Date of Award

1985

Embargo Period

8-1-2024

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Molecular and Cellular Biology and Pathobiology

College

College of Graduate Studies

First Advisor

Maria G. Buse

Second Advisor

Rosalie K. Crouch

Third Advisor

George E. Lindenmayer

Abstract

The role of the insulin receptor in muscle insulin resistance was studied. Denervation of skeletal muscle resulted in the rapid appearance of a decrease in basal and insulin stimulated glucose transport, glycogen synthesis and a defect in the activation of glycogen synthase which was independent of levels of glycogen in the muscle. The effect was seen in both predominantly red and predominantly white muscle. There was no change in the denervated muscles' sensitivity to epinephrine-mediated decrease in glycogen synthesis or glycogen content, nor was there a major change in the insulin binding capacity of denervated muscles. A technique was developed to investigate the properties of the insulin receptor solubilized from skeletal muscle. Freezing hindlimb muscles with subsequent powdering and homogenization with buffered Triton X-100 produced a fraction with specific insulin binding. Further purification on agarose-bound wheat germ agglutinin yielded insulin receptors with intact insulin binding, as well as insulin stimulatable autophosphorylation and exogenous substrate kinase activity. The autophosphorylation of the insulin receptor β-subunit in intact tissues was also demonstrated. The binding and kinase domains of insulin receptors solubilized from denervated skeletal muscle were identical to that from contralateral controls. No significant changes were seen in the autophosphorylation of the β-subunit in intact, denervated solei. Injection of insulin in the rat and subsequent partial purification of insulin receptors in the presence of phosphatase inhibitors allowed the demonstration of an increased kinase activity towards histone H2b as compared to saline treated rats. The increase was due to an increase in tyrosine phosphorylation attributed to the activation of the insulin receptor kinase. Again, no change in the insulin receptors from denervated muscles was observed. Streptozotocin diabetes resulted in an increase in specific insulin binding but a decrease in insulin-stimulatable autophosphorylation and tyrosine kinase activity. Structural analysis showed no change in the β-subunit but the β-subunit attained a component with a slower migration on electrophoresis. The β-subunit change was shown to be, in part, due to increased sialic acid content of the receptor subunit. The kinetic and structural changes were reversible with insulin treatment of the rat.

Rights

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