Agonist selective regulation of g proteins by cannabinoid CB1 and CB2 receptors
Agonist selective regulation of g proteins by cannabinoid CB1 and CB2 receptors
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DOI：
10.1124/mol.56.6.1362
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发表时间：
1999-12-01
影响因子：
3.6
通讯作者：
Northup, JK
中科院分区：
文献类型：
--
作者：
Glass, M;Northup, JK
我们通过直接测量 G 蛋白活化的原位重建技术检查了人大麻素 CB1 和 CB2 受体的配体调节和 G 蛋白选择性。从表达 CB1 和 CB2 受体的草地贪夜蛾细胞膜中提取离液剂，使内源性 GTP 结合蛋白变性。然后用纯化的牛脑 G(i) 和 G(o) 检查受体催化每种 G 蛋白的 GDP-GTP 交换的能力。 CB1 受体的激活对 G(i) 和 G(o) 产生高亲和力可饱和相互作用。 CB2 受体的激动剂刺激也导致与 G(i) 的高亲和力饱和相互作用。相反，CB2受体不能有效地与G(o)相互作用。然后用不同的配体组检查 G 蛋白的激活。对于 CB2 受体与 G(i) 的相互作用，HU210 是唯一表现出最大激活的测试化合物。相比之下，WIN55,212 (64%)、anandamide (42%) 和 Delta(9)-四氢大麻酚 (Delta(9)-THC) (44%) 均启动次最大水平的 G 蛋白激活。对于 CB1 受体催化的 G(i) 激活，HU210、WIN55,212 和 anandamide 均引发最大激活，而 Delta(9)-THC (56 +/- 6%) 仅引起部分 G(i) 激活。相反，仅HU210对G(o)产生最大CB1刺激，与HU210相比，anandamide、WIN55,212和Delta(9)-THC均刺激60%至75%。这些数据表明，不同的激动剂诱导 CB1 受体的不同构象，从而可以区分不同的 G 蛋白。因此，我们的数据证明了 CB1 受体的激动剂选择性 G 蛋白信号传导，并表明治疗剂可以设计为选择性调节个体 G 蛋白信号传导途径。
We have examined the ligand regulation and G protein selectivity of the human cannabinoid CB1 and CB2 receptors by an in situ reconstitution technique directly measuring G protein activation. Membranes from Spodoptera frugiperda cells expressing CB1 and CB2 receptors were chaotrope extracted to denature endogenous GTP-binding proteins. The ability of the receptors to catalyze the GDP-GTP exchange of each G protein was then examined with purified bovine brain G(i) and G(o). Activation of CB1 receptors produced a high-affinity saturable interaction for both G(i) and G(o). Agonist stimulation of CB2 receptors also resulted in a high-affinity saturable interaction with G(i). In contrast, CB2 receptors did not interact efficiently with G(o). G protein activation was then examined with a diverse group of ligands. For the interaction of CB2 receptors with G(i), HU210 was the only compound tested that demonstrated maximal activation. In contrast, WIN55,212 (64%), anandamide (42%), and Delta(9)-tetrahydrocannabinol (Delta(9)-THC) (44%) all initiated submaximal levels of G protein activation. For CB1 receptor-catalyzed activation of G(i), HU210, WIN55,212, and anandamide all elicited maximal activation, whereas Delta(9)-THC (56 +/- 6%) caused only partial G(i) activation. In contrast, only HU210 effected maximal CB1 stimulation of G(o), with anandamide, WIN55,212, and Delta(9)-THC all stimulating between 60 and 75% compared with HU210. These data demonstrate that different agonists induce different conformations of the CB1 receptor, which in turn can distinguish between different G proteins. Our data thus demonstrate agonist-selective G protein signaling by the CB1 receptor and suggest that therapeutic agents may be designed to regulate individual G protein-signaling pathways selectively.