Mapping of the Localization of Type 1 Angiotensin Receptor in Membrane Microdomains Using Bioluminescence Resonance Energy Transfer-based Sensors
Mapping of the Localization of Type 1 Angiotensin Receptor in Membrane Microdomains Using Bioluminescence Resonance Energy Transfer-based Sensors
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DOI：
10.1074/jbc.m111.293944
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发表时间：
2012-03-16
影响因子：
4.8
通讯作者：
Hunyady, Laszlo
中科院分区：
文献类型：
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作者：
Balla, Andras;Toth, Daniel J.;Hunyady, Laszlo
I 型血管紧张素受体 (AT(1)-R) 信号传导的启动和终止可导致其在质膜微域中的定位发生动态变化。最近开发了几种标记物来研究膜微区。在这里，我们使用几种 YFP 标记的融合结构（即筏或非筏质膜标记）来分析激动剂诱导的 AT(1)-R 区室化变化，包括使用生物发光共振能量转移测量响应刺激而在质膜区室之间内化或横向运动。我们的数据表明，血管紧张素 II (AngII) 刺激改变了 HEK293 细胞中与荧光探针共表达的野生型或突变型（DRY -> AAY 或 TSTS -> AAAA）AT(1)-Rs 的微域定位。将 AngII 刺激下 AT(1)-R 的运输与 [Sar(1),Ile(8)]AngII 或 [Sar(1),Ile(4),Ile(8)]AngII 刺激的运输进行比较，揭示了不同类型的变化，具体取决于配体的性质。观察到的 AT(1)-R 受体区室化变化与 5HT-2C 和 EGF 受体的变化显着不同，这证明了基于生物发光共振能量转移的测量在活细胞实验中质膜中受体运输研究中的有用性。
Initiation and termination of signaling of the type I angiotensin receptor (AT(1)-R) can lead to dynamic changes in its localization in plasma membrane microdomains. Several markers were recently developed to investigate membrane microdomains. Here, we used several YFP-labeled fusion constructs (i.e. raft or non-raft plasma membrane markers) to analyze the agonist-induced changes in compartmentalization of AT(1)-R, including internalization or lateral movement between plasma membrane compartments in response to stimulation using bioluminescence resonance energy transfer measurements. Our data demonstrate that angiotensin II (AngII) stimulus changes the microdomain localization of wild type or mutated (DRY -> AAY or TSTS -> AAAA) AT(1)-Rs co-expressed with the fluorescent probes in HEK293 cells. The comparison of the trafficking of AT(1)-R upon AngII stimulus with those of [Sar(1),Ile(8)]AngII or [Sar(1),Ile(4),Ile(8)]AngII stimulus revealed different types of changes, depending on the nature of the ligand. The observed changes in receptor compartmentalization of the AT(1)-R are strikingly different from those of 5HT-2C and EGF receptors, which demonstrate the usefulness of the bioluminescence resonance energy transfer-based measurements in the investigation of receptor trafficking in the plasma membrane in living cell experiments.