Vascular smooth muscle insulin resistance, but not hypertrophic signaling, is independent of angiotensin II-induced IRS-1 phosphorylation by JNK.

Angiotensin II (ANG II) has been implicated in the pathogenesis of diabetic micro- and macrovascular disease. In vascular smooth muscle cells (VSMCs), ANG II phosphorylates and degrades insulin receptor substrate-1 (IRS-1). While the pathway responsible for IRS-1 degradation in this system is unknown, c-Jun NH(2)-terminal kinase (JNK) has been linked with serine phosphorylation of IRS-1 and insulin resistance. We investigated the role of JNK in ANG II-induced IRS-1 phosphorylation, degradation, Akt activation, glucose uptake, and hypertrophic signaling, focusing on three IRS-1 phosphorylation sites: Ser302, Ser307, and Ser632. Maximal IRS-1 phosphorylation on Ser632 occurred at 5 min, on Ser307 at 30 min, and on Ser302 at 60 min. The JNK inhibitor SP600125 reduced ANG II-induced IRS-1 Ser307 phosphorylation (by 80%), IRS-1 Ser302 phosphorylation (by 70%), and IRS-1 Ser632 phosphorylation (by 50%). However, JNK inhibition had no effect on ANG II-mediated IRS-1 degradation, nor did it reverse the ANG II-induced decrease in Akt phosphorylation or glucose uptake. Transfection of VSMCs with mutants S307A, S302A, or S632A of IRS-1 did not block ANG II-mediated IRS-1 degradation. In contrast, JNK inhibition attenuated insulin-induced upregulation of collagen and smooth muscle α-actin in ANG II-pretreated cells. We conclude that phosphorylation of Ser307, Ser302, and Ser632 of IRS-1 is not involved in ANG II-mediated IRS-1 degradation, and that JNK alone does not mediate ANG II-stimulated IRS-1 degradation, but rather is responsible for the hypertrophic effects of insulin on smooth muscle.