Temporal dependence of shifts in mu opioid receptor mobility at the cell surface after agonist binding observed by single-particle tracking.
Temporal dependence of shifts in mu opioid receptor mobility at the cell surface after agonist binding observed by single-particle tracking.
复制标题
通过单粒子追踪观察到激动剂结合后细胞表面μ阿片受体迁移率变化的时间依赖性。
DOI：
10.1038/s41598-019-43657-x
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发表时间：
2019
影响因子：
4.6
通讯作者：
Hentges,ShaneT
中科院分区：
文献类型：
--
作者：
Metz,MarissaJ;Pennock,ReaganL;Krapf,Diego;Hentges,ShaneT
激动剂与mu阿片受体（MOR）结合导致构象变化，从而允许g蛋白的募集，下游效应物的激活以及最终的脱敏和内化，所有这些都可能影响受体的流动性。本研究采用量子点标记的flag标记MORs的单粒子跟踪（SPT）来检测激动剂结合后MOR迁移率的变化。在基础条件下，质膜上的FLAG-MORs处于移动和不移动状态。用DAMGO激活FLAG-MORs会导致可移动MORs的比例急剧增加，而可移动轨迹的自由部分部分依赖于与g蛋白的相互作用。相比之下，暴露于DAMGO或吗啡10分钟会增加固定的FLAG-MORs的比例。虽然长时间暴露于DAMGO导致的移动性下降与网格蛋白共定位的增加相对应，但在可移动和不可移动的FLAG-MORs中均存在共定位的增加。因此，受体的单一迁移状态不能解释与网格蛋白的共定位。这些发现表明，SPT可用于跟踪激动剂依赖性MOR迁移率随时间的变化，但观察到的迁移率状态可能是由一系列不同的相互作用引起的，当与特定的下游效应物一起检查时，将是最有信息的。
Agonist binding to the mu opioid receptor (MOR) results in conformational changes that allow recruitment of G-proteins, activation of downstream effectors and eventual desensitization and internalization, all of which could affect receptor mobility. The present study employed single particle tracking (SPT) of quantum dot labeled FLAG-tagged MORs to examine shifts in MOR mobility after agonist binding. FLAG-MORs on the plasma membrane were in both mobile and immobile states under basal conditions. Activation of FLAG-MORs with DAMGO caused an acute increase in the fraction of mobile MORs, and free portions of mobile tracks were partially dependent on interactions with G-proteins. In contrast, 10-minute exposure to DAMGO or morphine increased the fraction of immobile FLAG-MORs. While the decrease in mobility with prolonged DAMGO exposure corresponded to an increase in colocalization with clathrin, the increase in colocalization was present in both mobile and immobile FLAG-MORs. Thus, no single mobility state of the receptor accounted for colocalization with clathrin. These findings demonstrate that SPT can be used to track agonist-dependent changes in MOR mobility over time, but that the mobility states observed likely arise from a diverse set of interactions and will be most informative when examined in concert with particular downstream effectors.