Oligomerization of μ- and δ-Opioid Receptors

doi:10.1074/jbc.M000345200

Oligomerization of μ- and δ-Opioid Receptors

GENERATION OF NOVEL FUNCTIONAL PROPERTIES*

Abstract

The existence of dimers and oligomers for many G protein-coupled receptors has been described by us and others. Since many G protein-coupled receptor subtypes are highly homologous to each other, we examined whether closely related receptors may interact with each other directly and thus have the potential to create novel signaling units. Using μ- and δ-opioid receptors, we show that each receptor expressed individually was pharmacologically distinct and could be visualized following electrophoresis as monomers, homodimers, homotetramers, and higher molecular mass oligomers. When μ- and δ-opioid receptors were coexpressed, the highly selective synthetic agonists for each had reduced potency and altered rank order, whereas endomorphin-1 and Leu-enkephalin had enhanced affinity, suggesting the formation of a novel binding pocket. No heterodimers were visualized in the membranes coexpressing μ- and δ-receptors by the methods available. However, hetero-oligomers were identified by the ability to co-immunoprecipitate μ-receptors with δ-receptors and vice versa using differentially epitope-tagged receptors. In contrast to the individually expressed μ- and δ-receptors, the coexpressed receptors showed insensitivity to pertussis toxin and continued signal transduction, likely due to interaction with a different subtype of G protein. In this study, we provide, for the first time, evidence for the direct interaction of μ- and δ-opioid receptors to form oligomers, with the generation of novel pharmacology and G protein coupling properties.

Footnotes

↵* This work was supported by grants from the Medical Research Council of Canada, the National Institute on Drug Abuse, and the Smokeless Tobacco Research Council, Inc.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵‖ To whom correspondence should be addressed: Dept. of Pharmacology, University of Toronto, Medical Sciences Bldg., Rm. 4358, 1 King's College Circle, Toronto, Ontario M5S 1A8, Canada. Tel.: 416-978-3367; Fax: 416-971-2868; E-mail: s.george@utoronto.ca.
Published, JBC Papers in Press, June 6, 2000, DOI 10.1074/jbc.M000345200
Abbreviations:

GPCRs

G protein-coupled receptors

DAMGO

[d-Ala²,N-methyl-Phe⁴,Gly⁵-ol]enkephalin

DPDPE

[d- Pen^2,5]enkephalin

K_H and K_L

affinity constants for the agonist-detected high and low affinity sites, respectively

GTPγS

guanosine 5′-O-(3-thiotriphosphate)

Pen

penicillamine
- Received January 18, 2000.
- Revision received May 30, 2000.
The American Society for Biochemistry and Molecular Biology, Inc.