Sphingomyelinase Disables Inactivation in Endogenous PIEZO1 Channels

Jian Shi,Adam J Hyman,Dario De Vecchis,Jiehan Chong,Laeticia Lichtenstein,T Simon Futers,Myriam Rouahi,Anne Negre Salvayre,Nathalie Auge,Antreas C Kalli,David J Beech

doi:10.1016/j.celrep.2020.108225

Abstract

SummaryEndogenous PIEZO1 channels of native endothelium lack the hallmark inactivation often seen when these channels are overexpressed in cell lines. Because prior work showed that the force of shear stress activates sphingomyelinase in endothelium, we considered if sphingomyelinase is relevant to endogenous PIEZO1. Patch clamping was used to quantify PIEZO1-mediated signals in freshly isolated murine endothelium exposed to the mechanical forces caused by shear stress and membrane stretch. Neutral sphingomyelinase inhibitors and genetic disruption of sphingomyelin phosphodiesterase 3 (SMPD3) cause PIEZO1 to switch to profoundly inactivating behavior. Ceramide (a key product of SMPD3) rescues non-inactivating channel behavior. Its co-product, phosphoryl choline, has no effect. In contrast to ceramide, sphingomyelin (the SMPD3 substrate) does not affect inactivation but alters channel force sensitivity. The data suggest that sphingomyelinase activity, ceramide, and sphingomyelin are determinants of native PIEZO gating that enable sustained activity.

Highlights

Shear stress is a frictional force that arises when fluid flows along a cellular surface: it impacts many aspects of biology (Mammoto and Ingber, 2010)
We investigated the relevance of sphingomyelinase to endothelial PIEZO1 and sought to determine whether it might explain some of the distinct properties of native endothelial PIEZO1 channels
We investigated whether neutral sphingomyelinase, sphingomyelin phosphodiesterase 3 (SMPD3), regulates PIEZO1 channels and investigated the underlying cellular mechanism in endothelial cells

Summary

Introduction

Shear stress is a frictional force that arises when fluid flows along a cellular surface: it impacts many aspects of biology (Mammoto and Ingber, 2010). Endothelial PIEZO1 has been found to be important in numerous vascular and other cardiovascular biology that depends on mechanical force (Beech and Kalli, 2019), including vascular maturation in the embryo (Li et al, 2014), blood pressure regulation (Rode et al, 2017), vascular permeability (Friedrich et al, 2019), atherosclerosis (Albarran-Juarez et al, 2018), response to shear stress in endothelial cells in vitro and in vivo (Beech and Kalli, 2019; Li et al, 2014; Rode et al, 2017), lymphatic structure, and the disease of generalized lymphatic dysplasia (Fotiou et al, 2015). It raises the possibility that there is a process in endothelial cells that disables the intrinsic fast inactivation gate of PIEZO1 to enable compatibility with the sustained requirements of vascular flow sensing

Results

Discussion

Conclusion