Role of IRS-1-GRB-2 complexes in insulin signaling.
GRB-2 is a small SH2- and SH3 domain-containing adapter protein that associates with the mammalian SOS homolog to regulate p21ras during growth factor signaling. During insulin stimulation, GRB-2 binds to the phosphorylated Y895VNI motif of IRS-1. Substitution of Tyr-895 with phenylalanine (IRS-1F-895) prevented the IRS-1-GRB-2 association in vivo and in vitro. The myeloid progenitor cell line, 32-D, is insensitive to insulin because it contains few insulin receptors and no IRS-1. Coexpression of IRS-1 or IRS-1F-895 with the insulin receptor was required for insulin-stimulated mitogenesis in 32-D cells, while expression of the insulin receptor alone was sufficient to mediate insulin-stimulated tyrosine phosphorylation of Shc and activation of p21ras and mitogen-activated protein (MAP) kinase. The Shc-GRB-2 complex formed during insulin stimulation is a possible mediator of p21ras and MAP kinase activation in IRS-1-deficient 32-D cells. Interestingly, IRS-1, but not IRS-1F-895, enhanced the stimulation of MAP kinase by insulin in 32-D cells expressing insulin receptors. Thus, IRS-1 contributes to the stimulation of MAP kinase by insulin, probably through formation of the IRS-1-GRB-2 complex at Tyr-895. Our results suggest that the Shc-GRB-2 complex and the activation of p21ras-dependent signaling pathways, including MAP kinase, are insufficient for insulin-stimulated mitogenesis and that the essential function(s) of IRS-1 in proliferative signaling is largely unrelated to IRS-1-GRB-2 complex formation.
Myers
M G
MG
Research Division, Joslin Diabetes Center, Boston, Massachusetts 02215.
Wang
L M
LM
Sun
X J
XJ
Zhang
Y
Y
Yenush
L
L
Schlessinger
J
J
Pierce
J H
JH
White
M F
MF
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002477:Cells; D006224:Cricetinae; D017931:DNA Primers; D066246:ErbB Receptors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D051380:GRB2 Adaptor Protein; D015870:Gene Expression; D006412:Hematopoietic Stem Cells; D007328:Insulin / Q000494:pharmacology*; D055504:Insulin Receptor Substrate Proteins; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013014:SOS Response, Genetics; D015398:Signal Transduction
D048868:Adaptor Proteins, Signal Transducing; D017931:DNA Primers; D051380:GRB2 Adaptor Protein; C495950:Grb2 protein, rat; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; C526296:Irs1 protein, rat; D010750:Phosphoproteins; D011506:Proteins; D011993:Recombinant Fusion Proteins; D066246:ErbB Receptors; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D016283:Proto-Oncogene Proteins p21(ras)