OSBP-related protein 2 is a sterol receptor on lipid droplets that regulates the metabolism of neutral lipids

Riikka Hynynen,Monika Suchanek,Johanna Spandl,Nils Ba¨Ck,Christoph Thiele,Vesa M Olkkonen

doi:10.1194/jlr.m800661-jlr200

Abstract

Oxysterol binding protein-related protein 2 (ORP2) is a member of the oxysterol binding protein family, previously shown to bind 25-hydroxycholesterol and implicated in cellular cholesterol metabolism. We show here that ORP2 also binds 22(R)-hydroxycholesterol [22(R)OHC], 7-ketocholesterol, and cholesterol, with 22(R)OHC being the highest affinity ligand of ORP2 (K(d) 1.4 x 10(-8) M). We report the localization of ORP2 on cytoplasmic lipid droplets (LDs) and its function in neutral lipid metabolism using the human A431 cell line as a model. The ORP2 LD association depends on sterol binding: Treatment with 5 microM 22(R)OHC inhibits the LD association, while a mutant defective in sterol binding is constitutively LD bound. Silencing of ORP2 using RNA interference slows down cellular triglyceride hydrolysis. Furthermore, ORP2 silencing increases the amount of [(14)C]cholesteryl esters but only under conditions in which lipogenesis and LD formation are enhanced by treatment with oleic acid. The results identify ORP2 as a sterol receptor present on LD and provide evidence for its role in the regulation of neutral lipid metabolism, possibly as a factor that integrates the cellular metabolism of triglycerides with that of cholesterol.

Highlights

Oxysterol binding protein-related protein 2 (ORP2) is a member of the oxysterol binding protein family, previously shown to bind 25-hydroxycholesterol and implicated in cellular cholesterol metabolism
We employed a charcoal-dextran pull-down assay of [3H]oxysterols with glutathione S-transferase (GST)-tagged ORP2 produced in E. coli as previously
The starting point of this study is the novel finding that ORP2 localizes on intracellular lipid droplet (LD)

Summary

Introduction

Oxysterol binding protein-related protein 2 (ORP2) is a member of the oxysterol binding protein family, previously shown to bind 25-hydroxycholesterol and implicated in cellular cholesterol metabolism. When the cells were treated overnight with 350 mM oleic acid-BSA to enhance lipogenesis and LD formation, prominent ORP2 localization on Bodipy 493/503positive spherical structures was evident (Fig. 2D–F). A431 cells overexpressing ORP2 were treated overnight with 350 mM oleic acid-BSA to induce LD formation in the presence of 5 mM 22(R)OHC.

Results

Conclusion