Identification of protein kinase Calpha as an essential, but not sufficient, cytosolic factor for Ca2+-induced alpha- and dense-core granule secretion in platelets.
Upon activation, platelets release many active substances. Here, we have analyzed the mechanism governing Ca(2+)-induced secretion of von Willebrand factor stored in alpha-granules and 5-hydroxytryptamine in dense-core granules in permeabilized human platelets. Both secretions were dependent on ATP and cytosol. An essential factor for both granule secretions was purified from rat brain cytosol and identified to be protein kinase Calpha (PKCalpha) by partial amino acid sequencing. Purified PKCalpha efficiently stimulated both secretions in the presence of cytosol, whereas PKCalpha alone did not support the secretion of either type of granules, suggesting that PKCalpha is not a sufficient factor. Finally, in human platelet cytosol fractionated by a gel filtration column, the stimulatory activity for dense-core granule secretion paralleled with the concentration of PKC, suggesting that PKC could also be such a stimulatory factor in platelet cytosol. Thus, we identified PKCalpha as an essential, but not sufficient, cytosolic factor for the Ca(2+)-induced secretions of both alpha- and dense-core granules in platelets.
Yoshioka
A
A
Department of Geriatric Medicine, Graduate School of Medicine, Kyoto University, Kyoto, 606-8507, Japan.
Shirakawa
R
R
Nishioka
H
H
Tabuchi
A
A
Higashi
T
T
Ozaki
H
H
Yamamoto
A
A
Kita
T
T
Horiuchi
H
H
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001792:Blood Platelets / Q000201:enzymology* / Q000378:metabolism; D015153:Blotting, Western; D001921:Brain / Q000201:enzymology; D002118:Calcium / Q000378:metabolism*; D002850:Chromatography, Gel; D003600:Cytosol / Q000201:enzymology / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D007527:Isoenzymes / Q000378:metabolism* / Q000502:physiology*; D019169:Jurkat Cells; D008301:Maleimides / Q000494:pharmacology; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism* / Q000502:physiology*; D051571:Protein Kinase C-alpha; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007211:Indoles; D007527:Isoenzymes; D008301:Maleimides; D000255:Adenosine Triphosphate; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; C070515:bisindolylmaleimide I; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium