scholarly journals Activation of signal transduction in platelets by the tyrosine phosphatase inhibitor pervanadate (vanadyl hydroperoxide)

1992 ◽  
Vol 286 (2) ◽  
pp. 441-449 ◽  
Author(s):  
K M Pumiglia ◽  
L F Lau ◽  
C K Huang ◽  
S Burroughs ◽  
M B Feinstein
Keyword(s):  
Signal Transduction ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Tyrosine Phosphorylation ◽  
Actin Polymerization ◽  
Kinase Inhibitor ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine ◽  
Kinase C ◽  
Granule Secretion

The protein tyrosine phosphatase (PTPase) inhibitor pervanadate (vanadyl hydroperoxide) stimulated protein tyrosine phosphorylation 29-fold more than did thrombin in intact and saponin-permeabilized platelets. Increased tyrosine phosphorylation preceded, or was coincident with, a fall in PtdIns(4,5)P2 levels, production of PtdIns(3,4)P2 and phosphatidic acid, mobilization of intracellular Ca2+, stimulation of protein kinase C-dependent protein phosphorylation, secretion of dense and alpha-granules, increased actin polymerization, shape change and aggregation which required fibrinogen and was mediated by increased surface expression of GPIIb-IIIa. The tyrosine kinase inhibitor RG 50864 totally prevented induction of tyrosine phosphorylation by pervanadate, as well as all other responses measured; in contrast, the inactive structural analogue, tyrphostin #1, had no effect. Dense-granule secretion induced by pervanadate required protein kinase C activity; however, aggregation and alpha-granule secretion were independent of protein kinase C. In saponin-permeabilized platelets pervanadate and thrombin stimulated phospholipase C activity by GTP-independent and GTP-dependent mechanisms respectively. We conclude that PTPases are important regulators of signal transduction in platelets.

The protein kinase C inhibitor H-7 activates human neutrophils: effect on shape, actin polymerization, fluid pinocytosis and locomotion

1990 ◽  
Vol 96 (1) ◽  
pp. 99-106
Author(s):  
H.U. Keller ◽  
V. Niggli ◽  
A. Zimmermann ◽  
R. Portmann
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Actin Polymerization ◽  
Kinase Inhibitor ◽  
Human Neutrophils ◽  
Kinase C ◽  
Chemotactic Peptides ◽  
Protein Kinase C Inhibitor ◽  
Shape Changes ◽  
Stimulation Of

The present study demonstrates new properties of H-7. The protein kinase inhibitor H-7 is a potent activator of several neutrophil functions. Stimulation of initially spherical nonmotile neutrophils elicits vigorous shape changes within a few seconds, increases in cytoskeletal actin, altered F-actin distribution, increased adhesiveness and a relatively small increase in pinocytic activity. H-7 has also chemokinetic activities. Depending on the experimental condition, H-7 may elicit or inhibit neutrophil locomotion. It failed to induce chemotaxis. Thus, the response pattern elicited by H-7 is different from that of other leukocyte activators such as chemotactic peptides, PMA or diacylglycerols. The finding that H-7 can elicit shape changes, actin polymerization and pinocytosis suggests that these events can occur without activation of protein kinase C (PKC). PMA-induced shape changes and stimulation of pinocytosis were not inhibited by H-7.

Endothelin-1–Induced Interleukin-8 Production in Human Brain-Derived Endothelial Cells Is Mediated by the Protein Kinase C and Protein Tyrosine Kinase Pathways

Blood ◽  
1999 ◽  
Vol 94 (4) ◽  
pp. 1291-1299 ◽  
Author(s):  
R. Zidovetzki ◽  
P. Chen ◽  
M. Chen ◽  
F.M. Hofman
Keyword(s):  
Signal Transduction ◽  
Endothelial Cells ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Tyrosine Kinase ◽  
Protein Tyrosine Kinase ◽  
Protein Tyrosine ◽  
Kinase C ◽  
Eta Receptor ◽  
Specific Inhibitors

