Early insulin signaling cascade in a model of oxidative skeletal muscle: mouse Sol8 cell line.
Cell models provide important tools to investigate the mechanisms modulating the insulin-signaling cascade. Insulin interaction and subsequent signaling of cells is complex and regulated at multiple levels: receptor abundance, binding dynamics, phosphorylation/dephosphorylation of tyrosine and serine/threonine residues, and subsequent interactions of key intracellular messengers. We report early insulin signaling events in the mouse Sol8 myogenic cell line. Sol8 cells responded to insulin by increasing total IRS-1, p85 PI3-kinase and tyrosine phosphorylated IRS-1 (pY-IRS-1) at 10 min (P<0.05), but not at 1 min of insulin stimulation. The dose-response relationships at 10-min insulin (10 to 300 nM) stimulation showed that IRS-1 and pY-IRS-1 responded to 100 and 300 nM insulin, and the p85 PI3-kinase response peaked at 30 nM insulin. PI3-kinase appeared to be present in high abundance and, in response to insulin, recruitment to the insulin receptor tyrosine kinase (IR) of IRS-1 and PI3-kinase was observed. The increase in IRS-1 detected in IR immunoprecipitates was twofold, while the corresponding increase in PI3-kinase was threefold, suggesting direct recruitment of PI3-kinase to the IR. PI3-kinase detected in IRS-1 immunoprecipitates in response to insulin increased 1.7-fold. An ultimate target of this pathway, GLUT4 recruitment to the PM, was delayed (30 min), the increase in GLUT4 being of similar magnitude (1.6-fold) to the early signaling events. Saturation binding analysis indicated that IR in the plasma membrane was not down-regulated in response to insulin. The present study suggests that early signaling events in the insulin cascade are invoked in Sol8 myogenic cells and that this cell line provides a useful model to study insulin signaling.
Hill
Rodney A
RA
Department of Animal and Veterinary Science, University of Idaho, Moscow, Idaho 83844, USA. rodhill@uidaho.edu
Strat
A Lulu
AL
Hughes
Nikki J
NJ
Kokta
Theresa J
TJ
Dodson
Michael V
MV
Gertler
Arieh
A
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000378:metabolism*; D055504:Insulin Receptor Substrate Proteins; D051379:Mice; D008954:Models, Biological; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D009124:Muscle Proteins / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D011972:Receptor, Insulin / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*
D051275:Glucose Transporter Type 4; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D011972:Receptor, Insulin; C526295:Irs1 protein, mouse; C495693:Slc2a4 protein, mouse