Chemerin C9 peptide induces receptor internalization through a clathrin-independent pathway.
Chemerin C9 peptide induces receptor internalization through a clathrin-independent pathway.
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Chemerin C9 肽通过网格蛋白独立途径诱导受体内化
DOI：
10.1038/aps.2013.198
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发表时间：
2014-05
影响因子：
8.2
通讯作者：
Ye, Richard D.
中科院分区：
文献类型：
--
作者：
Zhou, Jun-xian;Liao, Dan;Zhang, Shuo;Cheng, Ni;He, Hui-qiong;Ye, Richard D.
关键词：
目的：趋化蛋白受体CMKLR1是一种G蛋白偶联受体，广泛存在于单核细胞来源的树突状细胞和巨噬细胞中，在HIV/SIV等免疫缺陷病毒进入淋巴细胞和巨噬细胞的过程中起关键作用。本工作的目的是研究CMKLR1是如何内化的，以及它的内化是否会在体外影响细胞信号转导。方法：使用大鼠嗜碱性白血病RBL-2H3细胞、HEK 293细胞和HeLa细胞。通过共聚焦显微镜成像或使用FACScan流式细胞仪观察CMKLR1的内化。利用定点突变技术，将CMKLR1中6个潜在的磷酸化位点(Ser337、Ser343、Thr352、Ser344、Ser347和Ser350)替换为丙氨酸。结果：在稳定表达CMKLR1的HEK 293细胞和RBL-2H3细胞中，Chmerin和趋化蛋白衍生的九肽(C9)诱导细胞表面CMKLR1-GFP融合蛋白的剂量依赖性丢失，并增加其在细胞内的积聚。经C9(1μ摩尔/L)处理后，CMKLR190%被内化。通过使用不同的试剂，证明了CMKLR1内化过程中存在不依赖于笼蛋白的机制。G蛋白偶联受体激酶磷酸化的Ser343突变和PKC磷酸化的Ser347突变导致CMKLR1内化。CMKLR1内化缺失部分增强了受体信号转导，表现为钙流增加和ERK磷酸化高峰潜伏期缩短。结论：CMKLR1内化是以不依赖于笼蛋白的方式发生的，对受体介导的钙流和ERK磷酸化具有负性调节作用。
Aim:The chemerin receptor CMKLR1 is one type of G protein-coupled receptors abundant in monocyte-derived dendritic cells and macrophages, which plays a key role in the entry of a subset of immunodeficiency viruses including HIV/SIV into lymphocytes and macrophages. The aim of this work was to investigate how CMKLR1 was internalized and whether its internalization affected cell signaling in vitro.Methods:Rat basophilic leukemia RBL-2H3 cells, HEK 293 cells, and HeLa cells were used. CMKLR1 internalization was visualized by confocal microscopy imaging or using a FACScan flow cytometer. Six potential phosphorylation sites (Ser337, Ser343, Thr352, Ser344, Ser347, and Ser350) in CMKLR1 were substituted with alanine using site-directed mutagenesis. Heterologous expression of wild type and mutant CMKLR1 allowed for functional characterization of endocytosis, Ca 2+ flux and extracellular signal-regulated kinase (ERK) phosphorylation.Results:Chemerin and the chemerin-derived nonapeptide (C9) induced dose-dependent loss of cell surface CMKLR1-GFP fusion protein and increased its intracellular accumulation in HEK 293 cells and RBL-2H3 cells stably expressing CMKLR1. Up to 90% of CMKLR1 was internalized after treatment with C9 (1 μmol/L). By using different agents, it was demonstrated that clathrin-independent mechanism was involved in CMKLR1 internalization. Mutations in Ser343 for G protein-coupled receptor kinase phosphorylation and in Ser347 for PKC phosphorylation abrogated CMKLR1 internalization. Loss of CMKLR1 internalization partially enhanced the receptor signaling, as shown by increased Ca 2+ flux and a shorter latency to peak level of ERK phosphorylation.Conclusion:CMKLR1 internalization occurs in a clathrin-independent manner, which negatively regulated the receptor-mediated Ca 2+ flux and ERK phosphorylation.
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DOI：
10.1083/jcb.127.5.1217
发表时间：
1994-12
期刊：
The Journal of cell biology
影响因子：
--
作者：
Schnitzer JE;Oh P;Pinney E;Allard J
通讯作者：
Allard J
DOI：
10.1073/pnas.84.24.8879
发表时间：
1987-12-01
影响因子：
11.1
作者：
BENOVIC, JL;KUHN, H;LEFKOWITZ, RJ
通讯作者：
LEFKOWITZ, RJ
影响因子：
4.8
作者：
Rapacciuolo, A;Suvarna, S;Rockman, HA
通讯作者：
Rockman, HA
DOI：
10.1073/pnas.0710487105
发表时间：
2008-01-08
影响因子：
11.1
作者：
Barnea, Gilad;Strapps, Walter;Lee, Kevin J.
通讯作者：
Lee, Kevin J.
影响因子：
5.5
作者：
Guillabert, Aude;Wittamer, Valerie;Communi, David
通讯作者：
Communi, David