scholarly journals Plasma membrane phosphatidylinositol (4,5)-bisphosphate promotes Weibel–Palade body exocytosis

2020 ◽  
Vol 3 (11) ◽  
pp. e202000788
Author(s):  
Tu Thi Ngoc Nguyen ◽  
Sophia N Koerdt ◽  
Volker Gerke
Keyword(s):  
Plasma Membrane ◽  
Von Willebrand Factor ◽  
Membrane Lipids ◽  
Phospholipid Composition ◽  
Dominant Negative ◽  
Interaction Sites ◽  
Membrane Contact Sites ◽  
Phospholipase D1 ◽  
Von Willebrand ◽  
Willebrand Factor

Weibel–Palade bodies (WPB) are specialized secretory organelles of endothelial cells that control vascular hemostasis by regulated, Ca2+-dependent exocytosis of the coagulation-promoting von-Willebrand factor. Some proteins of the WPB docking and fusion machinery have been identified but a role of membrane lipids in regulated WPB exocytosis has so far remained elusive. We show here that the plasma membrane phospholipid composition affects Ca2+-dependent WPB exocytosis and von-Willebrand factor release. Phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2] becomes enriched at WPB–plasma membrane contact sites at the time of fusion, most likely downstream of phospholipase D1-mediated production of phosphatidic acid (PA) that activates phosphatidylinositol 4-phosphate (PI4P) 5-kinase γ. Depletion of plasma membrane PI(4,5)P2 or down-regulation of PI4P 5-kinase γ interferes with histamine-evoked and Ca2+-dependent WPB exocytosis and a mutant PI4P 5-kinase γ incapable of binding PA affects WPB exocytosis in a dominant-negative manner. This indicates that a unique PI(4,5)P2-rich environment in the plasma membrane governs WPB fusion possibly by providing interaction sites for WPB-associated docking factors.

Phospholipase D1 is specifically required for regulated secretion of von Willebrand factor from endothelial cells

Blood ◽  
2009 ◽  
Vol 113 (4) ◽  
pp. 973-980 ◽  
Author(s):  
Jennifer Disse ◽  
Nicolas Vitale ◽  
Marie-France Bader ◽  
Volker Gerke
Keyword(s):  
Endothelial Cells ◽  
Plasma Membrane ◽  
Von Willebrand Factor ◽  
Inflammatory Responses ◽  
Endothelial Activation ◽  
Tissue Type ◽  
Type Plasminogen Activator ◽  
Phospholipase D1 ◽  
Von Willebrand ◽  
Willebrand Factor

AbstractEndothelial cells regulate thrombosis, hemostasis, and inflammatory responses by supplying the vasculature with several factors that include procoagulant von Willebrand factor (VWF) and fibrinolytic tissue-type plasminogen activator (tPA). Both proteins can be secreted in a Ca2+-regulated manner after endothelial activation but exhibit opposing physiologic effects. In search for factors that could modulate endothelial responses by selectively affecting the secretion of procoagulant or anticoagulant proteins, we identify here phospholipase D1 (PLD1) as a specific regulator of VWF secretion. PLD1 is translocated to the plasma membrane upon stimulation of endothelial secretion, and this process correlates with the generation of phosphatidic acid (PA) in the plasma membrane. Histamine-evoked secretion of VWF, but not tPA, is inhibited by blocking PLD-mediated production of PA, and this effect can be attributed to PLD1 and not PLD2. Thus, different mechanisms appear to control the agonist-induced secretion of VWF and tPA, with only the former requiring PLD1.

Platelet Density-Dependent Partition of Platelet- von Willebrand Factor Between Alpha Granule and Non-Alpha Granule Pools

1987 ◽  
Vol 58 (03) ◽  
pp. 911-914 ◽  
Author(s):  
R I Parker ◽  
Brenda C Shafer ◽  
H R Gralnick
Keyword(s):  
Plasma Membrane ◽  
Von Willebrand Factor ◽  
Plasma Proteins ◽  
Buoyant Density ◽  
Membrane Permeabilization ◽  
Density Profiles ◽  
Density Dependent ◽  
Plasma Membrane Permeabilization ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryIn this study, we demonstrate that platelets contain a small hut significant amount of platelet-von Willebrand factor(vWf) not associated with α-granules. When platelets free of plasma proteins are exposed to micromolar concentrations ofdigitonin, plasma membrane permeabilization occurs without disruption of platelet granules. Employing this technique, wehave found that upon exposure of a total platelet population to 8 μM digitonin, 5% of total platelet-vWf is released into the supernatant; this occurs without release of (β-TG from α-granules. When platelets of discrete buoyant density profiles are tested, this extragranular platelet-vWf increased with decreasing platelet density. These findings suggest that a redistribution of platelet-vWf from α-granule to non-granule sites occurs coincident with a decrease inplatelet buoyant density.

scholarly journals Fluvastatin inhibits regulated secretion of endothelial cell von Willebrand factor in response to diverse secretagogues

