Neoplastic transformation induced by insulin receptor substrate-1 overexpression requires an interaction with both Grb2 and Syp signaling molecules.
The insulin receptor substrate-1 (IRS-1) is the major intracellular substrate of insulin and insulin-like growth factor-I (IGF-I) receptor tyrosine kinase activity, and this protein has been found to be overexpressed in human hepatocellular carcinomas. IRS-1 contains several src homology 2 (SH2) binding motifs that interact following tyrosyl phosphorylation with SH2-containing proteins, and this interaction may be essential for transmitting the growth signal from the cell surface to the nucleus. We have previously reported that overexpression of IRS-1 may induce neoplastic transformation of NIH 3T3 cells. This study examines the role of two SH2-containing molecules, namely the Grb2 adapter and Syp tyrosine phosphatase proteins as important components of the cellular transforming activity of IRS-1. Mutations of tyrosine 897 in the YVNI motif (Y897F) and of tyrosine 1180 in the YIDL motif (Y1180F) reduced the intracellular interaction of IRS-1 with Grb2 and Syp proteins, respectively. Furthermore, a single mutation at either Phe-897 or Phe-1180 substantially but not completely reduced IGF-I-dependent transforming activity of IRS-1, whereas creation of a double mutation of both tyrosine residues (Y897F/Y1180F) strikingly attenuated the transforming activity of IRS-1. Stable expression of the IRS-1 mutant constructs in NIH 3T3 cells was associated with a lower level of activation of the mitogen-activated protein kinase kinase (MAPKK)/MAPK cascade following IGF-I stimulation compared with cells stably transfected with the "wild-type" IRS-1 gene. These results suggest that IRS-1-induced cellular transformation requires an interaction with both Grb2 and Syp signal transduction molecules since neither interaction alone appears to be required, and this event subsequently leads to activation of the MAPKK/MAPK cascade.
Tanaka
S
S
Molecular Hepatology Laboratory, Massachusetts General Hospital Cancer Center, Massachusetts General Hospital and Harvard Medical School, Charlestown, Massachusetts 02129, USA.
Ito
T
T
Wands
J R
JR
D016475:3T3 Cells; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D006528:Carcinoma, Hepatocellular; D002448:Cell Adhesion / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002471:Cell Transformation, Neoplastic; D004789:Enzyme Activation; D066246:ErbB Receptors / Q000096:biosynthesis; D051380:GRB2 Adaptor Protein; D015870:Gene Expression; D006801:Humans; D055504:Insulin Receptor Substrate Proteins; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D047908:Intracellular Signaling Peptides and Proteins; D008113:Liver Neoplasms; D051379:Mice; D020929:Mitogen-Activated Protein Kinase Kinases; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D010750:Phosphoproteins / Q000096:biosynthesis*; D011494:Protein Kinases / Q000378:metabolism; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; D015398:Signal Transduction; D014162:Transfection; D014407:Tumor Cells, Cultured; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C495949:Grb2 protein, mouse; C526292:IRS1 protein, human; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; C526295:Irs1 protein, mouse; D009838:Oligodeoxyribonucleotides; D010750:Phosphoproteins; D011506:Proteins; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D011494:Protein Kinases; D066246:ErbB Receptors; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C516724:PTPN11 protein, human; C509249:PTPN6 protein, human; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C516725:Ptpn11 protein, mouse; C509248:Ptpn6 protein, mouse; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases