Abstract
The quaternary structure of G protein-coupled receptors (GPCRs) can influence their trafficking and ability to transduce signals. GPCR oligomers are generally portrayed as long-lived entities, although the stability of these complexes has not been studied. Here we show that D2 dopamine receptor protomers interact transiently at a specific oligomer interface. Selective immobilization of cyan fluorescent protein-D2 receptors (C-D2Rs) in the plasma membrane failed to completely immobilize coexpressed D2-venus receptors (D2R-Vs), suggesting that the two did not form stable oligomers with each other. Oxidative cross-linking stabilized C-D2R-D2R-V oligomers such that immobilization of C-D2R also immobilized D2R-V. This stabilization required the presence in both C-D2R and D2R-V of a cysteine residue in transmembrane domain 4 (TM4), a region identified as a putative oligomer interface in these and other class A GPCRs. These results suggest that the interaction of D2 receptor protomers at TM4 is transient unless stabilized and that the quaternary structure of these receptors may thus be subject to physiological or pharmacological regulation.
Footnotes
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This work was supported by the National Institutes of Health National Institute of Neurological Disorders and Stroke [Grant NS045543]; the National Institutes of Health National Institute of General Medical Sciences [Grant GM078319]; and the National Science Foundation [Grant MCB-0620024].
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A report of these findings was presented at the 2008 Annual Meeting of the Biophysical Society, 2008 Feb 2-6, Long Beach, CA.
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ABBREVIATIONS: GPCR, G protein-coupled receptor; TM, transmembrane domain; FRAP, fluorescence recovery after photobleaching; CuP; Cu2+(phenanthroline)2; ECFP, enhanced cyan fluorescent protein; V, venus; C-D2R, cyan fluorescent protein-D2 receptor; D2R-V, D2-venus receptor; GFP, green fluorescent protein; PBS, phosphate-buffered saline; β2AR-V, β2-adrenoreceptor-venus fusion.
- Received December 1, 2008.
- Accepted March 9, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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