2007 ◽  
Vol 405 (3) ◽  
pp. 597-604 ◽  
Author(s):  
Richard J. Fish ◽  
Hong Yang ◽  
Christelle Viglino ◽  
Raoul Schorer ◽  
Sylvie Dunoyer-Geindre ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Intracellular Calcium ◽  
Von Willebrand Factor ◽  
Small Gtpases ◽  
Dominant Negative ◽  
Calcium Handling ◽  
Regulated Secretion ◽  
Nos Inhibitors ◽  
Von Willebrand ◽  
Willebrand Factor

Regulated secretion of EC (endothelial cell) vWF (von Willebrand factor) is part of the haemostatic response. It occurs in response to secretagogues that raise intracellular calcium or cAMP. Statins are cholesterol-lowering drugs used for the treatment of cardiovascular disease. We studied the effect of fluvastatin on regulated secretion of vWF from HUVEC (human umbilical-vein ECs). Secretion in response to thrombin, a protease-activated receptor-1 agonist peptide, histamine, forskolin and adrenaline (epinephrine) was inhibited. This inhibition was reversed by mevalonate or geranylgeranyl pyrophosphate, and mimicked by a geranylgeranyl transferase inhibitor, demonstrating that the inhibitory mechanism includes inhibition of protein geranylgeranylation. To investigate this mechanism further, calcium handling and NO (nitric oxide) regulation were studied in fluvastatin-treated HUVEC. Intracellular calcium mobilization did not correlate with vWF secretion. Fluvastatin increased eNOS [endothelial NOS (NO synthase)] expression, but NOS inhibitors failed to reverse the effect of fluvastatin on vWF secretion. Exogenous NO did not inhibit thrombin-induced vWF secretion. Many small GTPases are geranylgeranylated and some are activated by secretagogues. We overexpressed DN (dominant negative) Rho GTPases, RhoA, Rac1 and Cdc42 (cell division cycle 42), in HUVEC. DNCdc42 conferred inhibition of thrombin- and forskolin-induced vWF secretion. We conclude that, via inhibition of protein geranylgeranylation, fluvastatin is a broadspectrum inhibitor of regulated vWF secretion. Geranylgeranylated small GTPases with functional roles in regulated secretion, such as Cdc42, are potential targets for the inhibitory activity of fluvastatin.

scholarly journals Promotion of binding of von Willebrand factor to platelet glycoprotein Ib by dimers of ristocetin

1995 ◽  
Vol 306 (2) ◽  
pp. 453-463 ◽  
Author(s):  
M F Hoylaerts ◽  
K Nuyts ◽  
K Peerlinck ◽  
H Deckmyn ◽  
J Vermylen
Keyword(s):  
Von Willebrand Factor ◽  
Specific Binding ◽  
Platelet Glycoprotein ◽  
Glycoprotein Ib ◽  
Natural Interaction ◽  
Charge Neutralization ◽  
Interaction Sites ◽  
Platelet Receptor ◽  
Von Willebrand ◽  
Willebrand Factor

In the absence of high shear forces, the in vitro binding of human von Willebrand factor (vWF) to its platelet receptor glycoprotein Ib (GPIb) can be promoted by two well-characterized mediators, botrocetin and ristocetin. Using purified vWF and GPIb, we have investigated the mechanisms by which ristocetin mediates this binding. Specific binding of vWF monomers to GPIb occurred with a 1:1 stoichiometry, but high-affinity binding required the participation of two ristocetin dimers. Binding was strongly dependent on pH and inhibited by low poly-L-lysine concentrations, indicating ristocetin-dependent charge neutralization during the interaction. With increasing ristocetin concentrations, vWF binding depended progressively less on the involvement of its A1 loop, which is compatible with a model in which the two ristocetin dimers bridge the vWF-GPIb complex on secondary sites. In agreement with this model, the ristocetin-dimer-promoted stabilization of vWF on GPIb was abolished by low concentrations of poly(Pro-Gly-Pro), which is known to complex ristocetin dimers. Mechanistic analysis of the inhibition of vWF binding by the recombinant vWF fragment Leu504-Ser728 (VCL), which covers the entire A1 loop, revealed an affinity of VCL for GPIb comparable with that of the botrocetin-vWF complex for GPIb, and identified a specific but 20-fold lower affinity of VCL in the presence of ristocetin. The proline-rich peptides flanking the vWF A1 loop, Cys474-Val489 and Leu694-Asp709, inhibited vWF binding semispecifically by competitively interfering with the formation of the GPIb-vWF complex rather than by complexation of free ristocetin dimers. In conclusion, ristocetin-promoted binding of vWF to its GPIb receptor results from charge neutralization and interactions involving proline residues in the vicinity of the natural interaction sites present on both GPIb and the A1 domain of vWF.

