Endogenous Substrates of the Insulin Receptor: Studies with Cells Expressing Wild-Type and Mutant Receptors
The insulin receptor has an intrinsic tyrosine kinase activity which appears to be required for insulin to elicit its various biological responses (1). Identification of endogenous substrates of the insulin receptor kinase and other tyrosine kinases has been greatly advanced in the last few years by the development of high affinity polyclonal and monoclonal antibodies to phosphotyrosine (2–4). These reagents can be used to identify substrates of tyrosine kinases by immunoblotting and immunoprecipitation as well as to purify substrates by utilizing affinity columns composed of anti-phosphotyrosine antibodies. Via these and other techniques, numerous proteins have been identified as becoming phosphorylated on tyrosine residues. These include a number of proteins with unknown functions. One such protein, Mr ~160,000 to 180,000, has been reported to be tyrosine phosphorylated in response to both insulin and insulin-like growth factor I in a variety of cell types (5–6). Other proteins that have been identified as substrates of tyrosine kinases include various enzymes as well as other defined molecules. One such identified molecule is another tyrosine kinase, called pp60C-SRC. Activation of the platelet derived growth factor (PDGF) receptor tyrosine kinase was shown to increase the extent of tyrosine phosphorylation of the SRC protein and increase its enzymatic activity (7, 8). More recently, a phospholipase C (9–11), the type I phosphatidylinositol kinase (12–13), the GTPase activating protein of Ras (called GAP) (14, 15)and several serine/threonine kinases (the MAP2 kinase, the proto-oncogene Raf kinase and a cell cycle dependent kinase, CDC-2) (16–18) have all been shown to be tyrosine phosphorylated. The role of the tyrosine phosphorylation of these different molecules is not always clear. Only the physiological role of the tyrosine phosphorylation of the yeast cell cycle dependent kinase has been directly demonstrated (19).
KeywordsInsulin Receptor Tyrosine Phosphorylation Chinese Hamster Ovary Cell Thymidine Incorporation Mutant Receptor
Unable to display preview. Download preview PDF.
- 7.R. Ralston and J. M. Bishop, The product of the protooncogene c-src is modified during the cellular response to platelet-derived growth factor, Biochemistry 82:7845 (1985).Google Scholar
- 23.K. Yonezawa, G. Endemann, K.S. Kovacina, J.E. Chin, C. Stover and R.A. Roth, Substrates of the insulin receptor kinase, in: “The Biology and Medicine of Signal Transduction,” Y. Nishizuka et al., Publisher, New York (1990).Google Scholar
- 28.G. Steele-Perkins and R.A. Roth, Monoclonal antibody aIR-3 inhibits the ability of insulin-like growth factor II to stimulate a signal from the type I receptor without inhibiting its binding, Biochem. Biophvs. Res. Comm., in press.Google Scholar
- 35.J. Avruch, H.E. Tornqvist, J.R. Gunsalus, E.J. Yurkow, Insulin regulation of protein phosphorylation, im “Insulin,” P. Cuatrecasas and S. Jacobs, ed., Springer-Verlag Berlin Heidelberg (1990).Google Scholar
- 38.K. Yonezawa and R.A. Roth, Assessment of the in situ tyrosine kinase activity of mutant insulin receptors lacking tyrosine autophosphorylation sites 1162 and 1163, Mol. Endocrinol., Submitted (1990).Google Scholar
- 39.C. Reynet, M. Caron, J. Magre, G. Cherqui, E. Clauser, J. Picard and J. Capeau, Mutation of tyrosine residues 1162 and 1163 of the insulin receptor affects hormone and receptor internalization, Mol. Endocrinol., 304 (1990).Google Scholar