Disassembly of the actin network inhibits insulin-dependent stimulation of glucose transport and prevents recruitment of glucose transporters to the plasma membrane.
In muscle and fat tissues, insulin stimulates glucose transport through the translocation of glucose transporter proteins from an intracellular storage pool to the plasma membrane. The mechanism of this translocation is unknown. We have examined the possible role of the actin microfilament network in the stimulation of glucose transport by insulin and on the distribution of glucose transporters, in differentiated L6 rat skeletal muscle cells. Insulin (10(-7) M for 30 min) caused a major reorganization of the actin network of differentiated L6 myotubes. Cytochalasin D, a widely used inhibitor of actin filament formation, caused a dose- and time-dependent disassembly of the actin network, which was associated with an 80% inhibition of the insulin stimulation of glucose transport, without affecting the basal rate of glucose uptake. L6 myotubes express three glucose transporter isoforms, named GLUT1, GLUT3, and GLUT4. Disassembly of the actin network by cytochalasin D did not affect the number of basal glucose transporters in the plasma membrane but reduced the content of all three glucose transporters in intracellular membranes and prevented their appearance at the plasma membrane response to insulin. The inhibitory effect of cytochalasin D treatment on the insulin stimulation of glucose transport occurred downstream of tyrosine phosphorylation of the insulin receptor substrate-1 and of binding of phosphatidylinositol 3-kinase to the insulin receptor substrate-1. Using immunoprecipitation of intact membranes, we detected specific association of the actin-binding protein spectrin with GLUT4 glucose transporter-containing vesicles. We conclude that an intact actin network is required for the correct intracellular localization of glucose transporters, as well as for their incorporation into the plasma membrane in response to insulin. A direct interaction may exist between the actin network and the glucose transporter vesicles which may be mediated through a spectrin-containing skeleton attached to glucose transporter-containing vesicles.
Tsakiridis
T
T
Division of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada.
Vranic
M
M
Klip
A
A
D000199:Actins / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured / Q000378:metabolism; D015638:Cytochalasin D / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism*; D007328:Insulin / Q000494:pharmacology*; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009132:Muscles / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D011233:Precipitin Tests; D051381:Rats; D015398:Signal Transduction
D000199:Actins; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D015638:Cytochalasin D; D005947:Glucose