Functional and structural consequences of chemokine (C-X-C motif) receptor 4 activation with cognate and non-cognate agonists
Functional and structural consequences of chemokine (C-X-C motif) receptor 4 activation with cognate and non-cognate agonists
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DOI：
10.1007/s11010-017-3044-7
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发表时间：
2017-10-01
影响因子：
4.3
通讯作者：
Majetschak, Matthias
中科院分区：
文献类型：
--
作者：
Eby, Jonathan M.;Abdelkarim, Hazem;Majetschak, Matthias
趋化因子受体4（CXCR4）调节细胞运输，在免疫系统中起重要作用。泛素最近被鉴定为CXCR 4的内源性非同源激动剂，其通过与同源激动剂C-X-C基序趋化因子配体12（CXCL 12）的相互作用位点不同的相互作用位点激活CXCR 4。与CXCL12相比，泛素在原代人细胞中的趋化活性的特征很差。此外，缺乏CXCR4激动剂的功能选择性的证据，并且泛素与CXCR4结合的结构后果是未知的。在这里，我们表明，泛素和CXCL12在正常人外周血单核细胞，单核细胞，血管平滑肌和内皮细胞中具有相当的趋化活性。CXCR4配体的趋化活性可以用选择性CXCR4拮抗剂AMD 3100和CXCR4第二跨膜结构域的肽类似物抑制。在人单核细胞中，泛素和CXCL12诱导的趋化性可以被百日咳毒素和磷脂酶C、磷脂酰肌醇3激酶和细胞外信号调节激酶12的抑制剂抑制。两种激动剂均诱导血管平滑肌细胞产生三磷酸肌醇，AMD 3100可抑制这一过程。在β-抑制蛋白募集试验中，泛素不能充分地将β-抑制蛋白2募集到CXCR 4（EC50> 10 μ M），而CXCL 12的EC50为4.6 nM（95%置信区间为3.1 - 6.1 nM）。两种激动剂在膜中CXCR 4的C-13-H-1-杂原子单量子相关（HSQC）光谱中诱导了相似的化学位移变化，而CXCL 11没有显著改变CXCR 4的C-13-H-1-HSQC光谱。我们的研究结果指向泛素作为CXCR4的偏向性激动剂。
Chemokine (C-X-C motif) receptor 4 (CXCR4) regulates cell trafficking and plays important roles in the immune system. Ubiquitin has recently been identified as an endogenous non-cognate agonist of CXCR4, which activates CXCR4 via interaction sites that are distinct from those of the cognate agonist C-X-C motif chemokine ligand 12 (CXCL12). As compared with CXCL12, chemotactic activities of ubiquitin in primary human cells are poorly characterized. Furthermore, evidence for functional selectivity of CXCR4 agonists is lacking, and structural consequences of ubiquitin binding to CXCR4 are unknown. Here, we show that ubiquitin and CXCL12 have comparable chemotactic activities in normal human peripheral blood mononuclear cells, monocytes, vascular smooth muscle, and endothelial cells. Chemotactic activities of the CXCR4 ligands could be inhibited with the selective CXCR4 antagonist AMD3100 and with a peptide analogue of the second transmembrane domain of CXCR4. In human monocytes, ubiquitin- and CXCL12-induced chemotaxis could be inhibited with pertussis toxin and with inhibitors of phospholipase C, phosphatidylinositol 3 kinase, and extracellular signal-regulated kinase 1/2. Both agonists induced inositol trisphosphate production in vascular smooth muscle cells, which could be inhibited with AMD3100. In beta-arrestin recruitment assays, ubiquitin did not sufficiently recruit beta-arrestin2 to CXCR4 (EC50 > 10 mu M), whereas the EC50 for CXCL12 was 4.6 nM (95% confidence interval 3.1-6.1 nM). Both agonists induced similar chemical shift changes in the C-13-H-1-heteronuclear single quantum correlation (HSQC) spectrum of CXCR4 in membranes, whereas CXCL11 did not significantly alter the C-13-H-1-HSQC spectrum of CXCR4. Our findings point towards ubiquitin as a biased agonist of CXCR4.