Overexpression of Muscle Glycogen Phosphorylase in Cultured Human Muscle Fibers Causes Increased Glucose Consumption and Nonoxidative Disposal

Abstract

The effect of increased expression of glycogen phosphorylase on glucose metabolism in human muscle was examined in primary cultured fibers transduced with recombinant adenovirus AdCMV-MGP encoding muscle glycogen phosphorylase. Increments of 20-fold in total enzyme activity and of 14-fold of the active form of the enzyme were associated with a 30% reduction in basal glycogen levels. Total glycogen synthase activity was doubled in AdCMV-MGP-transduced cells even though the activity ratio was decreased. Incubation with forskolin, which inactivated glycogen synthase and activated glycogen phosphorylase, induced greater net glycogenolysis in engineered cells. In unstimulated fibers, lactate production was three times higher in AdCMV-MGP fibers as compared with controls, despite similar rates of glycogenolysis. In transduced fibers incubated with 2-deoxyglucose, the level of 2-deoxyglucose 6-phosphate was about 8-fold elevated over the control even though hexokinase activity was unmodified in AdCMV-MGP fibers. Overexpression of glycogen phosphorylase also led to enhancement of [U- CO 2 formation from [6-14 C]glucose was unmodified, in AdCMV-MGP fibers. Our data show that in human skeletal muscle cells in culture, the increase in glycogen phosphorylase activity is able to up-regulate glycogen synthase activity indicating the enhancement of glycogen turnover. We suggest that the increase in glycogen phosphorylase and, thereby, in glycogen metabolism, is sufficient to enhance glucose uptake in the muscle cell. Glucose taken up by engineered muscle cells is essentially disposed of through nonoxidative metabolism and converted into lactate and lipid.

Partly Ordered Synthesis and Degradation of Glycogen in Cultured Rat Myotubes

“…Glycogen degradation was followed for 4 h and initiated by a medium containing 1 mM glucose and 10 M forskolin, which is supposed to activate glycogen phosphorylase (15). Analogous to the synthesis, the degradation process could be well approximated by the monoexponential Equation 2 (see "Calculations") with a rate constant k deg of 1.39 Ϯ 0.14 h Ϫ1 , Glyc span-deg ϭ 0.913 Ϯ 0.016, Glyc t3ϱ ϭ 0.095 Ϯ 0.018, and Glyc max ϭ 1.008 Ϯ 0.011 (Table I).…”

Biochemical and physiological function of stearoyl-CoA desaturase

“…Increased glycogen phosphorylase activity would have been expected to promote glycogenolysis. However, overexpression of glycogen phosphorylase in cultured human skeletal muscle cells increased the level of glycogen synthase (2), which suggests that the amount of glycogen synthase and phosphorylase is controlled in a reciprocal manner by glycogen levels. Thus, the increase in both glycogen synthase and phosphorylase activities would also imply that there is increased glycogen metabolism in the SCD-1 Ϫ/Ϫ mice.…”

“…SCD-1 contains four transmembrane domains with both the NH 2 and COOH termini oriented toward the cytosol. The single cytoplasmic loop and the COOH terminus contain the eight histidine (His) residues known to form the His box, which binds iron within the catalytic center of the desaturase (Fig.…”

Shared control of hepatic glycogen synthesis by glycogen synthase and glucokinase

“…For example, the adenovirus-mediated overexpression of rabbit muscle glycogen phosphorylase in cultured human muscle fibres resulted in an increased level of total GS [19]. Transgenic mice that overexpress muscle GS also showed increased levels of glycogen phosphorylase in muscle fibres [3,20].…”