OVEREXPRESSION OF GLYCOGEN-PHOSPHORYLASE INCREASES GLUT4 EXPRESSION AND GLUCOSE-TRANSPORT IN CULTURED SKELETAL HUMAN MUSCLE

Skeletal muscle glucose utilization, a major factor in the control of whole-body glucose tolerance, is modulated in accordance with the musc le metabolic demand. For instance, it is increased in chronic contract ion or exercise training in association with elevated expression of GL UT4 and hexokinase II (HK-II). In this work, the contribution of incre ased metabolic flux to the regulation of the glucose transport capacit y was analyzed in cultured human skeletal muscle engineered to overexp ress glycogen phosphorylase (GP). Myocytes treated with an adenovirus- bearing muscle GP cDNA (AdCMV-MGP) expressed 10 times higher GP activi ty and exhibited a twofold increase in the V-max for 2-deoxy-D-[H-3]gl ucose (2-DG) uptake, with no effect on the apparent K-m. The stimulato ry effect of insulin on 2-DG uptake was also markedly enhanced in AdCM V-MGP-treated cells, which showed maximal insulin stimulation 2.8 time s higher than control cells. No changes in HKII total activity or the intracellular compartmentalization were found. GLUT4, protein, and mRN A were raised in AdCMV-MGP-treated cells, suggesting pretranslational activation. GLUT4 was immunodetected intracellularly with a perinuclea r predominance. Culture in glucose-free or high-glucose medium did not alter GLUT4 protein content in either control cells or AdCMV-MGP-trea ted cells. Control and GP-overexpressing cells showed similar autoinhi bition of glucose transport, although they appeared to differ in the m echanism(s) involved in this effect. Whereas GLUT1 protein increased i n control cells when they were switched from a high-glucose to a gluco se-free medium, GLUT1 remained unaltered in GP-expressing cells upon g lucose deprivation. Therefore, the increased intracellular metabolic ( glycogenolytic-glycolytie) flux that occurs in muscle cells overexpres sing GP causes an increase in GLUT4 expression and enhances basal and insulin-stimulated glucose transport, without significant changes in t he autoinhibition of glucose transport. This mechanism of regulation m ay be operative in the postexercise situation in which GLUT4 expressio n is upregulated in coordination with increased glycolytic flux and en ergy demand.