The protein kinase inhibitor staurosporine, like phorbol esters, induces the association of protein kinase C with membranes

1988 ◽  
Vol 154 (3) ◽  
pp. 1273-1279 ◽  
Author(s):  
Marlene Wolf ◽  
Marco Baggiolini
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Esters ◽  
Kinase Inhibitor ◽  
Protein Kinase Inhibitor ◽  
Kinase C

scholarly journals Rab6 is phosphorylated in thrombin-activated platelets by a protein kinase C-dependent mechanism: effects on GTP/GDP binding and cellular distribution

1999 ◽  
Vol 342 (2) ◽  
pp. 353-360 ◽  
Author(s):  
Michael L. FITZGERALD ◽  
Guy L. REED
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Esters ◽  
Kinase Inhibitor ◽  
Mitogen Activated Protein Kinase ◽  
Secretory Cells ◽  
Kinase C ◽  
Kd Values ◽  
Mitogen Activated Protein ◽  
Thrombin Activation

In platelets and other secretory cells, protein kinase C (PKC) plays a role in exocytosis stimulated by physiological extracellular signals, although its linkage to the secretory machinery is poorly understood. We investigated whether Rab6, a GTP-binding protein that fractionates with platelet α-granules, may be involved in linking these processes. We found that Rab6 contains two PKC consensus phosphorylation sites that are evolutionarily conserved. In platelets metabolically labelled with [32P]Pi, Rab6 phosphorylation was induced by phorbol esters or by thrombin. This phosphorylation was blocked by a specific PKC inhibitor (Ro-31-8220), but not by a p38 mitogen-activated protein kinase inhibitor (PD-169316). Physiological stimulation of platelets caused a PKC-dependent translocation of Rab6 from platelet particulate fractions, nearly doubling the fraction of Rab6 in the cytosol. A human Rab6 isoform (Rab6C) that is preferentially expressed in human platelet RNA was cloned and its phosphorylation by PKC was characterized. Rab6C incorporated up to 2 mol of [32P]Pi per mol of active protein. Rab6C bound GDP and GTP with Kd values of 113±12 and 119±27 nM respectively, and hydrolysed GTP at a rate of 100±15 μmol of GTP/mol of Rab6C per min. PKC phosphorylation of Rab6C increased the affinity for GTP by 3-fold, although it had lesser effects on GDP (1.6-fold). Phosphorylation did not alter the GTPase activity. In summary, thrombin activation of platelets leads to PKC-dependent phosphorylation of Rab6 and a translocation of Rab6 to the cytosol. We suggest that PKC phosphorylation may be an important mechanism through which Rab functional interactions in vesicle trafficking and secretion can be altered in response to an external stimulus.

Involvement of protein kinase C in serotonin-induced spike broadening and synaptic facilitation in sensorimotor connections of Aplysia

1992 ◽  
Vol 68 (2) ◽  
pp. 643-651 ◽  
Author(s):  
S. Sugita ◽  
J. R. Goldsmith ◽  
D. A. Baxter ◽  
J. H. Byrne
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Sensory Neurons ◽  
Transmitter Release ◽  
Phorbol Esters ◽  
Kinase Inhibitor ◽  
Model System ◽  
Short Term ◽  
Kinase C ◽  
Kinase A

1. Plasticity at the connections between sensory neurons and their follower cells in Aplysia has been used extensively as a model system to examine mechanisms of simple forms of learning. Earlier studies have concluded that serotonin (5-HT) is a key modulatory transmitter and that it exerts its short-term actions via cAMP-dependent activation of protein kinase A. Subsequently, it has become clear that other kinase systems such as protein kinase C (PKC) also may be involved in the actions of 5-HT. 2. Application of phorbol esters, which activate PKC, produced a slowly developing spike broadening but had little effect on excitability (a process known to be primarily cAMP dependent). Moreover, the effects of phorbol esters and 5-HT on spike duration were not additive, suggesting that they may share some common mechanisms. 3. The protein kinase inhibitor staurosporine suppressed both 5-HT-induced slowly developing spike broadening and, under certain conditions, facilitation of transmitter release. Staurosporine did not inhibit 5-HT-induced enhancement of excitability. The effectiveness of staurosporine on spike broadening was dependent on the time at which spike broadening was examined after application of 5-HT. Staurosporine appeared to have little effect on spike broadening 3 min after application of 5-HT, whereas it inhibited significantly 5-HT-induced spike broadening at later times. The staurosporine-insensitive component of 5-HT-induced spike broadening may be mediated by cAMP. 4. The results suggest that the activation of PKC plays a key role in components of both 5-HT-induced spike broadening and facilitation of synaptic transmission.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Regulation of T-cell-receptor-stimulated bivalent-cation entry in Jurkat E6 cells: role of protein kinase C

