BACKGROUND & AIMS:
:The chemokine receptor CXCR3 is expressed on the surface of both resting and activated T lymphocytes. We describe in this study the endocytosis of CXCR3 using T lymphocytes and CXCR3 transfectants. Chemokine-induced CXCR3 down-regulation occurred in a rapid, dose-dependent manner, with CXCL11 the most potent and efficacious ligand. Endocytosis was mediated in part by arrestins, but appeared to occur independently of clathrin and caveolae. In contrast to other chemokine receptors, which are largely recycled to the cell surface within an hour, cell surface replenishment of CXCR3 occurred over several hours and was dependent upon mRNA transcription, de novo protein synthesis, and transport through the endoplasmic reticulum and Golgi. Confocal microscopy and Western blotting confirmed the fate of endocytosed CXCR3 to be degradation, mediated in part by lysosomes and proteosomes. Site-directed mutagenesis of the CXCR3 C terminus revealed that internalization and degradation were independent of phosphorylation, ubiquitination, or a conserved LL motif. CXCR3 was found to be efficiently internalized in the absence of ligand, a process involving a YXXL motif at the extreme of the C terminus. Although freshly isolated T lymphocytes expressed moderate cell surface levels of CXCR3, they were only responsive to CXCL11 with CXCL9 and CXCL10 only having significant activity on activated T lymphocytes. Thus, the activities of CXCR3 are tightly controlled following mRNA translation. Because CXCR3(+) cells are themselves a source of IFN-gamma, which potently induces the expression of CXCR3 ligands, such tight regulation of CXCR3 may serve as a control to avoid the unnecessary amplification of activated T lymphocyte recruitment.
背景与目标:
: 趋化因子受体CXCR3在静止和活化的T淋巴细胞表面均表达。在这项研究中,我们使用T淋巴细胞和CXCR3转染子描述了CXCR3的内吞作用。趋化因子诱导的CXCR3下调以快速,剂量依赖性方式发生,其中CXCL11是最有效的配体。内吞作用部分由抑制素介导,但似乎独立于网格蛋白和小凹而发生。与其他趋化因子受体 (在一小时内大量回收到细胞表面) 相反,CXCR3的细胞表面补充发生了几个小时,并且取决于mRNA转录,从头蛋白质合成以及通过内质网和高尔基体的转运。共聚焦显微镜和Western印迹证实了内吞CXCR3降解的命运,部分由溶酶体和蛋白质体介导。CXCR3 C末端的定点诱变表明,内化和降解与磷酸化,泛素化或保守的LL基序无关。发现CXCR3在没有配体的情况下有效内化,该过程涉及C末端的YXXL基序。尽管新鲜分离的T淋巴细胞表达中等水平的CXCR3细胞表面,但它们仅对CXCL11有反应,而CXCL9和CXCL10仅对活化的T淋巴细胞具有显着活性。因此,CXCR3的活性在mRNA翻译后受到严格控制。由于CXCR3 () 细胞本身是IFN-γ 的来源,可以有效地诱导CXCR3配体的表达,因此对CXCR3的这种严格调节可以作为一种控制,以避免不必要的活化T淋巴细胞募集的扩增。