Cannabinoid CB1 and CB2 Receptor-Mediated Arrestin Translocation: Species, Subtype, and Agonist-Dependence
Cannabinoid CB1 and CB2 Receptor-Mediated Arrestin Translocation: Species, Subtype, and Agonist-Dependence
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DOI：
10.3389/fphar.2019.00350
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发表时间：
2019-04-10
影响因子：
5.6
通讯作者：
Grimsey, Natasha Lillia
中科院分区：
文献类型：
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作者：
Ibsen, Mikkel Soes;Finlay, David B.;Grimsey, Natasha Lillia
抑制蛋白转位和信号转导已成为G蛋白偶联受体分子药理学领域的前沿。一些受体-抑制蛋白相互作用被相对较好地理解，并被认为是特定治疗或不良结局的原因。已经报道了抑制素与大麻素受体1（CB 1）和2（CB 2）的偶联，尽管大多数研究还没有系统地表征这种活性的不同配体依赖性。此外，许多先前的研究利用牛（而不是人）抑制蛋白，最广泛应用的测定需要标记受体，这妨碍了受体类型之间有意义的比较。我们采用了生物发光共振能量转移（BRET）方法，不需要使用标记的受体，从而允许受体类型之间的arrestin易位的比较，以及与缺乏感兴趣的受体的细胞-一个重要的控制。评估了选择的CB 1和CB 2激动剂刺激人和牛β-抑制蛋白-1和-2的细胞表面易位的能力。我们发现，与加压素V-2受体（一种强大的抑制蛋白募集者）相比，一些CB 1配体诱导中度β-抑制蛋白-2易位;然而，CB 1与β-抑制蛋白-1偶联，CB 2与任一抑制蛋白偶联引起相对低的效力。对于受体和抑制蛋白，配体之间的一系列功效是明显的。内源性大麻素2-花生四烯酰甘油作为β-抑制蛋白-2通过CB 1易位的高效配体脱颖而出。δ（9）-四氢大麻酚通常不能引起抑制蛋白亚型通过CB 1或CB 2的易位;然而，对照实验显示在不表达CB 1/CB 2的细胞中的易位，这可能有助于解释与文献的一些差异。GRK 2过表达对CB 1/CB 2诱导的arrestin易位有一定影响。牛和人arrestins的结果在很大程度上是相似的，但牛和人arrestins之间的不一致的等级顺序效力/效力的少数情况下提出的可能性，这些配体稳定的受体构象的细微差异表现在两种arrestins物种的不同亲和力，在配体偏倚研究的解释具有重要的潜在后果。以及贡献的重要信息CB 1/CB 2配体依赖性arrestin耦合，我们的研究提出了一些点考虑在设计和解释arrestin招聘检测。
Arrestin translocation and signaling have come to the fore of the G protein-coupled receptor molecular pharmacology field. Some receptor-arrestin interactions are relatively well understood and considered responsible for specific therapeutic or adverse outcomes. Coupling of arrestins with cannabinoid receptors 1 (CB1) and 2 (CB2) has been reported, though the majority of studies have not systematically characterized the differential ligand dependence of this activity. In addition, many prior studies have utilized bovine (rather than human) arrestins, and the most widely applied assays require reporter-tagged receptors, which prevent meaningful comparison between receptor types. We have employed a bioluminescence resonance energy transfer (BRET) method that does not require the use of tagged receptors and thereby allows comparisons of arrestin translocation between receptor types, as well as with cells lacking the receptor of interest - an important control. The ability of a selection of CB1 and CB2 agonists to stimulate cell surface translocation of human and bovine beta-arrestin-1 and -2 was assessed. We find that some CB1 ligands induce moderate beta-arrestin-2 translocation in comparison with vasopressin V-2 receptor (a robust arrestin recruiter); however, CB1 coupling with beta-arrestin-1 and CB2 with either arrestin elicited low relative efficacies. A range of efficacies between ligands was evident for both receptors and arrestins. Endocannabinoid 2-arachidonoylglycerol stood out as a high efficacy ligand for translocation of beta-arrestin-2 via CB1. Delta(9)-tetrahydrocannabinol was generally unable to elicit translocation of either arrestin subtype via CB1 or CB2; however, control experiments revealed translocation in cells not expressing CB1/CB2, which may assist in explaining some discrepancy with the literature. Overexpression of GRK2 had modest influence on CB1/CB2-induced arrestin translocation. Results with bovine and human arrestins were largely analogous, but a few instances of inconsistent rank order potencies/efficacies between bovine and human arrestins raise the possibility that subtle differences in receptor conformation stabilized by these ligands manifest in disparate affinities for the two arrestin species, with important potential consequences for interpretation in ligand bias studies. As well as contributing important information regarding CB1/CB2 ligand-dependent arrestin coupling, our study raises a number of points for consideration in the design and interpretation of arrestin recruitment assays.