Insulin-induced desensitization of extracellular signal-regulated kinase activation results from all inhibition of Raf activity independent of Ras activation and dissociation of the Grb2-SOS complex

Previous studies have suggested that the interaction between the small adap tor protein Grb2 with the Ras guanyl nucleotide exchange factor SOS is func tionally important in the regulation of the Ras activation/inactivation cyc le. To examine the relationship between the Grb2-SOS complex and Ras activa tion, we observed that insulin stimulation results in a rapid but transient activation of Ras and the extracellular-signal regulated kinase (ERK) foll owed by dissociation of the Grb2-SOS complex. Although treatment with the p horbol myristate acetate resulted in ERK activation and complete dissociati on of the Grb2-SOS complex, there was no effect on subsequent insulin-stimu lated Ras activation. Similarly, insulin stimulation followed by insulin re moval resulted in a time-dependent restoration of the Grb2-SOS complex but which was significantly slower than the recovery of insulin-stimulated Ras activation. In addition, although insulin was able to activate Ras under th ese conditions, there was a complete desensitization of Raf and ERK activat ion. This apparent homologous desensitization of insulin action was specifi c for Raf and ERK as the insulin re-stimulation of insulin receptor autopho sphorylation and protein kinase B activation were unaffected. Together, the se data demonstrate the presence of a pathway independent of the Grb2-SOS c omplex that can lead to Ras activation but that the desensitization of Raf accounts for the homologous desensitization of ERK.