Different Effects of Insulin and Platelet‐Derived Growth Factor on Phosphatidylinositol 3‐Kinase at the Subcellular Level in 3T3‐L1 Adipocytes

Abstract

Insulin stimulates glucose uptake by induction of the translocation of vesicles that contain the glucose transporter Glut 4 to the plasma membrane. Phosphatidylinositol 3-kinase (Ptdlns 3-kinase), which is thought to be involved in intracellular trafficking, could play a critical role in insulin-induced glucose transport. In 3T3-Ll adipocytes, insulin and platelet-derived-growth-factor (PDGF) stimulated glucose uptake by 5.8-fold and 2.4-fold, respectively, but PDGF had no significant effect on Glut 4 translocation. Nevertheless, both hormones activated PtdIns 3-kinase activity in total cell extracts. However, insulin and PDGF had different effects on the stimulation of PtdIns 3-kinase activity in several subcellular fractions, and the movements of insulin-receptor substrate (IRS) 1 and the p85 subunit of PtdIns 3-kinase between subcellular compartments. PDGF stimulated PtdIns 3-kinase activity almost exclusively in the plasma membrane, and induced translocation of the p85 subunit from the cytosol to the plasma membrane, where the PDGF receptor was phosphorylated on tyrosine residues. In contrast, insulin stimulated Ptdlns 3-kinase activity in the plasma membrane, in low-density microsomes (LDM) and in cytosol. Furthermore, insulin induced the translocation of p85 from the cytosol to LDM and the translocation of IRS 1 from LDM to the cytosol. These data indicate that insulin and PDGF have different effects on the activation of Ptdlns 3-kinase and on the movement of IRS 1 and PtdIns 3-kinase between subcellular compartments. We would like to suggest that a crucial event in the stimulation of glucose uptake by insulin could be that insulin, but not PDGF, induces activation of PtdIns 3-kinase in the cytosol and in LDM, the compartment enriched in Glut-4-containing vesicles.

GLUT4 vesicle dynamics in living 3T3 L1 adipocytes visualized with green-fluorescent protein

“…Our data suggest that the primary site of insulin action must be to release GLUT4 vesicles from their tether, but it remains to be established whether insulin also acts at the translocation and fusion steps. This hypothesis is consistent with recent data suggesting that insulin-stimulated GLUT4 translocation may require targeting of PtdIns 3-kinase to the high-density microsomal fraction [23][24][25], and that additional PtdIns 3-kinase-independent events are also required [26]. Future work must be directed at resolving the identity of the intracellular site of attachment of GLUT4 vesicles, and determining whether PtdIns 3-kinase plays a role in the release of these vesicles, allowing them to translocate to the plasma membrane.…”

FGF-receptor substrate 2 functions as a molecular sensor integrating external regulatory signals into the FGF pathway

“…IRS-1 is the counterpart of FRS2 in the insulin signaling pathway and exhibits balanced PY and PT. Upon PS/T, either IRS-1 becomes a poor substrate for insulin receptor kinase [33] or its association with subcellular compartments and/or stability is probably altered [34,35]. FRS2 may also undergo similar changes.…”

Protein kinase B (c-Akt): a multifunctional mediator of phosphatidylinositol 3-kinase activation

“…This inhibition of insulin-stimulated glycogenesis may be explained in that persistent activation of this pathway could result in desensitization. Interestingly, PDGF, while able to activate PI-3K in 3T3-L1 adipocytes, is unable to stimulate GLUT4 translocation [74,75]. In these cells PDGF is also unable to induce phosphorylation of PKB, although activity measurements were not performed [76].…”

“…Activation of PI-3K by PDGF occurs only in plasma membranes, whereas insulin also induces activation in low-density microsomes, where GLUT4 expression is predominant [75,76]. Furthermore, okadaic acid, a potent stimulator of GLUT4 transport, induces PKB phosphorylation as efficiently as insulin without activation of PI-3K.…”