Phosphatidylinositol 3-kinase regulates Ca2+ signaling in pancreatic acinar cells through inhibition of sarco(endo)plasmic reticulum Ca2+-ATPase

2004 ◽  
Vol 287 (6) ◽  
pp. G1200-G1212 ◽  
Author(s):  
L. Fischer ◽  
A. S. Gukovskaya ◽  
S. H. Young ◽  
I. Gukovsky ◽  
A. Lugea ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Specific Inhibitor ◽  
Acinar Cells ◽  
Pi3 Kinase ◽  
Pancreatic Acinar Cells ◽  
Phosphatidylinositol 3 Kinase ◽  
Kinase Inhibition ◽  
Plasmic Reticulum ◽  
Genetic Deletion ◽  
Phosphatidylinositol 3

Calcium is a key mediator of hormone-induced enzyme secretion in pancreatic acinar cells. At the same time, abnormal Ca2+ responses are associated with pancreatitis. We have recently shown that inhibition of phosphatidylinositol 3-kinase (PI3-kinase) by LY-294002 and wortmannin, as well as genetic deletion of PI3-kinase-γ, regulates Ca2+ responses and the Ca2+-sensitive trypsinogen activation in pancreatic acinar cells. The present study sought to determine the mechanisms of PI3-kinase involvement in Ca2+ responses induced in these cells by CCK and carbachol. The PI3-kinase inhibitors inhibited both Ca2+ influx and mobilization from intracellular stores induced by stimulation of acini with physiological and pathological concentrations of CCK, as well as with carbachol. PI3-kinase inhibition facilitated the decay of cytosolic free Ca2+ concentration ([Ca2+]i) oscillations observed in individual acinar cells. The PI3-kinase inhibitors decreased neither CCK-induced inositol 1,4,5-trisphosphate [Ins( 1 , 4 , 5 )P3] production nor Ins( 1 , 4 , 5 )P3-induced Ca2+ mobilization, suggesting that the effect of PI3-kinase inhibition is not through Ins( 1 , 4 , 5 )P3 or Ins( 1 , 4 , 5 )P3 receptors. PI3-kinase inhibition did not affect Ca2+ mobilization induced by thapsigargin, a specific inhibitor of sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA). Moreover, SERCA blockade with thapsigargin abolished the effects of pharmacological and genetic PI3-kinase inhibition on [Ca2+]i signals, suggesting SERCA as a downstream target of PI3-kinase. Both pharmacological PI3-kinase inhibition and genetic deletion of PI3-kinase-γ increased the amount of Ca2+ in intracellular stores during CCK stimulation. Finally, addition of the PI3-kinase product phosphatidylinositol 3,4,5-trisphosphate to permeabilized acini significantly attenuated Ca2+ reloading into the endoplasmic reticulum. The results indicate that PI3-kinase regulates Ca2+ signaling in pancreatic acinar cells through its inhibitory effect on SERCA.

Phosphatidylinositol 3-kinase facilitates bile acid-induced Ca2+ responses in pancreatic acinar cells

2007 ◽  
Vol 292 (3) ◽  
pp. G875-G886 ◽  
Author(s):  
L. Fischer ◽  
A. S. Gukovskaya ◽  
J. M. Penninger ◽  
O. A. Mareninova ◽  
H. Friess ◽  
...  
Keyword(s):  
Bile Acid ◽  
Bile Acids ◽  
Acinar Cells ◽  
Pancreatic Acinar Cells ◽  
Phosphatidylinositol 3 Kinase ◽  
Pi3k Inhibitors ◽  
Trypsinogen Activation ◽  
Plasmic Reticulum ◽  
Intracellular Ca ◽  
Phosphatidylinositol 3

