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

Riikka Hynynen²^,*, Monika Suchanek²^,, Johanna Spandl, Nils Bäck, Christoph Thiele and Vesa M. Olkkonen¹^,*^,

^* National Institute for Health and Welfare, and FIMM, Institute for Molecular Medicine Finland, Biomedicum, FI-00290 University of Helsinki, Helsinki, Finland
Max-Planck-Institute of Molecular Cell Biology and Genetics, D-01307 Dresden, Germany
Institute of Biomedicine/Anatomy, University of Helsinki, FI-00014 Helsinki, Finland
^² R. Hynynen and M. Suchanek contributed equally to this work.

The online version of this article (available at http://www.jlr.org)contains supplementary data in the form of two figures.

This study was supported by the Academy of Finland (Grants 113013,118720, and 121457 to V.M.O.), the Sigrid Juselius Foundation,the Finnish Foundation for Cardiovascular Research, the MagnusEhrnrooth Foundation, the Finnish Cultural Foundation (V.M.O.),the Biomedicum Helsinki Foundation (R.H.), the PerklénFoundation, and the Liv och Hälsa Foundation (N.B). Moreover,the work was partially funded by the European Union programLIPIDOMICNET (Grant 202272). R.H. is a member of the HelsinkiBiomedical Graduate School, the support of which is acknowledged.

Published, JLR Papers in Press, February 17, 2009.

^¹ To whom correspondence should be addressed. e-mail: vesa.olkkonen{at}thl.fi

Oxysterol binding protein-related protein 2 (ORP2) is a memberof the oxysterol binding protein family, previously shown tobind 25-hydroxycholesterol and implicated in cellular cholesterolmetabolism. We show here that ORP2 also binds 22(R)-hydroxycholesterol[22(R)OHC], 7-ketocholesterol, and cholesterol, with 22(R)OHCbeing the highest affinity ligand of ORP2 (K_d 1.4 x 10^–8M). We report the localization of ORP2 on cytoplasmic lipiddroplets (LDs) and its function in neutral lipid metabolismusing the human A431 cell line as a model. The ORP2 LD associationdepends on sterol binding: Treatment with 5 µM 22(R)OHCinhibits the LD association, while a mutant defective in sterolbinding is constitutively LD bound. Silencing of ORP2 usingRNA interference slows down cellular triglyceride hydrolysis.Furthermore, ORP2 silencing increases the amount of [¹⁴C]cholesterylesters but only under conditions in which lipogenesis and LDformation are enhanced by treatment with oleic acid. The resultsidentify ORP2 as a sterol receptor present on LD and provideevidence for its role in the regulation of neutral lipid metabolism,possibly as a factor that integrates the cellular metabolismof triglycerides with that of cholesterol.

Supplementary key words cholesteryl ester • oxysterol • oxysterol binding protein • triglyceride

Abbreviations: 22(R)OHC, 22(R)-hydroxycholesterol; 25OHC, 25-hydroxycholesterol; CE, cholesteryl ester; CHO, Chinese hamster ovary; ER, endoplasmic reticulum; FCS, fetal calf serum; FFAT, two phenylalanines in an acidic tract; GST, glutathione S-transferase; LD, lipid droplet; mab, monoclonal mouse antibody; mβCD, methyl-β-cyclodextrin; ORD, oxysterol binding protein-related domain; ORP, oxysterol binding protein-related protein; OSBP, oxysterol binding protein; siRNA, short interfering RNA; SREBP, sterol regulatory element binding protein; TG, triglyceride

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