We have previously demonstrated that endothelin-1 (Et-1) induces human central nervous system-derived endothelial cells (CNS-EC) to produce and secrete the chemokine interleukin 8 (IL-8). In the present study, we use specific inhibitors and activators to elucidate the signal transduction pathways involved in this process. Et-1–induced IL-8 production was blocked by ETA receptor antagonist BQ610, but not by ETB receptor antagonist BQ788, demonstrating that CNS-EC activation is initiated by Et-1 binding to the ETA receptor. IL-8 mRNA expression is blocked by the protein kinase C inhibitor bisindolylmaleimide or protein tyrosine kinase inhibitors, genestein and geldanamycin, establishing the involvement of the protein kinase C and protein tyrosine kinase pathways in the activation process. The transcription factor, NF-κB, is involved in Et-1 activation as determined by specific inhibitors of translocation and direct analysis of DNA-binding proteins. Neither inhibition nor activation of cAMP-dependent protein kinase affected IL-8 production in the absence or presence of Et-1. Similarly, no effect was observed upon inhibition of protein phosphatases by okadaic acid. Thus, the signal transduction process induced by Et-1 in CNS-EC, leading to increased mRNA IL-8 expression, is initiated by Et-1 binding to ETA receptor followed by subsequent activation of protein kinase C, protein tyrosine kinase, and NF-κB. Because increased expression of Et-1 is associated with hypertension and stroke and IL-8 is likely to be involved in the accumulation of neutrophils causing tissue damage in ischemic/reperfusion injury, identification of the mechanism involved in the Et-1–induced increase in IL-8 production may have significant therapeutic value.

HC fragment (C-terminal portion of the heavy chain) of tetanus toxin activates protein kinase C isoforms and phosphoproteins involved in signal transduction

2001 ◽  
Vol 356 (1) ◽  
pp. 97-103 ◽  
Author(s):  
Carles GIL ◽  
Imane CHAIB-OUKADOUR ◽  
Juan BLASI ◽  
José AGUILERA
Keyword(s):  
Signal Transduction ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Tyrosine Phosphorylation ◽  
Heavy Chain ◽  
Tetanus Toxin ◽  
Signal Transduction Pathways ◽  
Terminal Portion ◽  
Kinase C ◽  
Pkc Isoforms

A recent report [Gil, Chaib-Oukadour, Pelliccioni and Aguilera (2000) FEBS Lett. 481, 177–182] describes activation of signal transduction pathways by tetanus toxin (TeTx), a Zn2+-dependent endopeptidase synthesized by the Clostridium tetani bacillus, which is responsible for tetanus disease. In the present work, specific activation of protein kinase C (PKC) isoforms and of intracellular signal-transduction pathways, which include nerve-growth-factor (NGF) receptor trkA, phospholipase C(PLC)γ-1 and extracellular regulated kinases (ERKs) 1 and 2, by the recombinant C-terminal portion of the TeTx heavy chain (HC-TeTx) is reported. The activation of PKC isoforms was assessed through their translocation from the soluble (cytosolic) compartment to the membranous compartment, showing that clear translocation of PKC-α, −β, −γ and −δ isoforms exists, whereas PKC-∊ showed a slight decrease in its soluble fraction immunoreactivity. The PKC-∊ isoform showed no consistent response. Using immunoprecipitation assays against phosphotyrosine residues, time- and dose-dependent increases in tyrosine phosphorylation were observed in the trkA receptor, PLCγ-1 and ERK-1/2. The effects shown by the HC-TeTx fragment on tyrosine phosphorylation were compared with the effects produced by NGF. The trkA and ERK-1/2 activation were corroborated using phospho-specific antibodies against trkA phosphorylated on Tyr490, and antibodies against Thr/Tyr phosphorylated ERK-1/2. Moreover, PLCγ-1 phosphorylation was supported by its HC-TeTx-induced translocation to the membranous compartment, an event related to PLCγ-1 activation. Since HC-TeTx is the domain responsible for membrane binding and lacks catalytic activity, the activations described here must be exclusively triggered by the interaction of TeTx with a membrane component.