1994 ◽  
Vol 303 (2) ◽  
pp. 671-677 ◽  
Author(s):  
L A Conroy ◽  
J E Merritt ◽  
T J Hallam
Keyword(s):  
Protein Kinase C ◽  
T Cell ◽  
Protein Kinase ◽  
Phospholipase C ◽  
T Cell Receptor ◽  
Phorbol Esters ◽  
Kinase Inhibitor ◽  
Cell Receptor ◽  
Kinase C ◽  
Bivalent Cations

Stimulation of Jurkat E6 cells with anti-CD3 antibody results in a characteristic rise in [Ca2+]i which is due to both the release of Ca2+ from intracellular stores and the entry of external Ca2+. Individual components of the [Ca2+]i increase were investigated by measuring intracellular Ca2+ release in the absence of external Ca2+ and determining influx of bivalent cations by following the entry of Mn2+. The increase in [Ca2+]i induced by anti-CD3 antibody in the presence or absence of extracellular Ca2+ could be inhibited by the non-selective kinase inhibitor staurosporine, which also inhibits anti-CD3-stimulated phospholipase C activity. Staurosporine also inhibits the influx of bivalent cations induced by anti-CD3 antibody, but not that induced by depletion of intracellular Ca2+ stores using thapsigargin. The effect of staurosporine was compared with that of Ro 31-8425, a potent and selective inhibitor of protein kinase C (PKC). Ro 31-8425, at concentrations up to 10 microM, has no inhibitory effect on the anti-CD3 antibody-induced [Ca2+]i increase or phospholipase C activity. These studies are consistent with the concept that augmentation of [Ca2+]i by stimulated T-cell receptors requires activation of a kinase, probably a tyrosine kinase such as p56lck, ZAP-70 or p59fyn, and is independent of PKC. Phorbol esters inhibit the anti-CD3-stimulated [Ca2+]i increase and phospholipase C activity, showing that this can be negatively regulated by PKC. A small potentiation of the anti-CD3 antibody-induced [Ca2+]i rise in the presence of extracellular Ca2+ was detected in the presence of Ro 31-8425; this suggests that T-cell-receptor ligation can also limit the increase in [Ca2+]i via PKC activation.

scholarly journals Tumor promotion by depleting cells of protein kinase C delta.

1997 ◽  
Vol 17 (6) ◽  
pp. 3418-3428 ◽  
Author(s):  
Z Lu ◽  
A Hornia ◽  
Y W Jiang ◽  
Q Zang ◽  
S Ohno ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Ester ◽  
Phorbol Esters ◽  
Kinase Inhibitor ◽  
Tumor Promotion ◽  
Dominant Negative ◽  
Prolonged Treatment ◽  
Bryostatin 1 ◽  
Kinase C

Tumor-promoting phorbol esters activate, but then deplete cells of, protein kinase C (PKC) with prolonged treatment. It is not known whether phorbol ester-induced tumor promotion is due to activation or depletion of PKC. In rat fibroblasts overexpressing the c-Src proto-oncogene, the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) induced anchorage-independent growth and other transformation-related phenotypes. The appearance of transformed phenotypes induced by TPA in these cells correlated not with activation but rather with depletion of expressed PKC isoforms. Consistent with this observation, PKC inhibitors also induced transformed phenotypes in c-Src-overexpressing cells. Bryostatin 1, which inhibited the TPA-induced down-regulation of the PKCdelta isoform specifically, blocked the tumor-promoting effects of TPA, implicating PKCdelta as the target of the tumor-promoting phorbol esters. Consistent with this hypothesis, expression of a dominant negative PKCdelta mutant in cells expressing c-Src caused transformation of these cells, and rottlerin, a protein kinase inhibitor with specificity for PKCdelta, like TPA, caused transformation of c-Src-overexpressing cells. These data suggest that the tumor-promoting effect of phorbol esters is due to depletion of PKCdelta, which has an apparent tumor suppressor function.

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