Phosphatidylinositol transfer proteins and protein kinase C make separate but non-interacting contributions to the phosphorylation state necessary for secretory competence in rat mast cells

2001 ◽  
Vol 356 (1) ◽  
pp. 287-296 ◽  
Author(s):  
Jef A. PINXTEREN ◽  
Bastien D. GOMPERTS ◽  
Danise ROGERS ◽  
Scott E. PHILLIPS ◽  
Peter E. R. TATHAM ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Mast Cells ◽  
Protein Kinase ◽  
Aminoglycoside Antibiotic ◽  
Pleckstrin Homology Domain ◽  
Kinase C ◽  
Streptolysin O ◽  
Regulatory Machinery ◽  
Phosphatidylinositol Transfer ◽  
Phosphatidylinositol Transfer Proteins

Mast cells permeabilized by streptolysin O undergo exocytosis when stimulated with Ca2+ and guanosine 5′-[γ-thio]triphosphate but become progressively refractory to this stimulus if it is delayed. This run-down of responsiveness occurs over a period of 20–30min, during which the cells leak soluble and tethered proteins. We show here that withdrawal of ATP during the process of run-down is strongly inhibitory but that as little as 25μM ATP can extend responsiveness significantly; this effect is maximal at 50μM. When phosphatidylinositol transfer proteins (PITPs) are provided to cells at the time of permeabilization, run-down is retarded. We conclude that in the presence of ATP they convey substrates for phosphorylation that are essential for exocytosis and thus interact with the regulatory machinery. Furthermore, we show that PITPα and PITPβ have additive effects in this mechanism, suggesting that they are not functionally redundant. Alternatively, secretion from run-down cells can be inhibited by the aminoglycoside antibiotic neomycin, which is understood to bind to phosphoinositide headgroups, and by a PH (pleckstrin homology) domain polypeptide that binds phosphoinositides. The apparent displacement of neomycin by exogenous PITPs suggests that these proteins screen essential lipids. Secretion from run-down cells is also inhibited by 1-O-hexadecyl-2-O-methyl-rac-glycerol (AMG-C16), an inhibitor of protein kinase C. The lack of synergy between neomycin and AMG-C16 suggests that protein kinase C independently provides a second essential component through protein phosphorylation and that there are two independent phosphorylation pathways necessary for secretion competence.

scholarly journals The Streptococcal Exotoxin Streptolysin O Activates Mast Cells To Produce Tumor Necrosis Factor Alpha by p38 Mitogen-Activated Protein Kinase- and Protein Kinase C-Dependent Pathways

2003 ◽  
Vol 71 (11) ◽  
pp. 6171-6177 ◽  
Author(s):  
Michael Stassen ◽  
Christian Müller ◽  
Christoph Richter ◽  
Christine Neudörfl ◽  
Lothar Hültner ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Mast Cells ◽  
Protein Kinase ◽  
Map Kinase ◽  
P38 Map Kinase ◽  
Necrosis Factor Alpha ◽  
Kinase C ◽  
Streptolysin O ◽  
Tnf Α ◽  
Mitogen Activated Protein

ABSTRACT Streptolysin O (SLO), a major virulence factor of pyogenic streptococci, binds to cholesterol in the membranes of eukaryotic cells and oligomerizes to form large transmembrane pores. While high toxin doses are rapidly cytocidal, low doses are tolerated because a limited number of lesions can be resealed. Here, we report that at sublethal doses, SLO activates primary murine bone marrow-derived mast cells to degranulate and to rapidly induce or enhance the production of several cytokine mRNAs, including tumor necrosis factor alpha (TNF-α). Mast cell-derived TNF-α plays an important protective role in murine models of acute inflammation, and the production of this cytokine was analyzed in more detail. Release of biologically active TNF-α peaked ∼4 h after stimulation with SLO. Production of TNF-α was blunted upon depletion of protein kinase C by pretreatment of the cells with phorbol-12 myristate-13 acetate. Transient permeabilization of mast cells with SLO also led to the activation of the stress-activated protein kinases p38 mitogen-activated protein (MAP) kinase and c-jun N-terminal kinase (JNK), and inhibition of p38 MAP kinase markedly reduced production of TNF-α. In contrast, secretion of preformed granule constituents triggered by membrane permeabilization was not dependent on p38 MAP kinase or on protein kinase C. Thus, transcriptional activation of mast cells following transient permeabilization might contribute to host defense against infections via the beneficial effects of TNF-α. However, hyperstimulation of mast cells might also lead to overproduction of TNF-α, which would then promote the development of toxic streptococcal syndromes.

scholarly journals Two G-proteins act in series to control stimulus-secretion coupling in mast cells: use of neomycin to distinguish between G-proteins controlling polyphosphoinositide phosphodiesterase and exocytosis.

1987 ◽  
Vol 105 (6) ◽  
pp. 2745-2750 ◽  
Author(s):  
S Cockcroft ◽  
T W Howell ◽  
B D Gomperts
Keyword(s):  
Protein Kinase C ◽  
Mast Cells ◽  
Protein Kinase ◽  
G Proteins ◽  
Kinase C ◽  
Streptolysin O ◽  
In Series ◽  
Sparing Effect ◽  
Necessary And Sufficient ◽  
Gamma S

Provision of GTP (or other nucleotides capable of acting as ligands for activation of G-proteins) together with Ca2+ (at micromolar concentrations) is both necessary and sufficient to stimulate exocytotic secretion from mast cells permeabilized with streptolysin-O. GTP and its analogues, through their interactions with Gp, also activate polyphosphoinositide-phosphodiesterase (PPI-pde generating inositol 1,4,5-trisphosphate and diglyceride [DG]). We have used mast cells labeled with [3H]inositol to test whether the requirement for GTP in exocytosis is an expression of Gp activity through the generation of DG and consequent activation of protein kinase C, or whether GTP is required at a later stage in the stimulus secretion sequence. Neomycin (0.3 mM) inhibits activation of PPI-pde, but maximal secretion due to optimal concentrations of guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-S) can still be evoked in its presence. When ATP is also provided the concentration requirement for GTP-gamma-S in support of exocytosis is reduced. This sparing effect of ATP is nullified when the PPI-pde reaction is inhibited by neomycin. We argue that the sparing effect of ATP occurs as a result of enhancement of DG production and through its action as a phosphoryl donor in the reactions catalyzed by protein kinase C.

