Activation of Glycogen Synthase by Insulin in 3T3-L1 Adipocytes Involves c-Cbl-associating Protein (CAP)-dependent and CAP-independent Signaling Pathways

Christian A Baumann,Matthew J Brady,Alan R Saltiel

doi:10.1074/jbc.c000856200

Abstract

In adipose and muscle, insulin stimulates glucose uptake and glycogen synthase activity. Phosphatidylinositol 3-kinase (PI3K) activation is necessary but not sufficient for these metabolic actions of insulin. The insulin-stimulated translocation of phospho-c-Cbl to lipid rafts, via its association with CAP, comprises a second pathway regulating GLUT4 translocation. In 3T3-L1 adipocytes, overexpression of a dominant negative CAP mutant (CAP Delta SH3) completely blocked the insulin-stimulated glucose transport and glycogen synthesis but only partially inhibited glycogen synthase activation. In contrast, CAP Delta SH3 expression did not affect glycogen synthase activation by insulin in the absence of extracellular glucose. Moreover, CAP Delta SH3 has no effect on the PI3K-dependent activation of protein phosphatase-1 or phosphorylation of glycogen synthase kinase-3. These results indicate blockade of the c-Cbl/CAP pathway directly inhibits insulin-stimulated glucose uptake, which results in secondary inhibition of glycogen synthase activation and glycogen synthesis.

Highlights

Activation of Glycogen Synthase by Insulin in 3T3-L1 Adipocytes Involves c-Cbl-associating Protein (CAP)-dependent and CAP-independent Signaling Pathways*
These results indicate blockade of the c-Cbl/CAP pathway directly inhibits insulin-stimulated glucose uptake, which results in secondary inhibition of glycogen synthase activation and glycogen synthesis
CAP⌬SH3 Expression Blocks Glucose Uptake and Glycogen Synthesis in 3T3-L1 Adipocytes—CAP⌬SH3 overexpression in 3T3-L1 adipocytes significantly inhibits the translocation of the insulin-stimulated glucose transporter GLUT4 to the plasma membrane, without affecting Phosphatidylinositol 3-kinase (PI3K)-dependent signals [19]

Summary

Accelerated Publication

CAP⌬SH3 has no effect on the PI3K-dependent activation of protein phosphatase-1 or phosphorylation of glycogen synthase kinase-3 These results indicate blockade of the c-Cbl/CAP pathway directly inhibits insulin-stimulated glucose uptake, which results in secondary inhibition of glycogen synthase activation and glycogen synthesis. In 3T3-L1 adipocytes, expression of a CAP mutant in which the SH3 domains have been deleted (CAP⌬SH3) disrupted the formation of a CAP1⁄7flotillin1⁄7Cbl complex in lipid rafts, blocked translocation of the insulin-stimulated glucose transporter GLUT4, and inhibited glucose transport, glycogen, and lipid synthesis [19] It remained unclear whether CAP⌬SH3 directly blocked the signaling pathways involving GS dephosphorylation that are independent of glucose transport. We demonstrate here that the CAP/Cbl pathway regulates GS activation exclusively through a glucosedependent mechanism

EXPERIMENTAL PROCEDURES

CAP Regulation of Glycogen Synthase Activation

RESULTS

DISCUSSION