Insulin-Stimulated Phosphorylation of the Akt Substrate AS160 Is Impaired in Skeletal Muscle of Type 2 Diabetic Subjects

Insulin-Stimulated Phosphorylation of the Akt Substrate AS160 Is Impaired in Skeletal Muscle of Type 2 Diabetic Subjects Håkan K.R. Karlsson 1 , Juleen R. Zierath 1 , Susan Kane 2 , Anna Krook 3 , Gustav E. Lienhard 2 and Harriet Wallberg-Henriksson 3 1 Department of Surgical Sciences, Integrative Physiology Section, Karolinska Institutet, Stockholm, Sweden 2 Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 3 Department of Physiology and Pharmacology, Integrative Physiology Section, Karolinska Institutet, Stockholm, Sweden Address correspondencereprint requests to Juleen R. Zierath, PhD, Karolinska Institutet, Department of Surgical Sciences, Integrative Physiology Section, S-171 77, Stockholm, Sweden. E-mail: juleen.zierath{at}fyfa.ki.se Abstract AS160 is a newly described substrate for the protein kinase Akt that links insulin signaling and GLUT4 trafficking. In this study, we determined the expression of and in vivo insulin action on AS160 in human skeletal muscle. In addition, we compared the effect of physiological hyperinsulinemia on AS160 phosphorylation in 10 lean−to−moderately obese type 2 diabetic and 9 healthy subjects. Insulin infusion increased the phosphorylation of several proteins reacting with a phospho-Akt substrate antibody. We focused on AS160, as this Akt substrate has been linked to glucose transport. A 160-kDa phosphorylated protein was identified as AS160 by immunoblot analysis with an AS160-specific antibody. Physiological hyperinsulinemia increased AS160 phosphorylation 2.9-fold in skeletal muscle of control subjects ( P < 0.001). Insulin-stimulated AS160 phosphorylation was reduced 39% ( P < 0.05) in type 2 diabetic patients. AS160 protein expression was similar in type 2 diabetic and control subjects. Impaired AS160 phosphorylation was related to aberrant Akt signaling; insulin action on Akt Ser 473 phosphorylation was not significantly reduced in type 2 diabetic compared with control subjects, whereas Thr 308 phosphorylation was impaired 51% ( P < 0.05). In conclusion, physiological hyperinsulinemia increases AS160 phosphorylation in human skeletal muscle. Moreover, defects in insulin action on AS160 may impair GLUT4 trafficking in type 2 diabetes. GAP, GTPase-activating protein GSK-3α, glycogen synthase kinase 3α IRS, insulin receptor substrate 1 mTOR, mammalian target of rapamycin PAS, anti-phospho-(Ser/Thr) Akt substrate PI, phosphatidylinositol TBST, Tris-buffered saline with 0.02% Tween Footnote