STXBP1 promotes Weibel-Palade body exocytosis through its interaction with the Rab27A effector Slp4-a.
Vascular endothelial cells contain unique rod-shaped secretory organelles, called Weibel-Palade bodies (WPBs), which contain the hemostatic protein von Willebrand factor (VWF) and a cocktail of angiogenic and inflammatory mediators. We have shown that the Rab27A effector synaptotagmin-like protein 4-a (Slp4-a) plays a critical role in regulating hormone-evoked WPB exocytosis. Using a nonbiased proteomic screen for targets for Slp4-a, we now identify syntaxin-binding protein 1 (STXBP1) and syntaxin-2 and -3 as endogenous Slp4-a binding partners in endothelial cells. Coimmunoprecipitations showed that STXBP1 interacts with syntaxin-2 and -3, but not with syntaxin-4. Small interfering RNA-mediated silencing of STXBP1 expression impaired histamine- and forskolin-induced VWF secretion. To further substantiate the role of STXBP1, we isolated blood outgrowth endothelial cells (BOECs) from an early infantile epileptic encephalopathy type 4 (EIEE4) patient carrying a de novo mutation in STXBP1. STXBP1-haploinsufficient EIEE4 BOECs contained similar numbers of morphologically normal WPBs compared with control BOECs of healthy donors; however, EIEE4 BOECs displayed significantly impaired histamine- and forskolin-stimulated VWF secretion. Based on these findings, we propose that the Rab27A-Slp4-a complex on WPB promotes exocytosis through an interaction with STXBP1, thereby controlling the release of vaso-active substances in the vasculature.
Plasma Proteins, Sanquin-Academisch Medisch Centrum Landsteiner Laboratory, Amsterdam, The Netherlands;
Dorothee
D
van Breevoort
Biomolecular Mass Spectrometry and Proteomics, Medical Research Council Clinical Sciences Centre, London, United Kingdom;
Ambrosius P
AP
Snijders
Physical Biochemistry, Medical Research Council National Institute for Medical Research, London, United Kingdom; Myocardial Function, National Heart and Lung Institute, Imperial College, London, United Kingdom;
Nicola
N
Hellen
Molecular Genetics, University of Antwerp, Belgium;
Sarah
http://orcid.org/0000-0003-2878-1147
S
Weckhuysen
Plasma Proteins, Sanquin-Academisch Medisch Centrum Landsteiner Laboratory, Amsterdam, The Netherlands;
Kathinka W E M
KW
van Hooren
Thrombosis and Hemostasis, Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, The Netherlands;
Jeroen
J
Eikenboom
Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands;
Karine
K
Valentijn
Molecular Cell Biology, Sanquin-Academisch Medisch Centrum Landsteiner Laboratory, Amsterdam, The Netherlands;
Mar
http://orcid.org/0000-0002-1503-433X
M
Fernandez-Borja
Pediatric Neurology, University Hospital, University of Antwerp, Belgium;
Berten
http://orcid.org/0000-0001-7818-0679
B
Ceulemans
Molecular Genetics, University of Antwerp, Belgium;
Peter
P
De Jonghe
Plasma Proteins, Sanquin-Academisch Medisch Centrum Landsteiner Laboratory, Amsterdam, The Netherlands;
Jan
J
Voorberg
Physical Biochemistry, Medical Research Council National Institute for Medical Research, London, United Kingdom; Public Health England, Colindale, London, United Kingdom; and.
Matthew
M
Hannah
Physical Biochemistry, Medical Research Council National Institute for Medical Research, London, United Kingdom; Division of Biomedical Sciences, St. Georges, University of London, United Kingdom.
Tom
T
Carter
Plasma Proteins, Sanquin-Academisch Medisch Centrum Landsteiner Laboratory, Amsterdam, The Netherlands; Physical Biochemistry, Medical Research Council National Institute for Medical Research, London, United Kingdom;
Ruben
http://orcid.org/0000-0002-1205-9689
R
Bierings
D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D005089:Exocytosis; D057809:HEK293 Cells; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D051938:Munc18 Proteins / Q000235:genetics / Q000378:metabolism*; D060066:Protein Interaction Maps; D050765:Qa-SNARE Proteins / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D050827:Syntaxin 1 / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000378:metabolism*; D020302:Weibel-Palade Bodies / Q000378:metabolism*; D020691:rab GTP-Binding Proteins / Q000378:metabolism*; D000075745:rab27 GTP-Binding Proteins
D051938:Munc18 Proteins; D050765:Qa-SNARE Proteins; D034741:RNA, Small Interfering; C497937:STXBP1 protein, human; C512779:SYTL4 protein, human; D050827:Syntaxin 1; D033921:Vesicular Transport Proteins; D000075745:rab27 GTP-Binding Proteins; C066694:RAB27A protein, human; D020691:rab GTP-Binding Proteins