Bile acids are known to induce Ca2+ signals in pancreatic acinar cells. We have recently shown that phosphatidylinositol 3-kinase (PI3K) regulates changes in free cytosolic Ca2+ concentration ([Ca2+]i) elicited by CCK by inhibiting sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA). The present study sought to determine whether PI3K regulates bile acid-induced [Ca2+]i responses. In pancreatic acinar cells, pharmacological inhibition of PI3K with LY-294002 or wortmannin inhibited [Ca2+]i responses to taurolithocholic acid 3-sulfate (TLC-S) and taurochenodeoxycholate (TCDC). Furthermore, genetic deletion of the PI3K γ-isoform also decreased [Ca2+]i responses to bile acids. Depletion of CCK-sensitive intracellular Ca2+ pools or application of caffeine inhibited bile acid-induced [Ca2+]i signals, indicating that bile acids release Ca2+ from agonist-sensitive endoplasmic reticulum (ER) stores via an inositol ( 1 , 4 , 5 )-trisphosphate-dependent mechanism. PI3K inhibitors increased the amount of Ca2+ in intracellular stores during the exposure of acinar cells to bile acids, suggesting that PI3K negatively regulates SERCA-dependent Ca2+ reloading into the ER. Bile acids inhibited Ca2+ reloading into ER in permeabilized acinar cells. This effect was augmented by phosphatidylinositol ( 3 , 4 , 5 )-trisphosphate (PIP3), suggesting that both bile acids and PI3K act synergistically to inhibit SERCA. Furthermore, inhibition of PI3K by LY-294002 completely inhibited trypsinogen activation caused by the bile acid TLC-S. Our results indicate that PI3K and its product, PIP3, facilitate bile acid-induced [Ca2+]i responses in pancreatic acinar cells through inhibition of SERCA-dependent Ca2+ reloading into the ER and that bile acid-induced trypsinogen activation is mediated by PI3K. The findings have important implications for the mechanism of acute pancreatitis since [Ca2+]i increases and trypsinogen activation mediate key pathological processes in this disorder.

scholarly journals c-Cbl Is Tyrosine-Phosphorylated by Interleukin-4 and Enhances Mitogenic and Survival Signals of Interleukin-4 Receptor by Linking With the Phosphatidylinositol 3′-Kinase Pathway

Blood ◽  
1998 ◽  
Vol 91 (1) ◽  
pp. 46-53 ◽  
Author(s):  
Hiroo Ueno ◽  
Ko Sasaki ◽  
Hiroaki Honda ◽  
Tetsuya Nakamoto ◽  
Tetsuya Yamagata ◽  
...  
Keyword(s):  
Specific Inhibitor ◽  
Pi3 Kinase ◽  
Interleukin 4 ◽  
Phosphatidylinositol 3 Kinase ◽  
Proliferation And Differentiation ◽  
Interleukin 4 Receptor ◽  
Pi3 Kinase Pathway ◽  
Kinase Pathway ◽  
Phosphatidylinositol 3 ◽  
Survival Signals

Interleukin-4 (IL-4) is a cytokine that induces both proliferation and differentiation and suppresses apoptosis of B cells. Although IL-4 has been shown to activate the phosphatidylinositol 3′ (PI3)-kinase pathway, the role of PI3 kinase in the IL-4 receptor (IL-4R) signaling remains unclear. In this study, we demonstrated that c-Cbl proto-oncogene product is inducibly phosphorylated on tyrosine residues and is associated with the p85 subunit of PI3-kinase by IL-4 stimulation. Overexpression of c-Cbl enhances the PI3-kinase activity and, at the same time, mitogenic activity and survival of cells in the presence of IL-4. However, these effects of c-Cbl were abolished by wortmannin, a specific inhibitor for the PI3 kinase pathway, or by a point mutation at tyrosine 731 of c-Cbl, which is a major binding site for p85. These results indicate that c-Cbl plays a role in linking IL-4R with the PI3 kinase pathway and thus enhancing the mitogenic and survival signals.

scholarly journals Both Src-Dependent and -Independent Mechanisms Mediate Phosphatidylinositol 3-Kinase Regulation of Colony-Stimulating Factor 1-Activated Mitogen-Activated Protein Kinases in Myeloid Progenitors

2000 ◽  
Vol 20 (18) ◽  
pp. 6779-6798 ◽  
Author(s):  
Angel W.-M. Lee ◽  
David J. States
Keyword(s):  
Kinase Inhibitors ◽  
Pi3 Kinase ◽  
Dominant Negative ◽  
Dependent Manner ◽  
Colony Stimulating Factor ◽  
Phosphatidylinositol 3 Kinase ◽  
Stimulating Factor ◽  
In Cells ◽  
Phosphatidylinositol 3 ◽  
Myeloid Progenitors

