Ubiquitination differentially regulates clathrin-dependent internalization of protease-activated receptor-1.
Ubiquitination differentially regulates clathrin-dependent internalization of protease-activated receptor-1.
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DOI：
10.1083/jcb.200610154
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发表时间：
2007-06-04
期刊：
影响因子：
--
通讯作者：
Trejo J
中科院分区：
文献类型：
--
作者：
Wolfe BL;Marchese A;Trejo J
蛋白酶激活受体1（PAR 1）是凝血酶的G蛋白偶联受体（GPCR），通过蛋白水解被不可逆地激活。因此，PAR 1运输对于凝血酶信号传导的保真度至关重要。PAR 1显示组成性和激动剂诱导的内化，这是网格蛋白和发动蛋白依赖，但不依赖于arrestins。网格蛋白接头AP 2（接头蛋白复合物-2）是组成型的关键，但不是激活PAR 1内化。在这项研究中，我们发现，泛素化负调控PAR 1组成性内化，并指定一个独特的网格蛋白适配器激活受体内化的要求。PAR 1在活化后基本上是泛素化和去泛素化的。PAR 1无赖氨酸突变体信号正常，但没有泛素化。泛素（Ub）缺陷型PAR 1的组成性内化显着增加，并通过Ub与胞质尾的融合来抑制。Ub缺陷型PAR 1组成性内化与野生型受体一样依赖于AP 2。然而，与野生型PAR 1不同，AP 2是激活的Ub缺陷型受体内化所必需的，这表明泛素化PAR 1的内化需要不同的内吞机制。这些研究揭示了泛素化在GPCR内化调节中的新功能。
Protease-activated receptor-1 (PAR1), a G protein–coupled receptor (GPCR) for thrombin, is irreversibly activated by proteolysis. Consequently, PAR1 trafficking is critical for the fidelity of thrombin signaling. PAR1 displays constitutive and agonist-induced internalization, which are clathrin and dynamin dependent but are independent of arrestins. The clathrin adaptor AP2 (adaptor protein complex-2) is critical for constitutive but not for activated PAR1 internalization. In this study, we show that ubiquitination negatively regulates PAR1 constitutive internalization and specifies a distinct clathrin adaptor requirement for activated receptor internalization. PAR1 is basally ubiquitinated and deubiquitinated after activation. A PAR1 lysineless mutant signaled normally but was not ubiquitinated. Constitutive internalization of ubiquitin (Ub)-deficient PAR1 was markedly increased and inhibited by the fusion of Ub to the cytoplasmic tail. Ub-deficient PAR1 constitutive internalization was AP2 dependent like the wild-type receptor. However, unlike wild-type PAR1, AP2 was required for the internalization of activated Ub-deficient receptor, suggesting that the internalization of ubiquitinated PAR1 requires different endocytic machinery. These studies reveal a novel function for ubiquitination in the regulation of GPCR internalization.
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