Kamiya Toshio, Saitoh Osamu, Yoshioka Kazuaki, Nakata Hiroyasu
Department of Molecular Cell Signaling, Tokyo Metropolitan Institute for Neuroscience, 2-6 Musashidai, Fuchu, 183-8526, Tokyo, Japan.
Biochem Biophys Res Commun. 2003 Jun 27;306(2):544-9. doi: 10.1016/s0006-291x(03)00991-4.
We investigated whether oligomerization of adenosine A(2A) receptor (A(2A)R) and dopamine D(2) receptor (D(2)R) exists in living cells using modified bioluminescence resonance energy transfer (BRET(2)) technology. Fusion of these receptors to a donor, Renilla luciferase (Rluc), and to an acceptor, modified green fluorescent protein (GFP(2)), did not affect the ligand binding affinity, subcellular distribution, and coimmunoprecipitation of the receptors. BRET was detected not only between Myc-D(2)R-Rluc and A(2A)R-GFP(2) but also between HA-tagged A(2A)R-Rluc and A(2A)R-GFP(2). These results indicate A(2A)R, either homomeric or heteromeric with D(2)R, exists as an oligomer in living cells.
我们使用改良的生物发光共振能量转移（BRET₂）技术，研究了腺苷A₂A受体（A₂AR）和多巴胺D₂受体（D₂R）在活细胞中是否存在寡聚化现象。将这些受体与供体海肾荧光素酶（Rluc）和受体改良型绿色荧光蛋白（GFP₂）融合，并不影响受体的配体结合亲和力、亚细胞分布和免疫共沉淀。不仅在Myc-D₂R-Rluc和A₂AR-GFP₂之间检测到了BRET，而且在HA标签的A₂AR-Rluc和A₂AR-GFP₂之间也检测到了BRET。这些结果表明，A₂AR无论是同聚体还是与D₂R形成的异聚体，在活细胞中均以寡聚体形式存在。
