Changes in tyrosine phosphorylation of insulin receptors and a 170,000 molecular weight nonreceptor protein in vivo in skeletal muscle of streptozotocin-induced diabetic rats: effects of insulin and glucose.
The effects of experimental diabetes on in vivo tyrosine phosphorylation of the insulin receptor (IR) and non-receptor proteins were investigated in rat skeletal muscle. Diabetes was induced in male Sprague-Dawley rats (200 g) by streptozotocin administration (100 mg/kg, ip). Diabetic animals were subsequently anesthetized, insulin was injected via cardiac puncture, and hindlimb skeletal muscles were removed, frozen in liquid N2, and homogenized in sodium dodecyl sulfate. Tyrosine phosphoproteins were first immunoprecipitated and then identified by immunoblotting with antiphosphotyrosine antibodies. In both control and diabetic rats, insulin stimulated tyrosine phosphorylation of the IR beta-subunit and a major nonreceptor 170,000 mol wt (Mr) endogenous protein (pp170) in a dose- and time-dependent manner. Total IR number (determined by immunoprecipitation and immunoblotting with an anti-IR antibody) increased 2.4-fold in diabetic muscle, but there was little change in phosphorylated insulin receptor beta-subunit (157 +/- 12% of control value; P less than 0.001). In contrast, pp170 phosphorylation increased markedly in diabetes (500 +/- 119% of control value; P less than 0.005), and the time course of its disappearance was delayed compared to that in control rats. These changes were reversed by insulin therapy (5 U, sc, twice daily), but not by correction of hyperglycemia with phlorizin (0.4 g/kg.day, sc). In conclusion, in rat skeletal muscle in vivo, streptozotocin-diabetes results in 1) increased total IR number, 2) reduced efficiency of IR phosphorylation, and 3) markedly enhanced tyrosine phosphorylation of a 170,000 Mr putative IR substrate. Hypoinsulinemia, but not hyperglycemia, appears to increase the level of the phophorylated 170,000 Mr protein in streptozotocin-diabetes.
Giorgino
F
F
Research Division, Joslin Diabetes Center, Boston, Massachusetts 02215.
Chen
J H
JH
Smith
R J
RJ
D000818:Animals; D001786:Blood Glucose / Q000378:metabolism / Q000502:physiology; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005947:Glucose / Q000494:pharmacology*; D015151:Immunoblotting; D007328:Insulin / Q000494:pharmacology*; D008297:Male; D008970:Molecular Weight; D009132:Muscles / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D010766:Phosphorylation / Q000187:drug effects; D011233:Precipitin Tests; D051381:Rats; D011919:Rats, Inbred Strains; D011972:Receptor, Insulin / Q000032:analysis / Q000378:metabolism*; D013311:Streptozocin; D013997:Time Factors; D014443:Tyrosine / Q000378:metabolism*
D001786:Blood Glucose; D007328:Insulin; D014443:Tyrosine; D013311:Streptozocin; D011972:Receptor, Insulin; D005947:Glucose