Stratifin, a keratinocyte specific 14–3-3 protein, harbors a pleckstrin homology (PH) domain and enhances protein kinase C activity

1995 ◽  
Vol 108 (11) ◽  
pp. 3569-3579
Author(s):  
E. Dellambra ◽  
M. Patrone ◽  
B. Sparatore ◽  
A. Negri ◽  
F. Ceciliani ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Terminal Differentiation ◽  
Pleckstrin Homology Domain ◽  
Ph Domain ◽  
Pleckstrin Homology ◽  
Protein Database ◽  
Kinase C ◽  
Human Keratinocyte ◽  
Protein Kinase C Activity

The intrinsic signal(s) responsible for the onset of human keratinocyte terminal differentiation is not yet fully understood. Evidence has been recently accumulated linking the phospholipase-mediated activation of protein kinase C to the coordinate changes in gene expression occurring during keratinocyte terminal differentiation. Here we report the purification of a keratinocyte-derived protein enhancing protein kinase C enzymatic activity. The stimulator eluted as a peak with estimated molecular mass of approximately 70 kDa, while analysis by SDS-PAGE showed a 30 kDa protein migrating as a distinct doublet, suggesting the formation of a 30 kDa homodimer. The amino acid sequence analysis allowed the unambigous identification of the protein kinase C stimulator as a mixture of the highly homologous sigma (stratifin) and zeta isoforms of 14–3-3 proteins, which are homodimers of identical 30 kDa subunits. Mono Q anion exchange chromatography and immunoblot analysis further confirmed that stratifin enhances protein kinase C activity. Stratifin was originally sequenced from a human keratinocyte protein database, but its function was unknown. The pleckstrin homology domain has been recently related to protein translocation to the cell membrane as well as to functional interactions of intracellular proteins involved in signal transduction. We show here that stratifin (and 14–3-3 zeta) harbors a pleckstrin homology domain, and the consequent functional implications will be discussed.

Ca2+ and protein kinase C activation of mucin granule exocytosis in permeabilized SPOC1 cells

1998 ◽  
Vol 275 (1) ◽  
pp. C285-C292 ◽  
Author(s):  
C. E. Scott ◽  
Lubna H. Abdullah ◽  
C. William Davis
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Fold Increase ◽  
Mucin Secretion ◽  
Granule Exocytosis ◽  
Kinase C ◽  
Streptolysin O ◽  
Intracellular Ca ◽  
Mucin Release ◽  
Protein Kinase C Activation

Mucin secretion by airway goblet cells is under the control of apical P2Y2, phospholipase C-coupled purinergic receptors. In SPOC1 cells, the mobilization of intracellular Ca2+ by ionomycin or the activation of protein kinase C (PKC) by phorbol 12-myristate 13-acetate (PMA) stimulates mucin secretion in a fully additive fashion [L. H. Abdullah, J. D. Conway, J. A. Cohn, and C. W. Davis. Am. J. Physiol. 273 ( Lung Cell. Mol. Physiol. 17): L201–L210, 1997]. This apparent independence between PKC and Ca2+ in the stimulation of mucin secretion was tested in streptolysin O-permeabilized SPOC1 cells. These cells were fully competent to secrete mucin when Ca2+ was elevated from 100 nM to 3.1 μM for 2 min following permeabilization; the Ca2+EC50 was 2.29 ± 0.07 μM. Permeabilized SPOC1 cells were exposed to PMA or 4α-phorbol at Ca2+ activities ranging from 10 nM to 10 μM. PMA, but not 4α-phorbol, increased mucin release at all Ca2+ activities tested: at 10 nM Ca2+ mucin release was 2.1-fold greater than control and at 4.7 μM Ca2+ mucin release was maximal (3.6-fold increase). PMA stimulated 27% more mucin release at 4.7 μM than at 10 nM Ca2+. Hence, SPOC1 cells possess Ca2+-insensitive, PKC-dependent, and Ca2+-dependent PKC-potentiated pathways for mucin granule exocytosis.

scholarly journals Steel Factor Enhances Supraoptimal Antigen-Induced IL-6 Production from Mast Cells via Activation of Protein Kinase C-β

2009 ◽  
Vol 182 (12) ◽  
pp. 7897-7905 ◽  
Author(s):  
Kerstin Fehrenbach ◽  
Eva Lessmann ◽  
Carolin N. Zorn ◽  
Marcel Kuhny ◽  
Gordon Grochowy ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Mast Cells ◽  
Protein Kinase ◽  
Steel Factor ◽  
Kinase C

scholarly journals Suppression of Microphthalmia Transcriptional Activity by Its Association with Protein Kinase C-interacting Protein 1 in Mast Cells

1999 ◽  
Vol 274 (48) ◽  
pp. 34272-34276 ◽  
Author(s):  
Ehud Razin ◽  
Zhao Cheng Zhang ◽  
Hovav Nechushtan ◽  
Shahar Frenkel ◽  
Yu-Nee Lee ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Mast Cells ◽  
Protein Kinase ◽  
Transcriptional Activity ◽  
Interacting Protein ◽  
Kinase C
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