A point mutation in the UDP-glucose pyrophosphorylase gene results in decreases of UDP-glucose and inactivation of glycogen synthase.
The regulatory role of UDP-glucose in glycogen biogenesis was investigated in fibroblasts containing a point mutation in the UDP-glucose pyrophosphorylase gene and, consequently, chronically low UDP-glucose levels (Qc). Comparisons were made with cells having the intact gene and restored UDP-glucose levels (G3). Glycogen was always very low in Qc cells. [(14)C]Glucose incorporation into glycogen was decreased and unaffected by insulin in Qc cells, whereas insulin stimulated glucose incorporation by approximately 50% in G3 cells. Glycogen synthase (GS) activity measured in vitro was virtually absent and the amount of enzyme in Qc cells was decreased by about 50%. The difference in GS activity between cells persisted even when G3 cells were devoid of glycogen. Incubation of G3 cell extracts with either exogenous UDP-glucose or glycogen resulted in increases in GS activity. Incubation of Qc cell extracts with exogenous UDP-glucose had no effect on GS activity; however, incubation with glycogen fully restored enzyme activity. Incubation of G3 cell extracts with radioactive UDP-glucose resulted in substantial binding of ligand to immunoprecipitated GS, whereas no binding was detected in Qc immunoprecipitates. Incubation of Qc cell extracts with exogenous glycogen fully restored UDP-glucose binding in the immunoprecipitate. These data suggest that chronically low UDP-glucose levels in cells result in inactivation of GS, owing to loss of the ability of GS to bind UDP-glucose.
Higuita
Juan-Carlos
JC
Microbiology and Tumor Biology Center, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Alape-Girón
Alberto
A
Thelestam
Monica
M
Katz
Abram
A
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D006224:Cricetinae; D004789:Enzyme Activation; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D006003:Glycogen / Q000378:metabolism; D006006:Glycogen Synthase / Q000378:metabolism*; D008168:Lung / Q000166:cytology; D017354:Point Mutation; D011485:Protein Binding; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase / Q000235:genetics / Q000378:metabolism*; D014532:Uridine Diphosphate Glucose / Q000737:chemistry / Q000378:metabolism*
D006003:Glycogen; D006006:Glycogen Synthase; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase; D014532:Uridine Diphosphate Glucose