Phosphorylation-independent regulation of metabotropic glutamate receptor signaling by G protein-coupled receptor kinase 2
Phosphorylation-independent regulation of metabotropic glutamate receptor signaling by G protein-coupled receptor kinase 2
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DOI：
10.1074/jbc.m203593200
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发表时间：
2002-07-12
影响因子：
4.8
通讯作者：
Ferguson, SSG
中科院分区：
文献类型：
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作者：
Dhami, GK;Anborgh, PH;Ferguson, SSG
G蛋白偶联受体激酶(GRK)介导的G蛋白偶联受体脱敏的公认范式包括GRK介导的受体磷酸化，然后与arrestin蛋白结合。虽然GRK有助于代谢性谷氨酸受体1(MGluR1)的失活，但13-拦阻蛋白似乎不是mGluR1G蛋白解偶联所必需的。因此，我们研究了位于mGluR1a C末端的丝氨酸和苏氨酸残基的磷酸化是否足以使GRK2介导的mGluR1a信号减弱。我们发现，mGluR1a C末端的截断阻止了mGluR1a的磷酸化，而GRK2对mGluR1剪接变异体(MGluR1b)的磷酸化没有贡献。然而，mGluR1a-866Delta和mGluR1b刺激的肌醇磷酸形成在GRK2表达后减弱。表达GRK2 C-末端结构域以阻止内源性GRK2的膜转位增加mGluR1a-866Delta和mGluR1b刺激的肌醇磷酸形成，可能是通过阻止GRK2的膜转位。相反，表达缺乏激酶的GRK2-K220R突变体可以通过这些非磷酸化受体抑制肌醇磷酸的形成。GRK2 N-末端结构域(残基45-185)的表达也减弱了结构性和激动剂刺激的mGluR1a、mGluR1a-866a和mGluR1b信号，GRK2N末端与mGluR1a共沉淀。综上所述，我们的观察表明，GRK2对mGluR1信号的减弱是不依赖于磷酸化的，GRK2的N末端结构域与mGluR1的相互作用有助于调节mGluR1G蛋白偶联。
The accepted paradigm for G protein-coupled receptor kinase (GRK)-mediated desensitization of G protein-coupled receptors involves GRK-mediated receptor phosphorylation followed by the binding of arrestin proteins. Although GRKs contribute to metabotropic glutamate receptor 1 (mGluR1) inactivation, 13-arrestins do not appear to be required for mGluR1 G protein uncoupling. Therefore, we investigated whether the phosphorylation of serine and threonine residues localized within the C terminus of mGluR1a is sufficient to allow GRK2-mediated attenuation of mGluR1a signaling. We find that the truncation of the mGluR1a C-terminal tail prevents mGluR1a phosphorylation and that GRK2 does not contribute to the phosphorylation of an mGluR1 splice variant (mGluR1b). However, mGluR1a-866Delta- and mGluR1b-stimulated inositol phosphate formation is attenuated following GRK2 expression. The expression of the GRK2 C-terminal domain to block membrane translocation of endogenous GRK2 increases mGluR1a-866Delta- and mGluR1b-stimulated inositol phosphate formation, presumably by blocking membrane translocation of GRK2. In contrast, expression of the kinase-deficient GRK2-K220R mutant inhibits inositol phosphate formation by these unphosphorylated receptors. Expression of the GRK2 N-terminal domain (residues 45-185) also attenuates both constitutive and agonist-stimulated mGluR1a, mGluR1a-866A, and mGluR1b signaling, and the GRK2 N terminus co-precipitates with mGluR1a. Taken together, our observations indicate that attenuation of mGluR1 signaling by GRK2 is phosphorylation-independent and that the interaction of the N-terminal domain of GRK2 with mGluR1 contributes to the regulation of mGluR1 G protein coupling.