Calcium/calmodulin transduces thrombin-stimulated secretion: studies in intact and minimally permeabilized human umbilical vein endothelial cells

doi:10.1083/jcb.118.6.1501

This Article

	Full Text (PDF, 1557K)
	Alert me when this article is cited
	Citation Map

Services

	Email this article
	Similar articles in this journal
	Similar articles in PubMed
	Alert me to new content in the JCB
	Download to citation manager

Citing Articles

	Citing Articles via HighWire
	Citing Articles via CrossRef
	Citing Articles via Google Scholar

Google Scholar

	Articles by Birch, K. A.
	Articles by Ewenstein, B. M.
	Search for Related Content

PubMed

	PubMed Citation
	Articles by Birch, K. A.
	Articles by Ewenstein, B. M.


Social Bookmarking

	What's this?

ARTICLES

Calcium/calmodulin transduces thrombin-stimulated secretion: studies in intact and minimally permeabilized human umbilical vein endothelial cells

KA Birch, JS Pober, GB Zavoico, AR Means and BM Ewenstein
Department of Microbiology, Biological Chemistry, and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115.

Thrombin stimulates cultured endothelial cells (EC) to secrete stored von Willebrand factor (vWF), but the signal transduction pathways are poorly defined. Thrombin is known to elevate the concentration of intracellular calcium ([Ca2+]i) and to activate protein kinase C (PKC) in EC. Since both calcium ionophores and phorbol esters release vWF, both second messenger pathways have been postulated to participate in vWF secretion in response to naturally occurring agonists. We find that in intact human EC, vWF secretion stimulated by either thrombin or by a thrombin receptor activating peptide, TR(42-55), can be correlated with agonist-induced elevations of [Ca2+]i. Further evidence implicating calcium in the signal transduction pathway is suggested by the finding that MAPTAM, a cell-permeant calcium chelator, in combination with the extracellular calcium chelator EGTA, can inhibit thrombin-stimulated secretion. In contrast, the observation that staurosporine (a pharmacological inhibitor of PKC) blocks phorbol ester- but not thrombin-stimulated secretion provides evidence against PKC-mediated signal transduction. To examine further the signal transduction pathway initiated by thrombin, we developed novel conditions for minimal permeabilization of EC with saponin (4-8 micrograms/ml for 5-15 min at 37 degrees C) which allow the introduction of small extracellular molecules without the loss of large intracellular proteins and which retain thrombin-stimulated secretion. These minimally permeabilized cells secrete vWF in response to exogenous calcium, and EGTA blocks thrombin-induced secretion. Moreover, in these cells, thrombin- stimulated secretion is blocked by a calmodulin-binding inhibitory peptide but not by a PKC inhibitory peptide. Taken together, these findings demonstrate that thrombin-stimulated vWF secretion is transduced by a rise in [Ca2+]i and provide the first evidence for the role of calmodulin in this process.
Add to CiteULike CiteULike Add to Complore Complore Add to Connotea Connotea Add to Del.icio.us Del.icio.us Add to Digg Digg Add to Reddit Reddit Add to Technorati Technorati What's this?

This article has been cited by other articles:

