THE PROTOONCOGENE PRODUCT C-CRK ASSOCIATES WITH INSULIN-RECEPTOR SUBSTRATE-1 AND 4PS - MODULATION BY INSULIN GROWTH-FACTOR-I (IGF) AND ENHANCED IGF-1 SIGNALING

The Crk proto-oncogene product is an SH2 and SH3 domain-containing ada ptor protein which we have previously shown to become rapidly tyrosine phosphorylated in response to stimulation with insulin-like growth fa ctor I (IGF-I) in NIH-3T3 cells, In order to further characterize the role of Crk in the IGF-I signaling pathway, NIH-3T3 and 293 cells were stably transfected with an expression vector containing the Crk cDNA, The various resultant 3T3-Crk clones expressed Crk at approximately 2 -15-fold higher levels than parental 3T3 cells, In 3T3-Crk cells, Crk immunoreactivity was detected in insulin receptor substrate-1 (IRS-1) immunoprecipitates, Stimulation with IGF-I resulted in a dissociation of Crk protein from IRS-1, In contrast, the association of the related adaptor protein Grb2 with IRS-1 was enhanced by IGF-I stimulation. Si milar results were obtained in stably transfected 293-Crk cells, which express both IRS-I and the IRS-1-related signaling protein 4PS. In th ese cells, IRS-1 and 4PS both associated with Crk, and this associatio n was also decreased by IGF-I treatment, whereas the association of Gr b2 with IRS-1 and 4PS was enhanced by IGF-I. Overexpression of Crk als o enhanced IGF-I-induced mitogenesis of NIH-3T3 cells, as measured by [H-3]thymidine incorporation. The levels of IGF-I-induced mitogenesis were proportional to the level of Crk expression, These results sugges t that Crk is a positive effector of IGF-I signaling, and may mediate its effects via interaction with IRS-1 and/or 4PS.