Activation of Protein Kinase C (α, β, and ζ) by Insulin in 3T3/L1 Cells

Abstract

We presently studied (a) insulin effects on protein kinase C (PKC) and (b) effects of transfection-induced, stable expression of PKC isoforms on glucose transport in 3T3/L1 cells. In both fibroblasts and adipocytes, insulin provoked increases in membrane PKC enzyme activity and membrane levels of PKC-alpha and PKC-beta. However, insulin-induced increases in PKC enzyme activity were apparent in both non-down-regulated adipocytes and adipocytes that were down-regulated by overnight treatment with 5 microM phorbol ester, which largely depletes PKC-alpha, PKC-beta, and PKC-epsilon, but not PKC-zeta. Moreover, insulin provoked increases in the enzyme activity of immunoprecipitable PKC-zeta. In transfection studies, stable overexpression of wild-type or constitutively active forms of PKC-alpha, PKC-beta1, and PKC-beta2 failed to influence basal or insulin-stimulated glucose transport (2-deoxyglucose uptake) in fibroblasts and adipocytes, despite inhibiting insulin effects on glycogen synthesis. In contrast, stable overexpression of wild-type PKC-zeta increased, and a dominant-negative mutant form of PKC-zeta decreased, basal and insulin-stimulated glucose transport in fibroblasts and adipocytes. These findings suggested that: (a) insulin activates PKC-zeta, as well as PKC-alpha and beta; and (b) PKC-zeta is required for, and may contribute to, insulin effects on glucose transport in 3T3/L1 cells.

Insulin Stimulates PKCζ-mediated Phosphorylation of Insulin Receptor Substrate-1 (IRS-1)

“…They are activated by insulin through a PI3K-dependent mechanism (27,38), and insulin-stimulated glucose transport depends upon the activation of one or both of these enzymes (27,28,38,39). In the present study we provide evidence that PKC also plays a major regulatory role in a negative feedback control mechanism induced by insulin to terminate its own signaling pathways.…”

“…Hence, phosphatidylinositol 1,4,5-trisphosphate may promote PKC signaling by colocalizing the enzyme in close proximity to its substrates. PKC can be activated by insulin, and this activation is blocked by inhibitors of PI3K and the expression of dominant negative p85 (27,28). Phosphorylation of PKC is required for its full activation; two of the regulatory phosphorylation sites on PKC are targets for phosphorylation by PDK-1 and PDK-2 (25), thus representing another mechanism by which activation of PI3K affects the activity of PKC.…”

Lysophosphatidylcholine Activates Adipocyte Glucose Uptake and Lowers Blood Glucose Levels in Murine Models of Diabetes

“…The involvement of conventional or atypical PKC activation in the promotion of glucose uptake by adipocytes and muscle cells is well established (30), but the activation of novel PKCs, especially of PKC␦, also controls glucose uptake. Stimulation of the translocation of GLUT4 to the plasma membrane and glucose uptake by insulin were found to be inhibited by rottlerin in rat skeletal muscle cells, and the overexpression of PKC␦ induced by GLUT4 translocation to the plasma membrane was found to increase basal glucose uptake to levels attained by insulin (31).…”

Molecular Basis of Insulin-stimulated GLUT4 Vesicle Trafficking

“…However, the specific downstream targets for PI 3-kinase have remained questionable with evidence both for and against protein kinase B, also known as Akt (20 -22). Alternatively, the atypical protein kinase C isoforms (protein kinase C and ) are downstream targets of PI 3-kinase and have also been implicated in the insulin stimulation of GLUT4 translocation (23,24,81).…”