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J. Biol. Chem., Vol. 280, Issue 24, 22827-22830, June 17, 2005

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 280/24/22827    most recent M412547200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Hannah, M. J. Articles by Carter, T. Search for Related Content PubMed PubMed Citation Articles by Hannah, M. J. Articles by Carter, T. Social Bookmarking                      What's this?

Differential Kinetics of Cell Surface Loss of von Willebrand Factor and Its Propolypeptide after Secretion from Weibel-Palade Bodies in Living Human Endothelial Cells^*

Matthew J. Hannah, Paul Skehel, Muriel Erent, Laura Knipe, David Ogden, and Tom Carter¶

From the National Institute for Medical Research, Mill Hill, London NW7 1AA, United Kingdom and Department of Neuroscience, University of Edinburgh, Edinburgh EH8 9JZ, United Kingdom

The time course for cell surface loss of von Willebrand factor (VWF) and the propolypeptide of VWF (proregion) following exocytosis of individual Weibel-Palade bodies (WPBs) from single human endothelial cells was analyzed. Chimeras of enhanced green fluorescent protein (EGFP) and full-length pre-pro-VWF (VWF-EGFP) or the VWF propolypeptide (proregion-EGFP) were made and expressed in human umbilical vein endothelial cells. Expression of VWF-EGFP or proregion-EGFP resulted in fluorescent rod-shaped organelles that recruited the WPB membrane markers P-selectin and CD63. The WPB secretagogue histamine evoked exocytosis of these fluorescent WPBs and extracellular release of VWF-EGFP or proregion-EGFP. Secreted VWF-EGFP formed distinctive extracellular patches of fluorescence that were labeled with an extracellular antibody to VWF. The half-time for dispersal of VWF-EGFP from extracellular patches was 323.5 ± 146.2 s (±S.D., n = 20 WPBs). In contrast, secreted proregion-EGFP did not form extracellular patches but dispersed rapidly from its site of release. The half-time for dispersal of proregion-EGFP following WPB exocytosis was 2.98 ± 1.88 s (±S.D., n = 32 WPBs). The slow rate of loss of VWF-EGFP is consistent with the adhesive nature of this protein for the endothelial membrane. The much faster rate of loss of proregion-EGFP indicates that this protein does not interact strongly with extracellular VWF or the endothelial membrane and consequently may not play an adhesive role at the endothelial cell surface.

Received for publication, November 5, 2004 , and in revised form, March 15, 2005.

^* This work was supported in part by the British Heart Foundation and the Medical Research Council. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains Supplemental Figs. 1 and 2 and Movies 1–4.

^¶ Held a British Heart Foundation Basic Sciences University Lectureship. To whom correspondence should be addressed. Tel.: 44-207-816-2743; Fax: 44-207-906-4477; E-mail: tcarter{at}nimr.mrc.ac.uk.

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