Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1996 Dec 15;320(Pt 3):871–877. doi: 10.1042/bj3200871

Insulin action in cultured human myoblasts: contribution of different signalling pathways to regulation of glycogen synthesis.

S J Hurel 1, J J Rochford 1, A C Borthwick 1, A M Wells 1, J R Vandenheede 1, D M Turnbull 1, S J Yeaman 1
PMCID: PMC1218009  PMID: 9003374

Abstract

A key metabolic action of insulin is the stimulation of non-oxidative glucose utilization in skeletal muscle, by increasing both glucose uptake and glycogen synthesis. The molecular mechanism underlying this process has been investigated using a variety of experimental systems. We report here the use of cultured human myoblasts to study insulin control of glycogen synthesis in humans. In these cells insulin stimulates glycogen synthesis approx. 2.2-fold, associated with a similar activation of glycogen synthase (GS) which occurs within 5-10 min of the addition of insulin. Insulin also causes inactivation of glycogen synthase kinase-3 (GSK-3) and activation of protein kinase B, both processes being sufficiently rapid to account for the effects of insulin on GS. Activation by insulin of the protein kinases p70s6K, p90s6K and extracellular signal-regulated kinase 2 (ERK2) is observed, but is significantly slower than the activation of GS. Selective inhibitors of the p70s6K pathway (rapamycin), the ERK2/p90s6K pathway (PD98059) and phosphatidylinositol 3-kinase (wortmannin) have been used to probe the contribution of these components to insulin signalling in human muscle. Wortmannin blocks activation of both glycogen synthesis and GS and inactivation of GSK-3. PD98059 is without effect on these events, while rapamycin is without effect on inactivation of GSK-3 but partially blocks activation of glycogen synthesis and GS. Taken together, these findings suggest that protein kinase B is responsible for the inactivation of GSK-3, but that an additional rapamycin-sensitive mechanism may contribute to the activation of GS and stimulation of glycogen synthesis.