ABSTRACT Colony-stimulating factor 1 (CSF-1) supports the proliferation, survival, and differentiation of bone marrow-derived cells of the monocytic lineage. In the myeloid progenitor 32D cell line expressing CSF-1 receptor (CSF-1R), CSF-1 activation of the extracellular signal-regulated kinase (ERK) pathway is both Ras and phosphatidylinositol 3-kinase (PI3-kinase) dependent. PI3-kinase inhibition did not influence events leading to Ras activation. Using the activity of the PI3-kinase effector, Akt, as readout, studies with dominant-negative and oncogenic Ras failed to place PI3-kinase downstream of Ras. Thus, PI3-kinase appears to act in parallel to Ras. PI3-kinase inhibitors enhanced CSF-1-stimulated A-Raf and c-Raf-1 activities, and dominant-negative A-Raf but not dominant-negative c-Raf-1 reduced CSF-1-provoked ERK activation, suggesting that A-Raf mediates a part of the stimulatory signal from Ras to MEK/ERK, acting in parallel to PI3-kinase. Unexpectedly, a CSF-1R lacking the PI3-kinase binding site (ΔKI) remained capable of activating MEK/ERK in a PI3-kinase-dependent manner. To determine if Src family kinases (SFKs) are involved, we demonstrated that CSF-1 activated Fyn and Lyn in cells expressing wild-type (WT) or ΔKI receptors. Moreover, CSF-1-induced Akt activity in cells expressing ΔKI is SFK dependent since Akt activation was prevented by pharmacological or genetic inhibition of SFK activity. The docking protein Gab2 may link SFK to PI3-kinase. CSF-1 induced Gab2 tyrosyl phosphorylation and association with PI3-kinase in cells expressing WT or ΔKI receptors. However, only in ΔKI cells are these events prevented by PP1. Thus in myeloid progenitors, CSF-1 can activate the PI3-kinase/Akt pathway by at least two mechanisms, one involving direct receptor binding and one involving SFKs.

scholarly journals c-Cbl Is Tyrosine-Phosphorylated by Interleukin-4 and Enhances Mitogenic and Survival Signals of Interleukin-4 Receptor by Linking With the Phosphatidylinositol 3′-Kinase Pathway

Blood ◽  
1998 ◽  
Vol 91 (1) ◽  
pp. 46-53 ◽  
Author(s):  
Hiroo Ueno ◽  
Ko Sasaki ◽  
Hiroaki Honda ◽  
Tetsuya Nakamoto ◽  
Tetsuya Yamagata ◽  
...  
Keyword(s):  
Specific Inhibitor ◽  
Pi3 Kinase ◽  
Interleukin 4 ◽  
Phosphatidylinositol 3 Kinase ◽  
Proliferation And Differentiation ◽  
Interleukin 4 Receptor ◽  
Pi3 Kinase Pathway ◽  
Kinase Pathway ◽  
Phosphatidylinositol 3 ◽  
Survival Signals

Abstract Interleukin-4 (IL-4) is a cytokine that induces both proliferation and differentiation and suppresses apoptosis of B cells. Although IL-4 has been shown to activate the phosphatidylinositol 3′ (PI3)-kinase pathway, the role of PI3 kinase in the IL-4 receptor (IL-4R) signaling remains unclear. In this study, we demonstrated that c-Cbl proto-oncogene product is inducibly phosphorylated on tyrosine residues and is associated with the p85 subunit of PI3-kinase by IL-4 stimulation. Overexpression of c-Cbl enhances the PI3-kinase activity and, at the same time, mitogenic activity and survival of cells in the presence of IL-4. However, these effects of c-Cbl were abolished by wortmannin, a specific inhibitor for the PI3 kinase pathway, or by a point mutation at tyrosine 731 of c-Cbl, which is a major binding site for p85. These results indicate that c-Cbl plays a role in linking IL-4R with the PI3 kinase pathway and thus enhancing the mitogenic and survival signals.

scholarly journals Neurite outgrowth of PC12 cells is suppressed by wortmannin, a specific inhibitor of phosphatidylinositol 3-kinase.

1994 ◽  
Vol 269 (29) ◽  
pp. 18961-18967
Author(s):  
K. Kimura ◽  
S. Hattori ◽  
Y. Kabuyama ◽  
Y. Shizawa ◽  
J. Takayanagi ◽  
...  
Keyword(s):  
Neurite Outgrowth ◽  
Pc12 Cells ◽  
Specific Inhibitor ◽  
Phosphatidylinositol 3 Kinase ◽  
Phosphatidylinositol 3

scholarly journals Inhibition of kinases impairs neutrophil activation and killing of Staphylococcus aureus

1998 ◽  
Vol 331 (2) ◽  
pp. 489-495 ◽  
Author(s):  
Bruno SCHNYDER ◽  
Paul C. MEUNIER ◽  
Bruce D. CAR
Keyword(s):  
Staphylococcus Aureus ◽  
Map Kinases ◽  
Specific Inhibitor ◽  
Neutrophil Function ◽  
Oxygen Radical ◽  
Polymorphonuclear Neutrophils ◽  
Detectable Effect ◽  
Phosphatidylinositol 3 Kinase ◽  
Radical Generation ◽  
Phosphatidylinositol 3

