Insulin resistance and muscle insulin receptor substrate-1 serine hyperphosphorylation

Abstract

Insulin resistance in metabolic syndrome subjects is profound in spite of muscle insulin receptor and insulin-responsive glucose transporter (GLUT4) expression being nearly normal. Insulin receptor tyrosine kinase phosphorylation of insulin receptor substrate-1 (IRS-1) at Tyr896 is a necessary step in insulin stimulation of translocation of GLUT4 to the cell surface. Serine phosphorylation of IRS-1 by some kinases diminishes insulin action in mice. We evaluated the phosphorylation status of muscle IRS-1 in 33 subjects with the metabolic syndrome and seventeen lean controls. Each underwent euglycemic insulin clamps and a thigh muscle biopsy before and after 8 weeks of either strength or endurance training. Muscle IRS-1 phosphorylation at six sites was quantified by immunoblots. Metabolic syndrome muscle IRS-1 had excess phosphorylation at Ser337 and Ser636 but not at Ser307, Ser789, or Ser1101. Ser337 is a target for phosphorylation by glycogen synthase kinase 3 (GSK3) and Ser636 is phosphorylated by c-Jun N-terminal kinase 1 (JNK1). Exercise training without weight loss did not change the IRS-1 serine phosphorylation. These data suggest that baseline hyperphosphorylation of at least two key serines within muscle IRS-1 diminishes the transmission of the insulin signal and thereby decreases the insulin-stimulated translocation of GLUT4. Excess fasting phosphorylation of muscle IRS-1 at Ser636 may be a major cause of the insulin resistance seen in obesity and might prevent improvement in insulin responsiveness when exercise training is not accompanied by weight loss.

Keywords: Insulin receptor substrate‐1; insulin resistance; metabolic syndrome; muscle.

© 2014 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

Figures

Figure 1.

IRS‐1 content of muscle from metabolic syndrome subjects. (Panel A) shows sample immunoblots for each of the antibodies used for quantifying total IRS‐1 and phosphorylation at six specific sites. Each blot shown here had lanes containing samples from metabolic syndrome subjects alternating with samples from controls. The band on the left of each image shown is a metabolic syndrome subject. Panel B summarizes the data for each antibody. All of the data are expressed in relation to the mean of the untrained sedentary control subjects for each assay. The data displayed are the mean and standard errors from at least two separate assays for each subject. The number of subjects that had complete data in each assay was variable due to sample limitations. The data shown in Panel B represent 10–12 controls and 14–18 metabolic syndrome subjects. The asterisk indicates significant difference (P < 0.05, t test) from the control subject data for that assay.

Figure 2.

The effect of exercise training on muscle IRS‐1 phosphorylation at Ser337 and Ser636. Pre‐ and postexercise training data are displayed in this figure. Eight weeks of training, either progressive resistance or a stationary bike program, did not significantly alter the level of baseline phosphorylation of Ser337 or Ser636 as quantified by western blots. Shown here are the mean and standard errors of results of at least two separate experiments including pre‐ and posttraining muscle specimens from 12 controls and 18 metabolic syndrome subjects. The asterisk denotes significant difference (P < 0.05) from the corresponding control data. There was no significant increase or decrease in phosphorylation of Ser337 or Ser636 after exercise training in controls or metabolic syndrome subjects.

Figure 3.

Correlation between insulin responsiveness and baseline phosphorylation at IRS‐1 serine 636. The data shown are the euglycemic clamp steady‐state glucose infusion rate (SSGIR) plotted against the relative amount of phosphorylation at IRS‐1 serine 636. The data shown are before and/or after training for 11 controls (open symbols) and 20 metabolic syndrome subjects (filled circles). The missing subjects had inadequate muscle left for this assay to be done. The correlation coefficient (R) and p value, calculated using Pearson Product Moment, are shown on the graph.