Rondaij, M. G., Bierings, R., van Agtmaal, E. L., Gijzen, K. A., Sellink, E., Kragt, A., Ferguson, S. S. G., Mertens, K., Hannah, M. J., van Mourik, J. A., Fernandez-Borja, M., Voorberg, J. (2008). Guanine exchange factor RalGDS mediates exocytosis of Weibel-Palade bodies from endothelial cells. Blood 112: 56-63 [Abstract] [Full Text]
Maliba, R., Brkovic, A., Neagoe, P.-E., Villeneuve, L. R., Sirois, M. G. (2008). Angiopoietin-mediated endothelial P-selectin translocation: cell signaling mechanisms. J. Leukoc. Biol. 83: 352-360 [Abstract] [Full Text]
Csortos, C., Kolosova, I., Verin, A. D. (2007). Regulation of vascular endothelial cell barrier function and cytoskeleton structure by protein phosphatases of the PPP family. Am. J. Physiol. Lung Cell. Mol. Physiol. 293: L843-L854 [Abstract] [Full Text]
Erent, M., Meli, A., Moisoi, N., Babich, V., Hannah, M. J., Skehel, P., Knipe, L., Zupancic, G., Ogden, D., Carter, T. (2007). Rate, extent and concentration dependence of histamine-evoked Weibel Palade body exocytosis determined from individual fusion events in human endothelial cells. J. Physiol. 583: 195-212 [Abstract] [Full Text]
Zhou, C., Chen, H., Lu, F., Sellak, H., Daigle, J. A., Alexeyev, M. F., Xi, Y., Ju, J., van Mourik, J. A., Wu, S. (2007). Cav3.1 ({alpha}1G) controls von Willebrand factor secretion in rat pulmonary microvascular endothelial cells. Am. J. Physiol. Lung Cell. Mol. Physiol. 292: L833-L844 [Abstract] [Full Text]
Rondaij, M. G., Bierings, R., Kragt, A., van Mourik, J. A., Voorberg, J. (2006). Dynamics and Plasticity of Weibel-Palade Bodies in Endothelial Cells. Arterioscler. Thromb. Vasc. Bio. 26: 1002-1007 [Abstract] [Full Text]
Cernuda-Morollon, E., Ridley, A. J. (2006). Rho GTPases and Leukocyte Adhesion Receptor Expression and Function in Endothelial Cells. Circ. Res. 98: 757-767 [Abstract] [Full Text]
Rondaij, M. G., Bierings, R., Kragt, A., Gijzen, K. A., Sellink, E., van Mourik, J. A., Fernandez-Borja, M., Voorberg, J. (2006). Dynein-Dynactin Complex Mediates Protein Kinase A-Dependent Clustering of Weibel-Palade Bodies in Endothelial Cells. Arterioscler. Thromb. Vasc. Bio. 26: 49-55 [Abstract] [Full Text]
Bhatia, R., Matsushita, K., Yamakuchi, M., Morrell, C. N., Cao, W., Lowenstein, C. J. (2004). Ceramide Triggers Weibel-Palade Body Exocytosis. Circ. Res. 95: 319-324 [Abstract] [Full Text]
Rondaij, M. G., Sellink, E., Gijzen, K. A., ten Klooster, J. P., Hordijk, P. L., van Mourik, J. A., Voorberg, J. (2004). Small GTP-Binding Protein Ral Is Involved in cAMP-Mediated Release of von Willebrand Factor From Endothelial Cells. Arterioscler. Thromb. Vasc. Bio. 24: 1315-1320 [Abstract] [Full Text]
Wu, S., Haynes, J. Jr, Taylor, J. T., Obiako, B. O., Stubbs, J. R., Li, M., Stevens, T. (2003). Cav3.1 ({alpha}1G) T-Type Ca2+ Channels Mediate Vaso-Occlusion of Sickled Erythrocytes in Lung Microcirculation. Circ. Res. 93: 346-353 [Abstract] [Full Text]
Romani de Wit, T., Rondaij, M. G., Hordijk, P. L., Voorberg, J., van Mourik, J. A. (2003). Real-Time Imaging of the Dynamics and Secretory Behavior of Weibel-Palade Bodies. Arterioscler. Thromb. Vasc. Bio. 23: 755-761 [Abstract] [Full Text]
Burgoyne, R. D., Morgan, A. (2003). Secretory Granule Exocytosis. Physiol. Rev. 83: 581-632 [Abstract] [Full Text]
Klarenbach, S. W., Chipiuk, A., Nelson, R. C., Hollenberg, M. D., Murray, A. G. (2003). Differential Actions of PAR2 and PAR1 in Stimulating Human Endothelial Cell Exocytosis and Permeability: The Role of Rho-GTPases. Circ. Res. 92: 272-278 [Abstract] [Full Text]
Nilius, B., Droogmans, G. (2001). Ion Channels and Their Functional Role in Vascular Endothelium. Physiol. Rev. 81: 1415-1459 [Abstract] [Full Text]
Methia, N., Andre, P., Denis, C. V., Economopoulos, M., Wagner, D. D. (2001). Localized reduction of atherosclerosis in von Willebrand factor-deficient mice. Blood 98: 1424-1428 [Abstract] [Full Text]
de Leeuw, H. P. J. C., Fernandez-Borja, M., Reits, E. A. J., Romani de Wit, T., Wijers-Koster, P. M., Hordijk, P. L., Neefjes, J., van Mourik, J. A., Voorberg, J. (2001). Small GTP-Binding Protein Ral Modulates Regulated Exocytosis of von Willebrand Factor by Endothelial Cells. Arterioscler. Thromb. Vasc. Bio. 21: 899-904 [Abstract] [Full Text]
Vischer, U. M., Barth, H., Wollheim, C. B. (2000). Regulated von Willebrand Factor Secretion Is Associated With Agonist-Specific Patterns of Cytoskeletal Remodeling in Cultured Endothelial Cells. Arterioscler. Thromb. Vasc. Bio. 20: 883-891 [Abstract] [Full Text]
Chen, Y. A., Duvvuri, V., Schulman, H., Scheller, R. H. (1999). Calmodulin and Protein Kinase C Increase Ca2+-stimulated Secretion by Modulating Membrane-attached Exocytic Machinery. J. Biol. Chem. 274: 26469-26476 [Abstract] [Full Text]
Vischer, U. M., Wollheim, C. B. (1998). Purine Nucleotides Induce Regulated Secretion of von Willebrand Factor: Involvement of Cytosolic Ca2+ and Cyclic Adenosine Monophosphate-Dependent Signaling in Endothelial Exocytosis. Blood 91: 118-127 [Abstract] [Full Text]
van den Eijnden-Schrauwen, Y., Atsma, D. E., Lupu, F., de Vries, R. E. M., Kooistra, T., Emeis, J. J. (1997). Involvement of Calcium and G Proteins in the Acute Release of Tissue-Type Plasminogen Activator and von Willebrand Factor From Cultured Human Endothelial Cells. Arterioscler. Thromb. Vasc. Bio. 17: 2177-2187 [Abstract] [Full Text]
Galbusera, M., Zoja, C., Donadelli, R., Paris, S., Morigi, M., Benigni, A., Figliuzzi, M., Remuzzi, G., Remuzzi, A. (1997). Fluid Shear Stress Modulates von Willebrand Factor Release From Human Vascular Endothelium. Blood 90: 1558-1564 [Abstract] [Full Text]
Datta, Y. H., Romano, M., Jacobson, B. C., Golan, D. E., Serhan, C. N., Ewenstein, B. M. (1995). Peptido-Leukotrienes Are Potent Agonists of von Willebrand Factor Secretion and P-Selectin Surface Expression in Human Umbilical Vein Endothelial Cells. Circulation 92: 3304-3311 [Abstract] [Full Text]
Chen, Y. A., Scales, S. J., Duvvuri, V., Murthy, M., Patel, S. M., Schulman, H., Scheller, R. H. (2001). Calcium Regulation of Exocytosis in PC12 Cells. J. Biol. Chem. 276: 26680-26687 [Abstract] [Full Text]