Full Text

The Full Text of this article is available as a PDF (717.5 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arcaro A., Wymann M. P. Wortmannin is a potent phosphatidylinositol 3-kinase inhibitor: the role of phosphatidylinositol 3,4,5-trisphosphate in neutrophil responses. Biochem J. 1993 Dec 1;296(Pt 2):297–301. doi: 10.1042/bj2960297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Azpiazu I., Saltiel A. R., DePaoli-Roach A. A., Lawrence J. C. Regulation of both glycogen synthase and PHAS-I by insulin in rat skeletal muscle involves mitogen-activated protein kinase-independent and rapamycin-sensitive pathways. J Biol Chem. 1996 Mar 1;271(9):5033–5039. doi: 10.1074/jbc.271.9.5033. [DOI] [PubMed] [Google Scholar]
  3. Barriocanal L. A., Borthwick A. C., Stewart M., Wells A., Hurel S. J., Yeaman S. J., Taylor R. The effect of acute (60 minute) insulin stimulation upon human skeletal muscle glycogen synthase and protein phosphatase-1 in non-insulin-dependent diabetic patients and control subjects. Diabet Med. 1995 Dec;12(12):1110–1115. doi: 10.1111/j.1464-5491.1995.tb00429.x. [DOI] [PubMed] [Google Scholar]
  4. Begum N. Stimulation of protein phosphatase-1 activity by insulin in rat adipocytes. Evaluation of the role of mitogen-activated protein kinase pathway. J Biol Chem. 1995 Jan 13;270(2):709–714. doi: 10.1074/jbc.270.2.709. [DOI] [PubMed] [Google Scholar]
  5. Borthwick A. C., Wells A. M., Rochford J. J., Hurel S. J., Turnbull D. M., Yeaman S. J. Inhibition of glycogen synthase kinase-3 by insulin in cultured human skeletal muscle myoblasts. Biochem Biophys Res Commun. 1995 May 25;210(3):738–745. doi: 10.1006/bbrc.1995.1721. [DOI] [PubMed] [Google Scholar]
  6. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  7. Cheatham B., Kahn C. R. Insulin action and the insulin signaling network. Endocr Rev. 1995 Apr;16(2):117–142. doi: 10.1210/edrv-16-2-117. [DOI] [PubMed] [Google Scholar]
  8. Cheng H. C., Kemp B. E., Pearson R. B., Smith A. J., Misconi L., Van Patten S. M., Walsh D. A. A potent synthetic peptide inhibitor of the cAMP-dependent protein kinase. J Biol Chem. 1986 Jan 25;261(3):989–992. [PubMed] [Google Scholar]
  9. Chung J., Kuo C. J., Crabtree G. R., Blenis J. Rapamycin-FKBP specifically blocks growth-dependent activation of and signaling by the 70 kd S6 protein kinases. Cell. 1992 Jun 26;69(7):1227–1236. doi: 10.1016/0092-8674(92)90643-q. [DOI] [PubMed] [Google Scholar]
  10. Cohen P. Dissection of the protein phosphorylation cascades involved in insulin and growth factor action. Biochem Soc Trans. 1993 Aug;21(3):555–567. doi: 10.1042/bst0210555. [DOI] [PubMed] [Google Scholar]
  11. Cross D. A., Alessi D. R., Cohen P., Andjelkovich M., Hemmings B. A. Inhibition of glycogen synthase kinase-3 by insulin mediated by protein kinase B. Nature. 1995 Dec 21;378(6559):785–789. doi: 10.1038/378785a0. [DOI] [PubMed] [Google Scholar]
  12. Cross D. A., Alessi D. R., Vandenheede J. R., McDowell H. E., Hundal H. S., Cohen P. The inhibition of glycogen synthase kinase-3 by insulin or insulin-like growth factor 1 in the rat skeletal muscle cell line L6 is blocked by wortmannin, but not by rapamycin: evidence that wortmannin blocks activation of the mitogen-activated protein kinase pathway in L6 cells between Ras and Raf. Biochem J. 1994 Oct 1;303(Pt 1):21–26. doi: 10.1042/bj3030021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dent P., Lavoinne A., Nakielny S., Caudwell F. B., Watt P., Cohen P. The molecular mechanism by which insulin stimulates glycogen synthesis in mammalian skeletal muscle. Nature. 1990 Nov 22;348(6299):302–308. doi: 10.1038/348302a0. [DOI] [PubMed] [Google Scholar]
  14. Denton R. M., Tavaré J. M. Does mitogen-activated-protein kinase have a role in insulin action? The cases for and against. Eur J Biochem. 1995 Feb 1;227(3):597–611. doi: 10.1111/j.1432-1033.1995.tb20179.x. [DOI] [PubMed] [Google Scholar]
  15. Dudley D. T., Pang L., Decker S. J., Bridges A. J., Saltiel A. R. A synthetic inhibitor of the mitogen-activated protein kinase cascade. Proc Natl Acad Sci U S A. 1995 Aug 15;92(17):7686–7689. doi: 10.1073/pnas.92.17.7686. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Guinovart J. J., Salavert A., Massagué J., Ciudad C. J., Salsas E., Itarte E. Glycogen synthase: a new activity ratio assay expressing a high sensitivity to the phosphorylation state. FEBS Lett. 1979 Oct 15;106(2):284–288. doi: 10.1016/0014-5793(79)80515-3. [DOI] [PubMed] [Google Scholar]
  17. Henry R. R., Abrams L., Nikoulina S., Ciaraldi T. P. Insulin action and glucose metabolism in nondiabetic control and NIDDM subjects. Comparison using human skeletal muscle cell cultures. Diabetes. 1995 Aug;44(8):936–946. doi: 10.2337/diab.44.8.936. [DOI] [PubMed] [Google Scholar]
