S1P in HDL promotes interaction between SR-BI and S1PR1 and activates S1PR1-mediated biological functions: calcium flux and S1PR1 internalization

Mary G Sorci-Thomas,Hesham M El-Shewy,Maria F Lopes-Virella,Samar M Hammad,Louis M Luttrell,Michael J Thomas,Richard L Klein,Mi-Hye Lee,Kathryn M Appleton

doi:10.1194/jlr.m070706

Abstract

HDL normally transports about 50-70% of plasma sphingosine 1-phosphate (S1P), and the S1P in HDL reportedly mediates several HDL-associated biological effects and signaling pathways. The HDL receptor, SR-BI, as well as the cell surface receptors for S1P (S1PRs) may be involved partially and/or completely in these HDL-induced processes. Here we investigate the nature of the HDL-stimulated interaction between the HDL receptor, SR-BI, and S1PR1 using a protein-fragment complementation assay and confocal microscopy. In both primary rat aortic vascular smooth muscle cells and HEK293 cells, the S1P content in HDL particles increased intracellular calcium concentration, which was mediated by S1PR1. Mechanistic studies performed in HEK293 cells showed that incubation of cells with HDL led to an increase in the physical interaction between the SR-BI and S1PR1 receptors that mainly occurred on the plasma membrane. Model recombinant HDL (rHDL) particles formed in vitro with S1P incorporated into the particle initiated the internalization of S1PR1, whereas rHDL without supplemented S1P did not, suggesting that S1P transported in HDL can selectively activate S1PR1. In conclusion, these data suggest that S1P in HDL stimulates the transient interaction between SR-BI and S1PRs that can activate S1PRs and induce an elevation in intracellular calcium concentration.

Highlights

HDL normally transports about 50–70% of plasma sphingosine 1-phosphate (S1P), and the S1P in HDL reportedly mediates several HDL-associated biological effects and signaling pathways
Intracellular calcium influx was substantially increased when cells were incubated with recombinant HDL (rHDL) particles supplemented with S1P but not when the cells were incubated with rHDL without S1P supplementation (Fig. 1A)
We have shown that S1P transported in HDL is biologically active and is a major effector of HDL-mediated increase in intracellular calcium

Summary

Introduction

HDL normally transports about 50–70% of plasma sphingosine 1-phosphate (S1P), and the S1P in HDL reportedly mediates several HDL-associated biological effects and signaling pathways. We investigate the nature of the HDLstimulated interaction between the HDL receptor, SR-BI, and S1PR1 using a protein-fragment complementation assay and confocal microscopy In both primary rat aortic vascular smooth muscle cells and HEK293 cells, the S1P content in HDL particles increased intracellular calcium concentration, which was mediated by S1PR1. The mass spectroscopy analyses were performed in the Mass Spectrometer Facility of the Comprehensive Cancer Center of Wake Forest School of Medicine supported in part by National Cancer Institute Center Grant 5P30CA12197 The contents of this manuscript do not represent the views of the Department of Veterans Affairs or the United States Government. Numerous studies have detailed the various signaling pathways generated by the interaction of HDL with cell surface receptors that have been shown to induce myriad

Methods

Results

Conclusion