Interactions between Src homology (SH) 2/SH3 adapter proteins and the guanylnucleotide exchange factor SOS are differentially regulated by insulin and epidermal growth factor.
Co-immunoprecipitation of whole cell extracts demonstrated that the guanylnucleotide exchange factor SOS was associated with the small adapter proteins Grb2, CrkII, and Nck. In vitro binding indicated a similar binding affinity of SOS for all three adapter proteins but with a slightly lower Kd for Grb2 (approximately 2.5-fold) compared with Nck and CrkII. Insulin stimulation resulted in co-immunoprecipitation of tyrosine-phosphorylated IRS1 with Grb2 and to a lesser extent CrkII. Although Grb2 also associated with tyrosine-phosphorylated Shc, there was no detectable interaction of CrkII with Shc. In contrast, EGF stimulation resulted in the predominant co-immunoprecipitation of Grb2 with the EGF receptor, whereas CrkII primarily associated with an unidentified 120-130-kDa protein. Similar to the ability of insulin to induce the dissociation of the Grb2-SOS complex, there was a concomitant time-dependent dissociation of the CrkII-SOS and Nck-SOS complexes. However, EGF stimulation had no effect on the association state of the Grb2-SOS or the Nck-SOS complexes but did result in a time-dependent dissociation of the CrkII from SOS. Together, these data demonstrate that different cellular pools of SOS associate with different adapter proteins forming various signaling complexes, each undergoing distinct patterns of assembly/disassembly following growth factor stimulation.
Okada
S
S
Department of Physiology and Biophysics, University of Iowa, Iowa City, Iowa 52242, USA.
Pessin
J E
JE
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D066246:ErbB Receptors / Q000096:biosynthesis / Q000502:physiology*; D051380:GRB2 Adaptor Protein; D020662:Guanine Nucleotide Exchange Factors; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D015513:Oncogene Proteins / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D050737:Proto-Oncogene Proteins c-crk; D011972:Receptor, Insulin / Q000096:biosynthesis / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; D020662:Guanine Nucleotide Exchange Factors; D007328:Insulin; C078332:Nck protein; D015513:Oncogene Proteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; D050737:Proto-Oncogene Proteins c-crk; D011993:Recombinant Fusion Proteins; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin