CLONING OF AN INTRACELLULAR RECEPTOR FOR PROTEIN-KINASE-C - A HOMOLOGOF THE BETA-SUBUNIT OF G-PROTEINS

Protein kinase C (PKC) translocates from the soluble to the cell parti culate fraction on activation. Intracellular receptors that bind activ ated PKC in the particulate fraction have been implicated by a number of studies. Previous work identified 30- to 36-kDa proteins in the par ticulate fraction of heart and brain that bound activated PKC in a spe cific and saturable manner. These proteins were termed receptors for a ctivated C-kinase, or RACKs. In the following study, we describe the c loning of a cDNA encoding a 36-kDa protein (RACK1) that fulfills the c riteria for RACKs. (i) RACK1 bound PKC in the presence of PKC activato rs, but not in their absence. (ii) PKC binding to the recombinant RACK 1 was not inhibited by a pseudosubstrate peptide or by a substrate pep tide derived from the pseudosubstrate sequence, indicating that the bi nding did not reflect simply PKC association with its substrate. (iii) Binding of PKC to RACK1 was saturable and specific; two other protein kinases did not bind to RACK1. (iv) RACK1 contains two short sequence s homologous to a PKC binding sequence previously identified in annexi n I and in the brain PKC inhibitor KCIP. Peptides derived from these s equences inhibited PKC binding to RACK1. Finally, RACK1 is a homolog o f the beta subunit of G proteins, which were recently implicated in me mbrane anchorage of the beta-adrenergic receptor kinase [Pitcher, J., Inglese, L., Higgins, J. B., Arriza, J. A., Casey, P. J., Kim, C., Ben ovic, J. L., Kwatra, M. M., Caron, M. G. & Lefkowitz, R. J. (1992) Sci ence 257, 1264-12671. Our in vitro data suggest a role for RACK1 in PK C-mediated signaling.