S1P induces proliferation of pulmonary artery smooth muscle cells by promoting YAP-induced Notch3 expression and activation

Jian Wang,Xin Yan,Wei Feng,Qingting Wang,Wenhua Shi,Limin Chai,Qianqian Zhang,Yuqian Chen,Jin Liu,Zhan Qu,Xinming Xie,Manxiang Li

doi:10.1016/j.jbc.2021.100599

Abstract

Sphingosine-1-phosphate (S1P), a natural multifunctional phospholipid, is highly increased in plasma from patients with pulmonary arterial hypertension and mediates proliferation of pulmonary artery smooth muscle cells (PASMCs) by activating the Notch3 signaling pathway. However, the mechanisms underpinning S1P-mediated induction of PASMCs proliferation remain unclear. In this study, using biochemical and molecular biology approaches, RNA interference and gene expression analyses, 5′-ethynyl-2′-deoxyuridine incorporation assay, and 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay, we demonstrated that S1P promoted the activation of signal transducers and activators of transcription 3 (STAT3) through sphingosine-1-phosphate receptor 2 (S1PR2), and subsequently upregulated the expression of the microRNA miR-135b, which further reduced the expression of E3 ubiquitin ligase β-transduction repeat-containing protein and led to a reduction in yes-associated protein (YAP) ubiquitinated degradation in PASMCs. YAP is the core effector of the Hippo pathway and mediates the expression of particular genes. The accumulation of YAP further increased the expression and activation of Notch3 and ultimately promoted the proliferation of PASMCs. In addition, we showed that preblocking S1PR2, prior silencing of STAT3, miR-135b, or YAP, and prior inhibition of Notch3 all attenuated S1P-induced PASMCs proliferation. Taken together, our study indicates that S1P stimulates PASMCs proliferation by activation of the S1PR2/STAT3/miR-135b/β-transduction repeat-containing protein/YAP/Notch3 pathway, and our data suggest that targeting this cascade might have potential value in ameliorating PASMCs hyperproliferation and benefit pulmonary arterial hypertension.

Highlights

Pulmonary arterial hypertension (PAH) is a progressive disease characterized by histological changes of the pulmonary vasculature, such as sustained vasoconstriction, uncontrolled pulmonary vascular remodeling, and thrombosis in situ, resulting in structural and functional changes of the pulmonary vasculature, continuous and notable elevation of pulmonary vascular resistance and pulmonary arterial pressure, and eventually leads to heart failure and death [1, 2]
To address above hypothesis and to clarify which S1P receptor (S1PR) is involved in this process, we intervened signal transducers and activators of transcription 3 (STAT3), miR135b, β-transduction repeat-containing protein (β-TrCP), Yes-associated protein (YAP), Notch3, and sphingosine-1-phosphate receptor 2 (S1PR2)/3 in primary cultured Pulmonary artery smooth muscle cells (PASMCs) stimulated with exogenous S1P, separately
STAT3 notably reduced the elevated protein level of YAP induced by S1P in PASMCs, which dropped from 1.62-fold over control in control siRNA and S1P-cotreated cells to 1.32-fold over control in STAT3 siRNA and S1P-cotreated cells

Summary

Introduction

Pulmonary arterial hypertension (PAH) is a progressive disease characterized by histological changes of the pulmonary vasculature, such as sustained vasoconstriction, uncontrolled pulmonary vascular remodeling, and thrombosis in situ, resulting in structural and functional changes of the pulmonary vasculature, continuous and notable elevation of pulmonary vascular resistance and pulmonary arterial pressure, and eventually leads to heart failure and death [1, 2]. Further experiments showed that the S1PR2/STAT3/miR-135b/β-TrCP signaling pathway mediated S1P induction of YAP protein and promoted PASMCs proliferation.

Results

Conclusion