Type II PI4-kinases control Weibel-Palade body biogenesis and von Willebrand factor structure in human endothelial cells.

Mafalda Lopes Da Silva,Janos Kriston-Vizi,J Paul Simons,Raya Al-Shawi,Julia Mössinger,Marie N O'Connor,Daniel F Cutler,Ian J White,Volker Haucke

doi:10.1242/jcs.187864

Abstract

ABSTRACTWeibel-Palade bodies (WPBs) are endothelial storage organelles that mediate the release of molecules involved in thrombosis, inflammation and angiogenesis, including the pro-thrombotic glycoprotein von Willebrand factor (VWF). Although many protein components required for WPB formation and function have been identified, the role of lipids is almost unknown. We examined two key phosphatidylinositol kinases that control phosphatidylinositol 4-phosphate levels at the trans-Golgi network, the site of WPB biogenesis. RNA interference of the type II phosphatidylinositol 4-kinases PI4KIIα and PI4KIIβ in primary human endothelial cells leads to formation of an increased proportion of short WPB with perturbed packing of VWF, as exemplified by increased exposure of antibody-binding sites. When stimulated with histamine, these cells release normal levels of VWF yet, under flow, form very few platelet-catching VWF strings. In PI4KIIα-deficient mice, immuno-microscopy revealed that VWF packaging is also perturbed and these mice exhibit increased blood loss after tail cut compared to controls. This is the first demonstration that lipid kinases can control the biosynthesis of VWF and the formation of WPBs that are capable of full haemostatic function.

Highlights

Endothelial cells line the inner layer of all blood vessels and play a crucial part in regulating vessel formation, function and structure
The expression of the kinases PI4KIIα and PI4KIIβ in human umbilical vein endothelial cells (HUVECs) was confirmed by western blotting (Fig. 1B) and quantitative real-time-PCR (Fig. 1C), and each kinase can be ablated by small interfering RNA (Fig. 1B,C)
RNA interference (RNAi)-meditated ablation of PI4KIIα and/or PI4KIIβ produces shorter Weibel-Palade bodies (WPBs) with abnormally folded von Willebrand factor (VWF) The elongated cigar-like shape of WPBs reflects the presence of VWF correctly folded into the tubules that are crucial to its function (Michaux et al, 2003)

Summary

Introduction

Endothelial cells line the inner layer of all blood vessels and play a crucial part in regulating vessel formation, function and structure. They are adaptable cells that can quickly respond to stimuli in the blood or tissue to control vessel tone, haemostasis and immune responses. Endothelial cells are characterised by the presence of large rod-shaped secretory organelles called Weibel-Palade bodies. In von Willebrand disease, mutations in VWF can perturb formation of WPBs and affect the release of VWF and the string formation, leading to excessive bleeding (Michaux et al, 2003; Sadler, 1998; Valentijn and Eikenboom, 2013), correct biogenesis of WPBs and storage of VWF is crucial for normal haemostasis

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