Centre for Cellular and Molecular Biology, Hyderabad, India.
Biophys J. 2011 Jan 19;100(2):361-8. doi: 10.1016/j.bpj.2010.12.3692.
The serotonin₁(A) receptor is a representative member of the GPCR superfamily and serves as an important drug target. The possible role of GPCR oligomerization in receptor function is an active area of research. We monitored the oligomerization state of serotonin₁(A) receptors using homo-FRET and fluorescence lifetime measurements. Homo-FRET is estimated by a reduction in fluorescence anisotropy and provides a superior approach for exploring oligomerization. In addition, homo-FRET offers the possibility of detecting higher-order oligomers. On the basis of an observed increase in fluorescence anisotropy upon progressive photobleaching and analysis of the difference between the extrapolated anisotropy and the predicted anisotropy of an immobile monomer, we propose the presence of constitutive oligomers of the serotonin₁(A) receptor. To the best of our knowledge, these results constitute the first report of higher-order oligomers for the serotonin₁(A) receptor. We further show that cholesterol depletion and antagonist treatment result in a reduced population of higher-order oligomers. In contrast, agonist stimulation and destabilization of the actin cytoskeleton lead to an increased contribution from higher oligomers. These results provide novel insight into the oligomerization status of the serotonin₁(A) receptor that could enhance the ability to design better therapeutic strategies to combat diseases related to malfunctioning of GPCRs.
5-羟色胺₁(A)受体是 G 蛋白偶联受体超家族的代表性成员，是重要的药物靶点。GPCR 寡聚化在受体功能中的可能作用是一个活跃的研究领域。我们使用同型荧光共振能量转移（homo-FRET）和荧光寿命测量来监测 5-羟色胺₁(A)受体的寡聚状态。homo-FRET 通过荧光各向异性的降低来估计，并为探索寡聚提供了一种优越的方法。此外，homo-FRET 还提供了检测更高阶寡聚体的可能性。基于荧光各向异性在逐步光漂白过程中的观察到的增加，以及对不可移动单体的外推各向异性和预测各向异性之间差异的分析，我们提出 5-羟色胺₁(A)受体存在组成性寡聚体。据我们所知，这些结果首次报道了 5-羟色胺₁(A)受体的更高阶寡聚体。我们进一步表明，胆固醇耗竭和拮抗剂处理导致更高阶寡聚体的种群减少。相比之下，激动剂刺激和肌动蛋白细胞骨架的不稳定导致更高的高阶寡聚体贡献。这些结果为 5-羟色胺₁(A)受体的寡聚状态提供了新的见解，这可能增强设计更好的治疗策略以对抗与 GPCR 功能障碍相关疾病的能力。
