Phosphorylation and Desensitization of G Protein-Coupled Receptors

我们研究了G蛋白偶联受体激酶（GRK）磷酸化与G蛋白偶联受体脱敏之间的关系，特别是在培养细胞中表达的毒蕈碱乙酰胆碱受体m2亚型（m2受体）上。我们发现，GRK2的共表达促进了中国仓鼠卵巢细胞（CHO细胞）中m2受体的内化、下调和解偶联，这很可能是通过m2受体的激动剂依赖性磷酸化来实现的。在COS-7细胞中表达的其他G蛋白偶联受体，包括m3、m4和m5毒蕈碱和多巴胺D2受体，也被发现通过GRK2的共表达促进内化，而m1毒蕈碱和多巴胺D4受体的内化不被GRK2的共表达促进。GRK2被激动剂结合受体和G蛋白β - γ亚基激活后，钙调素以Ca^<2+>依赖的方式减弱。尽管GRK2的底物特异性非常严格，已知的底物仅限于活化形式的G蛋白偶联受体，但研究发现微管蛋白与GRK2相互作用并被GRK2磷酸化。钙调蛋白和微管蛋白磷酸化抑制的生理意义仍有待阐明。

We have examined the relation between phosphorylation by G protein-coupled receptor kinases (GRK) of G protein-coupled receptors and their desensitization, particularly on muscarinic acetylcholine receptor m2 subtypes (m2 receptors) expressed in cultured cells. We found that internalization, down-regulation and uncoupling of m2 receptors expressed in chinese hamster ovary cells (CHO cells) were facilitated by coexpression of GRK2, most probably through agonist-dependent phosphorylation of m2 receptors. Internalization of other G protein-coupled receptors including m3, m4 and m5 muscarinic and dopamine D2 receptors, which were expressed in COS-7 cells, was also found to be facilitated by coexpression of GRK2, whereas internalization of m1 muscarinic and dopamine D4 receptors was not facilitated by coexpression of GRK2.Activation of GRK2 by agonist-bound receptros and G protein betagamma subunits was found to be attenuated by calmodulin in a Ca^<2+> dependent manner. Tubulin was found to interact with and be phosphorylated by GRK2, in spite of the fact that the substrate specificity of GRK2 is very strict and the known substrates are limited to activated forms of G protein-coupled receptors. Physiological meanings of inhibition by calmodulin and tubulin phosphorylation remain to be elucidated.