Insulin rapidly stimulates tyrosine phosphorylation of a Mr-185,000 protein in intact cells

Abstract

Phosphotyrosine-containing proteins are minor components of normal cells which appear to be associated primarily with the regulation of cellular metabolism and growth. The insulin receptor is a tyrosine-specific protein kinase, and one of the earliest detectable responses to insulin binding is activation of this kinase and autophosphorylation of its beta-subunit. Tyrosine autophosphorylation activates the phosphotransferase in the beta-subunit and increases its reactivity toward tyrosine phosphorylation of other substrates. When incubated in vitro with [gamma-32P]ATP and insulin, the purified insulin receptor phosphorylates various proteins on their tyrosine residues. However, so far no proteins other than the insulin receptor have been identified as undergoing tyrosine phosphorylation in response to insulin in an intact cell. Here, using anti-phosphotyrosine antibodies, we have identified a novel phosphotyrosine-containing protein of relative molecular mass (Mr) 185,000 (pp185) which appears during the initial response of hepatoma cells to insulin binding. In contrast to the insulin receptor, pp185 does not adhere to wheat-germ agglutininagarose or bind to anti-insulin receptor antibodies. Phosphorylation of pp185 is maximal within seconds after exposure of the cells to insulin and exhibits a dose-response curve similar to that of receptor autophosphorylation, suggesting that this protein represents the endogenous substrate for the insulin receptor kinase.

Crk protein binds to PDGF receptor and insulin receptor substrate-1 with different modulating effects on PDGF- and insulin-dependent signaling pathways

“…The autophosphorylation of the IR functions predominantly to up-regulate the activity of the tyrosine kinase catalytic domain (Herrera and Rosen, 1986). The IR phosphorylates the widely expressed cytosolic protein IRS-1, a major target for both the IR and the insulin-like growth factor-1 (IGF1) receptor (Myers et al, 1994a;White et al, 1985).…”

Insulin receptor substrates mediate distinct biological responses to insulin-like growth factor receptor activation in breast cancer cells

“…Though six IRS family members have been identified (IRS-1 to IRS-6) (White et al, 1985;Tobe et al, 1995;Lavan et al, 1997a, b;Cai et al, 2003), IRS-1 and -2 are the predominant signalling molecules utilised by the IGF-IR to mediate IGF-I action in breast cancer cells (Jackson et al, 1998(Jackson et al, , 2001. Indeed, human breast tumours express both IRS-1 and -2 (Jackson et al, 1998;Lee et al, 1999).…”

Defects in insulin signal transduction in liver and muscle of pregnant rats

“…This interaction catalyses the intramolecular autophosphorylation of specific tyrosine residues of the b subunit which further enhances the tyrosine kinase activity of the receptor toward other protein substrates [8]. In most cells, this primary event leads to the subsequent tyrosyl phosphorylation of a cytoplasmic protein with an apparent molecular weight of 160-185 kDa, called insulin receptor substrate 1 (IRS-1) [9][10][11]. Considerable evidence indicates that insulin receptor tyrosine Diabetologia (1997) 40: 179-186 Defects in insulin signal transduction in liver and muscle of pregnant rats Summary Pregnancy is known to induce insulin resistance, but the exact molecular mechanism involved is unknown.…”