SHIP-1 Associates with Tyrosine Phosphorylated Protein Kinase C-Delta at 155 Residue.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3004-3004
Author(s):  
Ramya Chari ◽  
Dheeraj Bhavanasi ◽  
James Daniel ◽  
Satya P. Kunapuli
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Tyrosine Phosphorylation ◽  
Conflicts Of Interest ◽  
Dense Granule ◽  
Protein Kinase C Delta ◽  
Tyrosine Residues ◽  
Kinase C ◽  
Dense Granule Secretion ◽  
Granule Secretion

Abstract Abstract 3004 Poster Board II-981 Protein Kinase C-delta (PKCδ) is a novel PKC isoform that differentially regulates platelet dense granule secretion. PKCδ positively regulates Protease activated receptor (PAR)-mediated dense granule secretion, whereas it negatively regulates glycoproteinVI (GPVI)-mediated dense granule secretion in platelets. PKCδ, a serine/threonine kinase is phosphorylated on its tyrosine residues. There are nine potential tyrosine phosphorylation sites in the regulatory domain of PKCδ. Phosphorylation at different tyrosine residues regulates its substrate specificity. We have previously shown that the association of PKCδ with Lyn and SHIP-1 negatively regulates GPVI-mediated dense granule secretion. However, the event leading to the association between PKCδ and SHIP-1 is not known. We hypothesize that the differential tyrosine phosphorylation of PKCδ downstream of PARs or GPVI receptors result in the preferential association with SHIP-1. In the current study, we show that PKCδ is phosphorylated at tyrosine residues Y332, Y523, Y525 and Y565 upon PAR or GPVI stimulation. Y311 residue is predominantly phosphorylated upon stimulation of PARs, whereas Y155 residue is preferentially phosphorylated upon GPVI stimulation. PAR-mediated Y311 phosphorylation peaks at later timepoint, whereas GPVI-mediated Y155 phosphorylation peaks at an early timepoint. correlating with dense granule secretion. Furthermore, we show that agarose-conjugated Y155 phosphorylated PKCδ peptide associates with SHIP-1 upon GPVI stimulation, and not PARs. These data suggest that the phosphorylation of PKCδ at distinct tyrosine residues differentially regulate its association with SHIP-1. Therefore, we conclude that the GPVI-mediated phosphorylation of PKCδ at 155 is required for its association with SHIP-1. This study is supported by pre-doctoral fellowship to Ramya Chari from American Heart Association, Pennsylvania-Delaware affiliate. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Lactosylceramide-enriched glycosphingolipid signaling domain mediates superoxide generation from human neutrophils

Blood ◽  
2002 ◽  
Vol 100 (4) ◽  
pp. 1454-1464 ◽  
Author(s):  
Kazuhisa Iwabuchi ◽  
Isao Nagaoka
Keyword(s):  
Signal Transduction ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Kinase Inhibitor ◽  
Signal Transduction Pathway ◽  
Transduction Pathway ◽  
Phosphatidylinositol 3 Kinase ◽  
Superoxide Generation ◽  
Kinase C ◽  
Signaling Domain

This study is focused on the functional significance of neutrophil lactosylceramide (LacCer)–enriched microdomains, which are involved in the initiation of a signal transduction pathway leading to superoxide generation. Treatment of neutrophils with anti-LacCer antibody, T5A7 or Huly-m13, induced superoxide generation from the cells, which was blocked by PP1, a Src kinase inhibitor; wortmannin, a phosphatidylinositol-3 kinase inhibitor; SB203580, a p38 mitogen-activated protein kinase (MAPK) inhibitor; and H7, an inhibitor for protein kinase C. When promyelocytic leukemia HL-60 cells were differentiated into neutrophilic lineage by dimethyl sulfoxide (DMSO) treatment, they acquired superoxide-generating activity but did not respond to anti-LacCer antibodies. Density gradient centrifugation revealed that LacCer and Lyn were recovered in detergent-insoluble membrane (DIM) of neutrophils and DMSO-treated HL-60 cells. However, immunoprecipitation experiments indicated that LacCer was associated with Lyn in neutrophils but not in DMSO-treated HL-60 cells. Interestingly, T5A7 induced the phosphorylation of Lyn in neutrophils but not in DMSO-treated HL-60 cells. Moreover, T5A7 induced the phosphorylation of p38 MAPK in neutrophils. T5A7-induced Lyn phosphorylation in neutrophil DIM fraction was significantly enhanced by cholesterol depletion or sequestration with methyl-β-cyclodextrin or nystatin. Collectively, these data suggest that neutrophils are characterized by the presence of cell surface LacCer-enriched glycosphingolipid signaling domain coupled with Lyn and that the ligand binding to LacCer induces the activation of Lyn, which may be suppressibly regulated by cholesterol, leading to superoxide generation through the phosphatidylinositol-3 kinase–, p38 MAPK–, and protein kinase C–dependent signal transduction pathway.

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