Intracellular phosphorylations polymorphonuclear neutrophils are mediated by kinases, including mitogen activated-protein (MAP) kinases and phosphatidylinositol 3-kinase. In the present study we demonstrate their effector functions upon both ligation of cell-surface seven-transmembrane-spanning receptors by bacterial peptide formylmethionyl-leucylphenylalanine as well as in the process of destruction of Staphylococcus aureus. To regulate neutrophil MAP kinases p38 and p44/42, specifically, we made use of their specific inhibitors 10 µM SK&F 86002 (for p38) and PD 098059 (for activating kinase of p44/42). SK&F 86002 was a potent inhibitor (by 70%) of induced antimicrobial oxygen-radical generation compared with PD 098059 (by 20%). SK&F 86002 and PD 098059 inhibited mobilization of a dominant neutrophil adhesion molecule, β2 integrin, from cytoplasmic granules to the plasma membrane by 40 and 10% respectively, and the combination of the two drugs resulted in a 90% effect. The combined effect of both drugs was moderate inhibition of bacterial destruction, despite the fact that neither compound had detectable effect on bactericidal activity if applied individually. Bacterial destruction was also inhibited by wortmannin (0.1 µM), the specific inhibitor of phosphatidylinositol 3-kinase, which had previously been described to target various other activations of the neutrophil, including oxygen-radical generation. Although the relative contribution of p38 and p44/42 MAP kinases varied, the marked effects of the combined inhibition of the kinases revealed their concerted actions to be critical for normal neutrophil function.

scholarly journals Regulation of Hepatic Insulin-Like Growth Factor-Binding Protein-1 Gene Expression by Insulin: Central Role for Mammalian Target of Rapamycin Independent of Forkhead Box O Proteins

Endocrinology ◽  
2006 ◽  
Vol 147 (5) ◽  
pp. 2383-2391 ◽  
Author(s):  
Catherine Mounier ◽  
Victor Dumas ◽  
Barry I. Posner
Keyword(s):  
Gene Expression ◽  
Rat Liver ◽  
Binding Protein ◽  
Mammalian Target Of Rapamycin ◽  
Pi3 Kinase ◽  
Target Of Rapamycin ◽  
Mrna Levels ◽  
Phosphatidylinositol 3 Kinase ◽  
Insulin Inhibition ◽  
Phosphatidylinositol 3

The expression of IGF-binding protein-1 (IGFBP-1) is induced in rat liver by dexamethasone and glucagon and is completely inhibited by 100 nm insulin. Various studies have implicated phosphatidylinositol 3-kinase, protein kinase B (Akt), phosphorylation of the transcription factors forkhead in rhabdomyosarcoma 1 (Foxo1)/Foxo3, and the mammalian target of rapamycin (mTOR) in insulin’s effect. In this study we examined insulin regulation of IGFBP-1 in both subconfluent and confluent hepatocytes. In subconfluent hepatocytes, insulin inhibition of IGFBP-1 mRNA levels was blocked by inhibiting PI3 kinase activation, and there was a corresponding inhibition of Foxo1/Foxo3 phosphorylation. In these same cells, inhibition of the insulin effect by rapamycin occurred in the presence of insulin-induced Foxo1/Foxo3 phosphorylation. In confluent hepatocytes, insulin could not activate the phosphatidylinositol 3-kinase (PI3 kinase)-Akt-Foxo1/Foxo3 pathway, but still inhibited IGFBP-1 gene expression in an mTOR-dependent manner. In subconfluent hepatocytes, the serine/threonine phosphatase inhibitor okadaic acid (100 nm) partially inhibited IGFBP-1 gene expression by 40%, but did not produce phosphorylation of either Akt or Foxo proteins. In contrast, 1 nm insulin inhibited the IGFBP-1 mRNA level by 40% and correspondingly activated Akt and Foxo1/Foxo3 phosphorylation to a level comparable to that observed with 100 nm insulin. These results suggest a potential role for a serine/threonine phosphatase(s) in the regulation of IGFBP-1 gene transcription, which is not downstream of mTOR and is independent of Akt. In conclusion, we have found that in rat liver, insulin inhibition of IGFBP-1 mRNA levels can occur in the absence of the phosphorylation of Foxo1/Foxo3, whereas activation of the mTOR pathway is both necessary and sufficient.

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