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J Biol Chem. 2014 Nov 14;289(46):32353-63. doi: 10.1074/jbc.M114.603506. Epub 2014 Sep 29.

A direct interaction between the sigma-1 receptor and the hERG voltage-gated K+ channel revealed by atomic force microscopy and homogeneous time-resolved fluorescence (HTRF®).

Author information

1
From the Department of Pharmacology, University of Cambridge, Cambridge CB2 1PD, United Kingdom.
2
CisBio Bioassays, Parc Marcel Boiteux BP 84175, 30200 Codolet, France, and.
3
the Institut de Biologie de Valrose (iBV), CNRS UMR 7277, INSERM U1091 UNS, Faculté des Sciences, Université de Nice Sophia Antipolis, 06108 Nice Cedex 2, France.
4
the Institut de Biologie de Valrose (iBV), CNRS UMR 7277, INSERM U1091 UNS, Faculté des Sciences, Université de Nice Sophia Antipolis, 06108 Nice Cedex 2, France olivier.soriani@unice.fr.
5
From the Department of Pharmacology, University of Cambridge, Cambridge CB2 1PD, United Kingdom, jme1000@cam.ac.uk.

Abstract

The sigma-1 receptor is an endoplasmic reticulum chaperone protein, widely expressed in central and peripheral tissues, which can translocate to the plasma membrane and modulate the function of various ion channels. The human ether-à-go-go-related gene encodes hERG, a cardiac voltage-gated K(+) channel that is abnormally expressed in many human cancers and is known to interact functionally with the sigma-1 receptor. Our aim was to investigate the nature of the interaction between the sigma-1 receptor and hERG. We show that the two proteins can be co-isolated from a detergent extract of stably transfected HEK-293 cells, consistent with a direct interaction between them. Atomic force microscopy imaging of the isolated protein confirmed the direct binding of the sigma-1 receptor to hERG monomers, dimers, and tetramers. hERG dimers and tetramers became both singly and doubly decorated by sigma-1 receptors; however, hERG monomers were only singly decorated. The distribution of angles between pairs of sigma-1 receptors bound to hERG tetramers had two peaks, at ∼90 and ∼180° in a ratio of ∼2:1, indicating that the sigma-1 receptor interacts with hERG with 4-fold symmetry. Homogeneous time-resolved fluorescence (HTRF®) allowed the detection of the interaction between the sigma-1 receptor and hERG within the plane of the plasma membrane. This interaction was resistant to sigma ligands, but was decreased in response to cholesterol depletion of the membrane. We suggest that the sigma-1 receptor may bind to hERG in the endoplasmic reticulum, aiding its assembly and trafficking to the plasma membrane.

KEYWORDS:

Atomic Force Microscopy (AFM); Fluorescence Resonance Energy Transfer (FRET); Molecular Imaging; Protein Complex; Sigma Receptor; hERG

PMID:
25266722
PMCID:
PMC4231707
DOI:
10.1074/jbc.M114.603506
[Indexed for MEDLINE]
Free PMC Article

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