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OSBP-related protein-2 (ORP2): a novel Akt effector that controls cellular energy metabolism

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Abstract

ORP2 is a ubiquitously expressed OSBP-related protein previously implicated in endoplasmic reticulum (ER)—lipid droplet (LD) contacts, triacylglycerol (TG) metabolism, cholesterol transport, adrenocortical steroidogenesis, and actin-dependent cell dynamics. Here, we characterize the role of ORP2 in carbohydrate and lipid metabolism by employing ORP2-knockout (KO) hepatoma cells (HuH7) generated by CRISPR-Cas9 gene editing. The ORP2-KO and control HuH7 cells were subjected to RNA sequencing, analyses of Akt signaling, carbohydrate and TG metabolism, the extracellular acidification rate, and the lipidome, as well as to transmission electron microscopy. The loss of ORP2 resulted in a marked reduction of active phosphorylated Akt(Ser473) and its target Glycogen synthase kinase 3β(Ser9), consistent with defective Akt signaling. ORP2 was found to form a physical complex with the key controllers of Akt activity, Cdc37, and Hsp90, and to co-localize with Cdc37 and active Akt(Ser473) at lamellipodial plasma membrane regions, in addition to the previously reported ER–LD localization. ORP2-KO reduced glucose uptake, glycogen synthesis, glycolysis, mRNA-encoding glycolytic enzymes, and SREBP-1 target gene expression, and led to defective TG synthesis and storage. ORP2-KO did not reduce but rather increased ER–LD contacts under basal culture conditions and interfered with their expansion upon fatty acid loading. Together with our recently published work (Kentala et al. in FASEB J 32:1281–1295, 2018), this study identifies ORP2 as a new regulatory nexus of Akt signaling, cellular energy metabolism, actin cytoskeletal function, cell migration, and proliferation.

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Abbreviations

CRISPR:

Clustered regularly interspaced short palindromic repeats

ECAR:

Extracellular acidification rate

EM:

Electron microscopy

ER:

Endoplasmic reticulum

FA:

Fatty acid

GSK:

Glycogen synthase kinase

HUVEC:

Human umbilical vein endothelial cell

IPA:

Ingenuity® pathway analysis

KO:

Knockout

LD:

Lipid droplet

LDL:

Low-density lipoprotein

OSBP:

Oxysterol-binding protein

ORP:

OSBP-related protein

PIP:

Phosphatidylinositol phosphate

TEM:

Transmission electron microscopy

TG:

Triacylglycerol

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Acknowledgements

We thank Riikka Kosonen and Mervi Lindman for expert technical assistance, and Adj. Prof. Reijo Käkelä (Department of Biosciences, University of Helsinki) for valuable comments on the manuscript. Personnel of the Genome Biology Unit (Biocenter Finland) and the Biomedicum Functional Genomics Unit (Helsinki Institute of Life Science, HiLIFE) are acknowledged for help in generating the recombinant lentiviruses. Prof. Feng Zhang (Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, MA, USA) is thanked for kindly providing components of the CRISPR-Cas9 vector system.

Funding

This study was supported by grants from the Academy of Finland (285223 to V.M.O., 307415 and 312491 to E.I.), the University of Helsinki Doctoral Programme in Biomedicine (H.K.), the Finnish Concordia Fund (H.K.), the Ida Montin Foundation (H.K), the Finnish-Norwegian Medical Foundation (H.K.), the Aarne Koskelo Foundation (H.K.), the Orion Research Foundation sr (H.K.), the Päivikki and Sakari Sohlberg Foundation (H.K.), the Sigrid Juselius Foundation, the Magnus Ehrnrooth Foundation, and the Finnish Foundation for Cardiovascular Research (V.M.O.). Electron Microscopy Unit is supported by Biocenter Finland and Helsinki Institute of Life Science. The funding bodies played no role in the study design, analysis or interpretation of the data, writing of the report or the decision to submit the article for publication.

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Authors and Affiliations

  1. Minerva Foundation Institute for Medical Research, Biomedicum 2U, Tukholmankatu 8, 00290, Helsinki, Finland

    Henriikka Kentala, Annika Koponen, Juho Pirhonen, Annukka Kivelä, Shiqian Li, Eeva Jääskeläinen, Elina Ikonen, You Zhou & Vesa M. Olkkonen

  2. Electron Microscopy Unit, Institute of Biotechnology, University of Helsinki, 00014, Helsinki, Finland

    Helena Vihinen, Leena Meriläinen & Eija Jokitalo

  3. Department of Anatomy, Faculty of Medicine, University of Helsinki, 00014, Helsinki, Finland

    Juho Pirhonen, Shiqian Li, Leena Karhinen, Elina Ikonen & Vesa M. Olkkonen

  4. Institute of Clinical Chemistry and Laboratory Medicine, University Hospital Regensburg, 93053, Regensburg, Germany

    Gerhard Liebisch, Zoltan Pataj & Silke Matysik

  5. Systems Immunity Research Institute, Cardiff University, Cardiff, CF14 4XN, UK

    Robert Andrews & You Zhou

  6. Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff, CF14 4XN, UK

    You Zhou

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  1. Henriikka Kentala
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  17. Vesa M. Olkkonen
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Correspondence to Vesa M. Olkkonen.

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Kentala, H., Koponen, A., Vihinen, H. et al. OSBP-related protein-2 (ORP2): a novel Akt effector that controls cellular energy metabolism. Cell. Mol. Life Sci. 75, 4041–4057 (2018). https://doi.org/10.1007/s00018-018-2850-8

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