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Cellular and Molecular Life Sciences

, Volume 75, Issue 21, pp 4041–4057 | Cite as

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

  • Henriikka Kentala
  • Annika Koponen
  • Helena Vihinen
  • Juho Pirhonen
  • Gerhard Liebisch
  • Zoltan Pataj
  • Annukka Kivelä
  • Shiqian Li
  • Leena Karhinen
  • Eeva Jääskeläinen
  • Robert Andrews
  • Leena Meriläinen
  • Silke Matysik
  • Elina Ikonen
  • You Zhou
  • Eija Jokitalo
  • Vesa M. OlkkonenEmail author
Original Article

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.

Keywords

Akt signaling CRISPR-Cas9 Glycolysis OSBPL2 OSBP-related protein Triacylglycerol 

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

Notes

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.

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

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Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  • Henriikka Kentala
    • 1
  • Annika Koponen
    • 1
  • Helena Vihinen
    • 2
  • Juho Pirhonen
    • 1
    • 3
  • Gerhard Liebisch
    • 4
  • Zoltan Pataj
    • 4
  • Annukka Kivelä
    • 1
  • Shiqian Li
    • 1
    • 3
  • Leena Karhinen
    • 3
  • Eeva Jääskeläinen
    • 1
  • Robert Andrews
    • 5
  • Leena Meriläinen
    • 2
  • Silke Matysik
    • 4
  • Elina Ikonen
    • 1
    • 3
  • You Zhou
    • 1
    • 5
    • 6
  • Eija Jokitalo
    • 2
  • Vesa M. Olkkonen
    • 1
    • 3
    Email author
  1. 1.Minerva Foundation Institute for Medical ResearchHelsinkiFinland
  2. 2.Electron Microscopy Unit, Institute of BiotechnologyUniversity of HelsinkiHelsinkiFinland
  3. 3.Department of Anatomy, Faculty of MedicineUniversity of HelsinkiHelsinkiFinland
  4. 4.Institute of Clinical Chemistry and Laboratory MedicineUniversity Hospital RegensburgRegensburgGermany
  5. 5.Systems Immunity Research InstituteCardiff UniversityCardiffUK
  6. 6.Division of Infection and ImmunityCardiff University School of MedicineCardiffUK

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