Role of the PI3K/PKB signaling pathway in cAMP-mediated translocation of rat liver Ntcp

1999 ◽  
Vol 277 (6) ◽  
pp. G1165-G1172 ◽  
Author(s):  
Cynthia R. L. Webster ◽  
M. Sawkat Anwer
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Signaling Pathway ◽  
Mitogen Activated Protein Kinase ◽  
Phosphatidylinositol 3 Kinase ◽  
Pi3k Inhibitors ◽  
Mapk Activity ◽  
Mitogen Activated Protein ◽  
Pi3k Signaling Pathway ◽  
Stimulation Of

cAMP stimulates Na+-taurocholate (TC) cotransport by translocating the Na+-TC-cotransporting peptide (Ntcp) to the plasma membrane. The present study was undertaken to determine whether the phosphatidylinositol-3-kinase (PI3K)-signaling pathway is involved in cAMP-mediated translocation of Ntcp. The ability of cAMP to stimulate TC uptake declined significantly when hepatocytes were pretreated with PI3K inhibitors wortmannin or LY-294002. Wortmannin inhibited cAMP-mediated translocation of Ntcp to the plasma membrane. cAMP stimulated protein kinase B (PKB) activity by twofold within 5 min, an effect inhibited by wortmannin. Neither basal mitogen-activated protein kinase (MAPK) activity nor cAMP-mediated inhibition of MAPK activity was affected by wortmannin. cAMP also stimulated p70S6K activity. However, rapamycin, an inhibitor of p70S6K, failed to inhibit cAMP-mediated stimulation of TC uptake, indicating that the effect of cAMP is not mediated via p70S6K. Cytochalasin D, an inhibitor of actin filament formation, inhibited the ability of cAMP to stimulate TC uptake and Ntcp translocation. Together, these results suggest that the stimulation of TC uptake and Ntcp translocation by cAMP may be mediated via the PI3K/PKB signaling pathway and requires intact actin filaments.

scholarly journals Insulin-Activated Protein Kinase Cβ Bypasses Ras and Stimulates Mitogen-Activated Protein Kinase Activity and Cell Proliferation in Muscle Cells

2000 ◽  
Vol 20 (17) ◽  
pp. 6323-6333 ◽  
Author(s):  
Pietro Formisano ◽  
Francesco Oriente ◽  
Francesca Fiory ◽  
Matilde Caruso ◽  
Claudia Miele ◽  
...  
Keyword(s):  
Growth Factor ◽  
Protein Kinase ◽  
Dna Synthesis ◽  
Antisense Oligonucleotide ◽  
Kinase Activity ◽  
Mitogen Activated Protein Kinase ◽  
Insulin Stimulation ◽  
Mapk Activity ◽  
Mitogen Activated Protein ◽  
Stimulation Of

ABSTRACT In L6 muscle cells expressing wild-type human insulin receptors (L6hIR), insulin induced protein kinase Cα (PKCα) and β activities. The expression of kinase-deficient IR mutants abolished insulin stimulation of these PKC isoforms, indicating that receptor kinase is necessary for PKC activation by insulin. In L6hIR cells, inhibition of insulin receptor substrate 1 (IRS-1) expression caused a 90% decrease in insulin-induced PKCα and -β activation and blocked insulin stimulation of mitogen-activated protein kinase (MAPK) and DNA synthesis. Blocking PKCβ with either antisense oligonucleotide or the specific inhibitor LY379196 decreased the effects of insulin on MAPK activity and DNA synthesis by >80% but did not affect epidermal growth factor (EGF)- and serum-stimulated mitogenesis. In contrast, blocking c-Ras with lovastatin or the use of the L61,S186 dominant negative Ras mutant inhibited insulin-stimulated MAPK activity and DNA synthesis by only about 30% but completely blocked the effect of EGF. PKCβ block did not affect Ras activity but almost completely inhibited insulin-induced Raf kinase activation and coprecipitation with PKCβ. Finally, blocking PKCα expression by antisense oligonucleotide constitutively increased MAPK activity and DNA synthesis, with little effect on their insulin sensitivity. We make the following conclusions. (i) The tyrosine kinase activity of the IR is necessary for insulin activation of PKCα and -β. (ii) IRS-1 phosphorylation is necessary for insulin activation of these PKCs in the L6 cells. (iii) In these cells, PKCβ plays a unique Ras-independent role in mediating insulin but not EGF or other growth factor mitogenic signals.

scholarly journals Activation of B-Raf and Regulation of the Mitogen-activated Protein Kinase Pathway by the Go α chain

2000 ◽  
Vol 11 (4) ◽  
pp. 1129-1142 ◽  
Author(s):  
Valeria Antonelli ◽  
Francesca Bernasconi ◽  
Yung H. Wong ◽  
Lucia Vallar
Keyword(s):  
Protein Kinase ◽  
Egf Receptor ◽  
Mapk Pathway ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Mitogen Activated Protein Kinase ◽  
Phosphatidylinositol 3 Kinase ◽  
Ovary Cells ◽  
Mapk Activity ◽  
Mitogen Activated Protein

Many receptors coupled to the pertussis toxin-sensitive Gi/o proteins stimulate the mitogen-activated protein kinase (MAPK) pathway. The role of the α chains of these G proteins in MAPK activation is poorly understood. We investigated the ability of Gαo to regulate MAPK activity by transient expression of the activated mutant Gαo-Q205L in Chinese hamster ovary cells. Gαo-Q205L was not sufficient to activate MAPK but greatly enhanced the response to the epidermal growth factor (EGF) receptor. This effect was not associated with changes in the state of tyrosine phosphorylation of the EGF receptor. Gαo-Q205L also potentiated MAPK stimulation by activated Ras. In Chinese hamster ovary cells, EGF receptors activate B-Raf but not Raf-1 or A-Raf. We found that expression of activated Gαo stimulated B-Raf activity independently of the activation of the EGF receptor or Ras. Inactivation of protein kinase C and inhibition of phosphatidylinositol-3 kinase abolished both B-Raf activation and EGF receptor-dependent MAPK stimulation by Gαo. Moreover, Gαo-Q205L failed to affect MAPK activation by fibroblast growth factor receptors, which stimulate Raf-1 and A-Raf but not B-Raf activity. These results suggest that Gαo can regulate the MAPK pathway by activating B-Raf through a mechanism that requires a concomitant signal from tyrosine kinase receptors or Ras to efficiently stimulate MAPK activity. Further experiments showed that receptor-mediated activation of Gαo caused a B-Raf response similar to that observed after expression of the mutant subunit. The finding that Gαo induces Ras-independent and protein kinase C- and phosphatidylinositol-3 kinase-dependent activation of B-Raf and conditionally stimulates MAPK activity provides direct evidence for intracellular signals connecting this G protein subunit to the MAPK pathway.

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