GLUT-4 phosphorylation and its intrinsic activity. Mechanism of Ca(2+)-induced inhibition of insulin-stimulated glucose transport

• Published in: The Journal of biological chemistry Vol. 268; no. 5; pp. 3352 - 3356
• Main Authors: Begum, N, Leitner, W, Reusch, J E, Sussman, K E, Draznin, B
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 15.02.1993
• Subjects: Adenosine Triphosphate - metabolism; Adenosine Triphosphate - pharmacology; Adipose Tissue - drug effects; Adipose Tissue - metabolism; Animals; Biological Transport - drug effects; Calcium - metabolism; Calcium-Transporting ATPases - antagonists & inhibitors; Cell Membrane - drug effects; Cell Membrane - metabolism; Cells, Cultured; Cyclic AMP - analogs & derivatives; Cyclic AMP - metabolism; Cyclic AMP - pharmacology; Deoxyglucose - metabolism; Glucose - metabolism; Insulin - pharmacology; Kinetics; Male; Monosaccharide Transport Proteins - metabolism; Nitrendipine - pharmacology; Phosphates - metabolism; Phosphorus Radioisotopes; Phosphorylation; Rats; Rats, Sprague-Dawley; Terpenes - pharmacology; Thapsigargin; Thionucleotides - pharmacology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)53701-7

In this study, we examined the influence of high levels of cytosolic calcium on phosphorylation status and function of GLUT-4 in isolated rat adipocytes. Intracellular calcium was elevated by exposing adipocytes to either extracellular ATP (1.6 mM) or thapsigargin (100 nM). Both agents increased cytosolic calcium 2-3 fold. While basal glucose uptake was unaffected, both ATP and thapsigargin reduced insulin-stimulated glucose transport by 40-70% (p < 0.05). Neither ATP nor thapsigargin affected GLUT-4 content or its translocation from the low density microsomes to the plasma membrane (PM). In contrast, GLUT-4 immunoprecipitated from the PM of adipocytes exposed to either ATP or thapsigargin was phosphorylated to a greater extent than the GLUT-4 isolated from control cells. ATP and thapsigargin also abolished insulin-stimulated dephosphorylation of GLUT-4. At the same time, GLUT-4 intrinsic activity was significantly reduced in adipocytes with high levels of cytosolic calcium (p < 0.05). Preincubation of adipocytes with cAMP antagonist, RpcAMP (10(-4) M), and calcium channel blocker, nitrendipine (30 microM), improved the ability of insulin to dephosphorylate GLUT-4 and restored insulin-stimulated GLUT-4 intrinsic activity. We conclude that elevated levels of cytosolic calcium interfere with insulin's ability to dephosphorylate GLUT-4, thus reducing its intrinsic activity.