Insulin promotes formation of polymerized microtubules by a phosphatidylinositol 3-kinase-independent, actin-dependent pathway in 3T3-L1 adipocytes.
Direct demonstrations implicating the microtubule cytoskeleton in insulin-mediated adipose/muscle-specific glucose transporter (GLUT4) translocation are beginning to emerge, and one role of the microtubule network appears to be the provision of a solid support for GLUT4 vesicle movement. In the current study we show that insulin treatment increases total polymerized alpha-tubulin in microtubules in a time- and dose-dependent manner that coincides with established insulin-mediated changes in GLUT4 translocation. Insulin stimulates the growth of microtubules through a pathway that requires tyrosine kinase activity, as indicated by inhibition of the effect after treatment with genistein. Insulin-mediated growth was not inhibited by treatment with the MAPK kinase (MEK) inhibitor, PD98059 or by wortmannin, indicating that the effect does not require activation of extracellular signal-regulated kinase 1/2 or phosphatidylinositide 3-kinase. Depolymerization of the actin cytoskeleton with latrunculin B abrogated the effect of insulin on microtubule polymerization, indicating that an intact actin network is a requirement for insulin-dependent modulation of microtubules. Using methods that measure insulin-dependent GLUT4 translocation in populations of adipocytes as opposed to individual cells, we show a statistically significant reduction in translocation (30% inhibition) in the presence of low concentrations of nocodazole (2 mum). This concentration incompletely depolymerizes the microtubule network, revealing that partial depolymerization of microtubules is sufficient to inhibit GLUT4 translocation. It is likely that stabilization of the microtubule network contributes to insulin stimulation of GLUT4 translocation.
Olson
Ann Louise
AL
Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, P.O. Box 26901, Room 853-BMSB, Oklahoma City, Oklahoma 73190, USA. ann-olson@ouhsc.edu
Eyster
Craig A
CA
Duggins
Quwanza S
QS
Knight
John B
JB
D016475:3T3 Cells; D000199:Actins / Q000502:physiology*; D017667:Adipocytes / Q000502:physiology*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000494:pharmacology*; D051379:Mice; D008870:Microtubules / Q000187:drug effects* / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D009124:Muscle Proteins; D015739:Nocodazole / Q000008:administration & dosage; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D011108:Polymers / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D013844:Thiazoles / Q000494:pharmacology; D053778:Thiazolidines; D013997:Time Factors; D014404:Tubulin / Q000378:metabolism*
D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D051275:Glucose Transporter Type 4; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D011108:Polymers; C495693:Slc2a4 protein, mouse; D013844:Thiazoles; D053778:Thiazolidines; D014404:Tubulin; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D020928:Mitogen-Activated Protein Kinases; C037068:latrunculin B; D015739:Nocodazole