  18. Häring H. U., Mehnert H. Pathogenesis of type 2 (non-insulin-dependent) diabetes mellitus: candidates for a signal transmitter defect causing insulin resistance of the skeletal muscle. Diabetologia. 1993 Mar;36(3):176–182. doi: 10.1007/BF00399946. [DOI] [PubMed] [Google Scholar]
  19. Kahn C. R. Banting Lecture. Insulin action, diabetogenes, and the cause of type II diabetes. Diabetes. 1994 Aug;43(8):1066–1084. doi: 10.2337/diab.43.8.1066. [DOI] [PubMed] [Google Scholar]
  20. Lawrence J. C., Jr, Zhang J. N. Control of glycogen synthase and phosphorylase by amylin in rat skeletal muscle. Hormonal effects on the phosphorylation of phosphorylase and on the distribution of phosphate in the synthase subunit. J Biol Chem. 1994 Apr 15;269(15):11595–11600. [PubMed] [Google Scholar]
  21. Lazar D. F., Wiese R. J., Brady M. J., Mastick C. C., Waters S. B., Yamauchi K., Pessin J. E., Cuatrecasas P., Saltiel A. R. Mitogen-activated protein kinase kinase inhibition does not block the stimulation of glucose utilization by insulin. J Biol Chem. 1995 Sep 1;270(35):20801–20807. doi: 10.1074/jbc.270.35.20801. [DOI] [PubMed] [Google Scholar]
  22. Lin T. A., Lawrence J. C., Jr Activation of ribosomal protein S6 kinases does not increase glycogen synthesis or glucose transport in rat adipocytes. J Biol Chem. 1994 Aug 19;269(33):21255–21261. [PubMed] [Google Scholar]
  23. Moule S. K., Edgell N. J., Welsh G. I., Diggle T. A., Foulstone E. J., Heesom K. J., Proud C. G., Denton R. M. Multiple signalling pathways involved in the stimulation of fatty acid and glycogen synthesis by insulin in rat epididymal fat cells. Biochem J. 1995 Oct 15;311(Pt 2):595–601. doi: 10.1042/bj3110595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Parker P. J., Caudwell F. B., Cohen P. Glycogen synthase from rabbit skeletal muscle; effect of insulin on the state of phosphorylation of the seven phosphoserine residues in vivo. Eur J Biochem. 1983 Jan 17;130(1):227–234. doi: 10.1111/j.1432-1033.1983.tb07140.x. [DOI] [PubMed] [Google Scholar]
  25. Sakaue H., Hara K., Noguchi T., Matozaki T., Kotani K., Ogawa W., Yonezawa K., Waterfield M. D., Kasuga M. Ras-independent and wortmannin-sensitive activation of glycogen synthase by insulin in Chinese hamster ovary cells. J Biol Chem. 1995 May 12;270(19):11304–11309. doi: 10.1074/jbc.270.19.11304. [DOI] [PubMed] [Google Scholar]
  26. Sarabia V., Lam L., Burdett E., Leiter L. A., Klip A. Glucose transport in human skeletal muscle cells in culture. Stimulation by insulin and metformin. J Clin Invest. 1992 Oct;90(4):1386–1395. doi: 10.1172/JCI116005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Sarabia V., Ramlal T., Klip A. Glucose uptake in human and animal muscle cells in culture. Biochem Cell Biol. 1990 Feb;68(2):536–542. doi: 10.1139/o90-076. [DOI] [PubMed] [Google Scholar]
  28. Seger R., Krebs E. G. The MAPK signaling cascade. FASEB J. 1995 Jun;9(9):726–735. [PubMed] [Google Scholar]
  29. Shepherd P. R., Navé B. T., Siddle K. Insulin stimulation of glycogen synthesis and glycogen synthase activity is blocked by wortmannin and rapamycin in 3T3-L1 adipocytes: evidence for the involvement of phosphoinositide 3-kinase and p70 ribosomal protein-S6 kinase. Biochem J. 1995 Jan 1;305(Pt 1):25–28. doi: 10.1042/bj3050025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Shulman G. I., Rothman D. L., Jue T., Stein P., DeFronzo R. A., Shulman R. G. Quantitation of muscle glycogen synthesis in normal subjects and subjects with non-insulin-dependent diabetes by 13C nuclear magnetic resonance spectroscopy. N Engl J Med. 1990 Jan 25;322(4):223–228. doi: 10.1056/NEJM199001253220403. [DOI] [PubMed] [Google Scholar]
  31. Sutherland C., Cohen P. The alpha-isoform of glycogen synthase kinase-3 from rabbit skeletal muscle is inactivated by p70 S6 kinase or MAP kinase-activated protein kinase-1 in vitro. FEBS Lett. 1994 Jan 24;338(1):37–42. doi: 10.1016/0014-5793(94)80112-6. [DOI] [PubMed] [Google Scholar]
  32. Welsh G. I., Foulstone E. J., Young S. W., Tavaré J. M., Proud C. G. Wortmannin inhibits the effects of insulin and serum on the activities of glycogen synthase kinase-3 and mitogen-activated protein kinase. Biochem J. 1994 Oct 1;303(Pt 1):15–20. doi: 10.1042/bj3030015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Welsh G. I., Proud C. G. Glycogen synthase kinase-3 is rapidly inactivated in response to insulin and phosphorylates eukaryotic initiation factor eIF-2B. Biochem J. 1993 Sep 15;294(Pt 3):625–629. doi: 10.1042/bj2940625. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES