Regulation of G protein-coupled receptor kinases by caveolin
Regulation of G protein-coupled receptor kinases by caveolin
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DOI：
10.1074/jbc.274.13.8858
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发表时间：
1999-03-26
影响因子：
4.8
通讯作者：
Benovic, JL
中科院分区：
文献类型：
--
作者：
Carman, CV;Lisanti, MP;Benovic, JL
G 蛋白偶联受体激酶 (GRK) 的主要特征在于其磷酸化 G 蛋白偶联受体并使 G 蛋白偶联受体脱敏的能力。然而，最近的研究表明，GRK 在细胞中可能具有更多样化的蛋白质/蛋白质相互作用。基于GRK2的pleckstrin同源结构域内共有的caveolin结合基序的鉴定，我们测试了纯化的全长GRK2与各种谷胱甘肽S-转移酶-caveolin-1融合蛋白的直接结合，并且我们发现了GRK2与caveolin支架结构域的特异性相互作用。有趣的是，对缺乏普莱克斯特林同源结构域的GRK1和GRK5的分析揭示了与GRK2相似的体外结合特性，麦芽糖结合蛋白caveolin和谷胱甘肽S-转移酶-GRK融合蛋白被用来绘制GRK2和GRK5 N末端的重叠区域，这些重叠区域似乎介导保守的GRK/caveolin相互作用。 GRK2和caveolin在A431和NIH-3T3细胞中的共分级表明GRK2和caveolin的体内关联，并且通过GRK2和caveolin在COS-1细胞中的免疫共沉淀得到进一步支持。 Caveolin-1和-3支架结构域肽对GRK介导的受体和肽底物磷酸化的有效抑制证明了GRK/caveolin相互作用的功能意义。这些数据揭示了GRK调节的新模式，可能在其细胞功能中发挥重要作用。
G protein-coupled receptor kinases (GRKs) have been principally characterized by their ability to phosphorylate and desensitize G protein-coupled receptors, However, recent studies suggest that GRKs may have more diverse protein/protein interactions in cells. Based on the identification of a consensus caveolin binding motif within the pleckstrin homology domain of GRK2, we tested the direct binding of purified full-length GRK2 to various glutathione S-transferase-caveolin-1 fusion proteins, and we discovered a specific interaction of GRK2 with the caveolin scaffolding domain. Interestingly, analysis of GRK1 and GRK5, which lack a pleckstrin homology domain, revealed in vitro binding properties similar to those of GRK2, Maltose-binding protein caveolin and glutathione S-transferase-GRK fusion proteins were used to map overlapping regions in the N termini of both GRK2 and GRK5 that appear to mediate conserved GRK/caveolin interactions. In vivo association of GRK2 and caveolin was suggested by co-fractionation of GRK2 with caveolin in A431 and NIH-3T3 cells and was further supported by co-immunoprecipitation of GRK2 and caveolin in COS-l cells. Functional significance for the GRK/caveolin interaction was demonstrated by the potent inhibition of GRK-mediated phosphorylation of both receptor and peptide substrates by caveolin-1 and -3 scaffolding domain peptides, These data reveal a novel mode for the regulation of GRKs that is likely to play an important role in their cellular function.