Subtype-specific regulation of receptor internalization and recycling by the carboxyl-terminal domains of the human A1 and rat A3 adenosine receptors:: Consequences for agonist-stimulated translocation of arrestin3
Subtype-specific regulation of receptor internalization and recycling by the carboxyl-terminal domains of the human A1 and rat A3 adenosine receptors:: Consequences for agonist-stimulated translocation of arrestin3
复制标题
DOI：
10.1021/bi0262911
复制
发表时间：
2002-12-17
期刊：
影响因子：
2.9
通讯作者：
Palmer, TM
中科院分区：
文献类型：
--
作者：
Ferguson, G;Watterson, KR;Palmer, TM
在这项研究中，我们的特点是不同的影响，抑制腺苷受体（AR）贩运破坏预测网站棕榈酰化和磷酸化内每个受体的羧基末端。虽然Cys（302，305）Ala突变的大鼠A（3）AR突变体在响应激动剂暴露时比野生型（WT）受体更快地内化，但人类A（1）AR的类似突变（CyS（309）Ala）对受体内化没有影响。此外，与WT A3AR不同，在激动剂去除后，内化的突变体A（3）AR的整个池能够再循环回到质膜。这些特性并不反映替代运输途径的利用，因为内化的WT和突变A（3）AR都累积到转铁蛋白受体阳性内体区室中。然而，受体在核内体中的蓄积依赖于先前G蛋白偶联受体激酶（GRK）介导的受体羧基末端磷酸化，因为用大鼠A（3）AR的14个GRK磷酸化氨基酸取代人A（1）AR的羧基末端结构域可使所得嵌合A（1）CT3AR在激动剂介导的核内体中快速蓄积。对GRK介导的磷酸化的敏感性也决定了激动剂暴露后观察到的arrestin3的不同再分布。因此，当非磷酸化的A（1）AR将arrestin3从细胞质重新分布到质膜上的点状聚集体时，GRK磷酸化的WT和Cys（302，305）Ala突变的A（3）AR以及A（1）CT3AR嵌合体均诱导arrestin3重新分布到质膜和细胞质内的点状聚集体中。无论是人类AIAR还是大鼠A3 AR与arrestin3在基础或激动剂刺激条件下共定位。总之，这些结果表明抑制AR介导的arrestin3分布的变化是亚型特异性的，特异性与受体羧基末端结构域对GRK磷酸化的敏感性相关。在大鼠A3AR的情况下，GRK介导的内化的敏感性似乎部分由其GRK磷酸受体位点上游的假定棕榈酸酯附着位点的完整性调节。
In this study, we have characterized the differential effects on inhibitory adenosine receptor (AR) trafficking of disrupting predicted sites for palmitoylation and phosphorylation within each receptor's carboxyl terminus. While a Cys(302,305)Ala-mutated rat A(3)AR mutant internalizes significantly faster than the wild-type (WT) receptor in response to agonist expo sure, analogous mutation of the human A(1)AR (CyS(309)Ala) had no effect on receptor internalization. Moreover, unlike the WT A3AR, the entire pool of internalized mutant A(3)AR is able to recycle back to the plasma membrane following agonist removal. These properties do not reflect utilization of an alternative trafficking pathway, as internalized WT and mutant A(3)ARs both accumulate into transferrin receptor-positive endosomal compartments. However, receptor accumulation into endosomes is dependent upon prior G-protein-coupled receptor kinase (GRK)-mediated phosphorylation of the receptor's carboxyl terminus, as replacement of the carboxyl-terminal domain of the human A(1)AR with the 14 GRK-phosphorylated amino acids of the rat A(3)AR confers rapid agonist-mediated endosomal accumulation of the resulting chimeric A(1)CT3AR. Sensitivity to GRK-mediated phosphorylation also dictates the distinct redistribution of arrestin3 observed upon agonist exposure. Thus, while the nonphosphorylated A(1)AR redistributes arrestin3 from the cytoplasm to punctate clusters at the plasma membrane, GRK-phosphorylated WT and Cys(302,305)Ala-mutated A(3)ARs, as well as the A(1)CT3AR chimera, each induce the redistribution of arrestin3 into punctate accumulations both at the plasma membrane and within the cytoplasm. Neither the human AIAR nor the rat A3AR colocalized with arrestin3 under basal or agonist-stimulated conditions. Together, these results demonstrate that inhibitory AR-mediated changes in arrestin3 distribution are subtype-specific, with specificity correlating with the sensitivity of the receptor's carboxyl-terminal domain to GRK phosphorylation. In the case of the rat A3AR, sensitivity to GRK-mediated internalization appears to be regulated in part by the integrity of putative palmitate attachment sites upstream of its GRK phosphoacceptor sites.