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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2006;26:49.)
© 2006 American Heart Association, Inc.

Vascular Biology

Dynein–Dynactin Complex Mediates Protein Kinase A–Dependent Clustering of Weibel-Palade Bodies in Endothelial Cells

Mariska G. Rondaij; Ruben Bierings; Astrid Kragt; Karina A. Gijzen; Erica Sellink; Jan A. van Mourik; Mar Fernandez-Borja; Jan Voorberg

From the Departments of Plasma Proteins (M.G.R., R.B., A.K., K.A.G., E.S., J.A.v.M., J.V.) and Molecular Cell Biology (M.F-B.), Sanquin Research and Landsteiner Laboratory, and the Department of Vascular Medicine (J.A.v.M.), Academic Medical Centre, University of Amsterdam, The Netherlands.

Correspondence to Jan Voorberg, Department of Plasma Proteins, Sanquin Research at CLB and Landsteiner Laboratory, Academic Medical Centre, University of Amsterdam, Plesmanlaan 125, 1066 CX Amsterdam, The Netherlands. E-mail j.voorberg{at}sanquin.nl

Objective— Perinuclear clustering is observed for several different organelles and illustrates dynamic regulation of the secretory pathway and organelle distribution. Previously, we observed that a subset of Weibel-Palade bodies (WPBs), endothelial cell–specific storage organelles, undergo centralization when endothelial cells are stimulated with cAMP-raising agonists of von Willebrand factor (vWF) secretion. In this study, we investigated this phenomenon of WPB clustering in more detail.

Methods and Results— Our results demonstrate that the clustered WPBs are localized at the microtubule organizing center and that cluster formation depends on an intact microtubule network. Disruption of the microtubules by nocodazole completely abolished clustering, whereas treatment with the actin depolymerizing compound cytochalasin B had no effect on WPB clustering. Interfering with the dynein–dynactin interaction by overexpression of the p50 dynamitin subunit or the CC1 domain of the p150^glued subunit of the dynactin complex completely inhibited perinuclear clustering of WPBs, suggesting that dynein activity mediates this process. Furthermore, we found that inhibition of dephosphorylation resulted in an increase in clustering, whereas inhibition of protein kinase A (PKA) markedly reduced WPB clustering.

Conclusions— These results suggest that perinuclear clustering of WPBs involves PKA-dependent regulation of the dynein–dynactin complex. Endothelial cell stimulation with epinephrine results in retrograde movement of a subset of WPBs to the microtubule organizing center. This minus-end directed transport requires an intact microtubular network and is mediated by the motor protein dynein. Together, our results suggest that epinephrine-induced clustering of WPBs involves PKA-dependent regulation of the dynein–dynactin complex.

Perinuclear clustering is observed for several different organelles and illustrates dynamic regulation of the secretory pathway and organelle distribution. Previously, we observed that a subset of Weibel-Palade bodies (WPBs), endothelial cell–specific storage organelles, undergo centralization when endothelial cells are stimulated with cAMP-raising agonists of von Willebrand factor secretion. In this study, we investigated this phenomenon of WPB clustering in more detail.

Key Words: organelle trafficking • dynein • Weibel-Palade bodies • PKA • endothelial cells

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