RACK1 regulates Ki-Ras-mediated signaling and morphological transformation of NIH 3T3 cells

Activating Ras mutations are involved in a significant fraction of human tumors. A suppressor screen using a retroviral mouse fibroblast cDNA library was performed to identify novel factors in Ras-mediated transformation. We identified a novel potent inhibitor of Ras-mediated morphological transformation encoded by a truncated version of the receptor for activated C-kinase (RACK1). The truncated protein, designated RACK1DeltaWD1, lacked the N-terminal 49 amino acids encoding the first of the 7 WD40 repeats in RACK1. RACK1DeltaWD1 expression restored contact inhibition, stress fiber formation and reduced ERK phosphorylation in Ki-Ras transformed NIH 3T3 cells. We demonstrate that truncated RACK1 is involved in complexes consisting of wild-type RACK1 and protein kinase C isoforms alpha, betaI and delta, compromising the transduction of an activated Ras signal to the Raf-MEK-ERK pathway. The cellular localization of RACK1DeltaWD1 differed from wtRACK1, indicating that signaling complexes containing the truncated version of RACK1 are incorrectly localized. Notably, 12-O-tetradecanoyl-13-phorbol acetate (TPA) mediated intracellular translocation of RACK1-interacting PKC alpha and delta was abrogated in RACK1DeltaWD1-expressing cells. Our data support a model where RACK1 acts as a key factor in Ki-Ras-mediated morphological transformation.