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J. Biol. Chem., Vol. 279, Issue 15, 15541-15549, April 9, 2004
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1D-Adrenergic Receptors Is Controlled by Heterodimerization with 1B-Adrenergic Receptors*
From the Department of Pharmacology, Emory University, Atlanta, Georgia 30322
1-Adrenergic receptors (ARs) belong to the large Class I G protein-coupled receptor superfamily and comprise three subtypes (1A, 1B, and 1D). Previous work with heterologously expressed C-terminal green fluorescent protein (GFP)-tagged 1-ARs showed that 1A- and 1B-ARs localize to the plasma membrane, whereas 1D-ARs accumulate intracellularly. We recently showed that 1D- and 1B-ARs form heterodimers, whereas 1D- and 1A-ARs do not. Here, we examined the role of heterodimerization in regulating 1D-AR localization using both confocal imaging of GFP- or CFP-tagged 1-ARs and a luminometer-based surface expression assay in HEK293 cells. Co-expression with 1B-ARs caused 1D-ARs to quantitatively translocate to the cell surface, but co-expression with 1A-ARs did not. Truncation of the 1B-AR extracellular N terminus or intracellular C terminus had no effect on surface expression of 1D-ARs, suggesting primary involvement of the hydrophobic core. Co-transfection with an uncoupled mutant 1B-AR (
121B) increased both 1D-AR surface expression and coupling to norepinephrine-stimulated Ca2+ mobilization. Finally, GFP-tagged 1D-ARs were not detected on the cell surface when expressed in rat aortic smooth muscle cells that express no endogenous ARs, but were almost exclusively localized on the surface when expressed in DDT1MF-2 cells, which express endogenous 1B-ARs. These studies demonstrate that 1B/1D-AR heterodimerization controls surface expression and functional coupling of 1D-ARs, the N- and C-terminal domains are not involved in this interaction, and that 1B-AR G protein coupling is not required. These observations may be relevant to many other Class I G protein-coupled receptors, where the functional consequences of heterodimerization are still poorly understood.
Received for publication, December 22, 2003 , and in revised form, January 12, 2004.
* The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Supported by grants from the National Institutes of Health (NIH) and by a Distinguished Young Scholar in Medical Research Award from the W.M. Keck Foundation.
¶ Supported by grants from the NIH.
To whom correspondence should be addressed: Dept. of Pharmacology, Emory University, Atlanta, GA 30322. Tel.: 404-727-0363; Fax: 404-727-0365; E-mail: chague{at}emory.edu.
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