The role of glycoprotein Ibalpha and von Willebrand factor interaction in stroke development

2010 ◽  
Vol 30 (03) ◽  
pp. 136-138 ◽  
Author(s):  
C. Kleinschnitz ◽  
B. Nieswandt ◽  
G. Stoll
Keyword(s):  
Ischemic Stroke ◽  
Von Willebrand Factor ◽  
Treatment Options ◽  
Serum Levels ◽  
Artery Occlusion ◽  
Downstream Signaling ◽  
Phospholipase D1 ◽  
Induced Changes ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryIschaemic stroke is a devastating disease with limited treatment options due to numerous uncertainties regarding the underlying pathophysiology. The contribution of glycoprotein (GP)Ibα and von Willebrand factor (VWF) in stroke development has only recently been established in mice. Complete blockade of GPIbα led to a significant reduction of infarct volumes in mice undergoing one hour of transient middle cerebral artery occlusion (tMCAO). High shear-induced changes in VWF confirmation are a prerequisite for VWF binding to collagen and GPIbα expressed on platelets. Importantly, transgenic VWF−/−mice were similarly protected against ischemic stroke after tMCAO, and hydrodynamic injection of a VWF-encoding plasmid restored VWF serum levels and the susceptibility towards stroke. Secreted VWF is rapidly cleaved by ADAMTS13. Accordingly, ADAMTS13 deficient mice developed larger infarction after tMCAO, while infusion of recombinant ADAMTS13 into wild-type mice was strokeprotective. In conclusion, there is compelling evidence that GPIbα/VWF interactions and downstream signaling via phospholipase D1 (PLD1) provide new therapeutic targets in ischemic stroke.

Release from Intracellular Stores Is Responsible for Calcium Signaling with von Willebrand Factor in Human Platelets

2002 ◽  
Vol 87 (04) ◽  
pp. 699-705 ◽  
Author(s):  
Elizabeth Milner ◽  
Qi Zheng ◽  
John Kermode
Keyword(s):  
Plasma Membrane ◽  
Intracellular Calcium ◽  
Von Willebrand Factor ◽  
Extracellular Calcium ◽  
Lag Phase ◽  
Minor Role ◽  
Calcium Stores ◽  
Intracellular Calcium Stores ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryInteraction of von Willebrand factor (VWF) with the platelet promotes hemostasis upon vascular injury. Such interaction raises intracellular free calcium concentration ([Ca2+]i) and induces platelet activation. The platelet [Ca2+]i increase is generally attributed to influx across the plasma membrane. The present study defined the contribution of intracellular calcium stores. Platelet [Ca2+]i was monitored with Fura-PE3. Ristocetin-mediated binding of VWF transiently elevated [Ca2+]i after a lag phase. Studies with 63 healthy donors consistently revealed a VWF-induced platelet [Ca2+]i signal in the absence of extracellular calcium; there was only modest enhancement with extracellular calcium. Blockade of plasma membrane calcium channels did not diminish the signal, whereas depletion or blockade of the intracellular calcium stores abolished it. These findings imply that release from intracellular stores is responsible for the VWF-induced platelet [Ca2+]i increase. Influx across the plasma membrane plays no more than a minor role, probably representing “capacitative entry” to refill the intracellular stores.

The dominant-negative von Willebrand factor gene deletion p.P1127_C1948delinsR: molecular mechanism and modulation

Blood ◽  
2010 ◽  
Vol 116 (24) ◽  
pp. 5371-5376 ◽  
Author(s):  
Caterina Casari ◽  
Mirko Pinotti ◽  
Stefano Lancellotti ◽  
Elena Adinolfi ◽  
Alessandra Casonato ◽  
...  
Keyword(s):  
Von Willebrand Factor ◽  
Molecular Mechanisms ◽  
Von Willebrand Disease ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Wild Type ◽  
Collagen Binding ◽  
Cellular Models ◽  
Von Willebrand ◽  
Willebrand Factor

AbstractUnderstanding molecular mechanisms in the dominant inheritance of von Willebrand disease would improve our knowledge of pathophysiologic processes underlying its prevalence. Cellular models of severe type 2 von Willebrand disease, caused by a heterozygous deletion in the von Willebrand factor (VWF) gene, were produced to investigate the altered biosynthesis. Coexpression of the wild-type and in-frame deleted (p.P1127_C1948delinsR) VWF forms impaired protein secretion, high molecular weight multimer formation and function (VWF collagen-binding 1.9% ± 0.5% of wild-type), which mimicked the patient's phenotype. mRNA, protein, and cellular studies delineated the highly efficient dominant-negative mechanism, based on the key role of heterodimers as multimer terminators. The altered VWF, synthesized in large amounts with the correctly encoded “cysteine knot” domain, formed heterodimers and heterotetramers with wild-type VWF, in addition to deleted homodimers. Impaired multimerization was associated with reduced amounts of VWF in late endosomes. Correction of the dominant-negative effect was explored by siRNAs targeting the mRNA breakpoint, which selectively inhibited the in-frame deleted VWF expression. Although the small amount of the deleted protein synthesized after inhibition still exerted dominant, even though weakened, negative effects, the siRNA treatment restored secretion of large multimers with improved function (VWF collagen-binding 28.0% ± 3.3% of wild-type).

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