Redox priming of the insulin receptor ß-chain associated with altered tyrosine kinase activity and insulin responsiveness in the absence of tyrosine autophosphorylation

^a Deutsches Krebsforschungszentrum, Division of Immunochemistry, D-69120 Heidelberg, Germany
^b INSERM, U294 et Labo Immuno-Hemato, 75018 Paris, France

Induction of tyrosine kinase activity of the insulin receptor (IR) ß-chain is believed to require its autophosphorylation at Tyr¹¹⁶², Tyr¹¹⁶³, and Tyr¹¹⁵⁸. However, the mechanism of the initial phosphorylation is poorly understood. We show that treatment of IR-transfected Chinese hamster ovary cells with antioxidants inhibits insulin responsiveness. Conversely, partial inhibition of glutathione biosynthesis by buthionine sulfoximine (BSO) and glutathione reductase by 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU), i.e., procedures that intracellularly induce mildly oxidative conditions, caused a decrease in IR ß-chain sulfhydryl groups and enhanced synergistically the induction of IR tyrosine phosphorylation by insulin. The IR ß-chain from cells treated with BSO/BCNU in the absence of insulin was not detectably tyrosine phosphorylated, but nevertheless was functionally altered, as demonstrated in vitro by a moderate kinase activity at low ATP concentrations (5 nM) and a strong kinase activity at 25 µM ATP. This activity was found to be specific for tyrosine (not for serine or threonine), and tryptic peptide maps indicated that it is more selective than that induced by insulin. Moreover, the kinase activity from BSO/BCNU-treated cells showed a spontaneous decay that was not prevented by the phosphatase inhibitor vanadate. Together, these results suggest that optimal insulin responsiveness may require a process of `redox priming' of the IR ß-chain that involves structural and functional changes in the absence of detectable tyrosine phosphorylation of the ß-chain.—Schmid, E., El Benna, J., Galter, D., Klein, G., Dröge, W. Redox priming of the insulin receptor ß-chain associated with altered tyrosine kinase activity and insulin responsiveness in the absence of tyrosine autophosphorylation. FASEB J. 12, 863–870 (1998)

Key Words: redox regulation • signal transduction • glutathione • CHO-HIR • BCNU

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