited v-A bl-induced AP-1 activation. Together, these data indicate that Grb4 In conj unction with Bcr-Abl may be capable of modulating the cytoskeletal structur e and negatively interfering with the signaling of oncogenic Abl kinases, G rb4 may therefore play a role in the molecular pathogenesis of chronic myel ogenous leukemia. (C) 2000 by The American Society of Hematology.