Mastoparan, a G protein agonist peptide, differentially modulates TLR4-and TLR2-mediated signaling in human endothelial cells and murine macrophages
Mastoparan, a G protein agonist peptide, differentially modulates TLR4-and TLR2-mediated signaling in human endothelial cells and murine macrophages
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DOI：
10.4049/jimmunol.174.7.4252
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发表时间：
2005-04-01
影响因子：
4.4
通讯作者：
Arditi, M
中科院分区：
文献类型：
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作者：
Lentschat, A;Karahashi, H;Arditi, M
先前的研究表明，异源三聚体G蛋白偶联信号传导在脂多糖（LPS）刺激的B细胞、单核细胞和巨噬细胞中起作用，并且已显示G蛋白与膜结合型CD14共同免疫沉淀。在本研究中，我们在缺乏膜结合型CD14的人皮肤微血管内皮细胞（HMEC）以及小鼠巨噬细胞中扩展了这些观察结果，以进一步明确异源三聚体G蛋白在Toll样受体（TLR）信号传导中的作用。利用黄蜂毒液衍生的肽——蜂毒肽来破坏G蛋白偶联信号传导，我们在TLR4激动剂刺激的HMEC中确定了一条依赖G蛋白的信号传导通路，该通路对于p38磷酸化和激酶活性的激活、核因子-κB（NF - κB）和白细胞介素 - 6（IL - 6）的反式激活以及IL - 6的分泌是必需的。相比之下，TLR2激动剂、肿瘤坏死因子 - α（TNF - α）或白细胞介素 - 1β（IL - 1β）对HMEC的激活对蜂毒肽不敏感。在小鼠巨噬细胞系RAW 264.7以及原代小鼠巨噬细胞中，蜂毒肽引起的G蛋白失调导致LPS诱导的信号传导受到显著抑制，从而影响MyD88依赖和MyD88非依赖的基因表达，而TLR2介导的基因表达未受到显著抑制。除了抑制巨噬细胞中TLR4介导的丝裂原活化蛋白激酶（MAPK）磷酸化外，蜂毒肽还减弱了LPS诱导的白细胞介素 - 1受体相关激酶 - 1（IL - 1R - associated kinase - 1）的激酶活性，但不影响TLR2激动剂诱导的该激酶活性，然而它不影响磷脂酰肌醇 - 3 - 激酶（phospboinositol - 3 - kinase）在TLR2或TLR4介导的信号传导中诱导的Akt磷酸化。这些数据证实了异源三聚体G蛋白在使用可溶性或膜相关CD14的细胞中TLR4介导的反应中的重要性，并揭示了一种此前未被重视的Toll样受体和信号传导通路特异性水平。
Previous studies have implicated a role for heterotrimeric G protein-coupled signaling in B cells, monocytes, and macrophages stimulated with LPS and have shown that G proteins coimmunoprecipitate with membrane-bound CD14. In this study, we have extended these observations in human dermal microvessel endothelial cells (HMEC) that lack membrane-bound CD14 and in murine macrophages to define further the role of heterotrimeric G proteins in TLR signaling. Using the wasp venom-derived peptide, mastoparan, to disrupt G protein-coupled signaling, we identified a G protein-dependent signaling pathway in HMEC stimulated with TLR4 agonists that is necessary for the activation of p38 phosphorylation and kinase activity, NF-kappa B and IL-6 transactivation, and IL-6 secretion. In contrast, HMEC activation by TLR2 agonists, TNF-alpha, or IL-1 beta was insensitive to mastoparan. In the murine macrophage cell line, RAW 264.7, and in primary murine macrophages, G protein dysregulation by mastoparan resulted in significant inhibition of LPS-induced signaling leading to both MyD88-dependent and MyD88-independent gene expression, while TLR2-mediated gene expression was not significantly inhibited. In addition to inhibition of TLR4-mediated MAPK phosphorylation in macrophages, mastoparan blunted IL-1R-associated kinase-1 kinase activity induced by LPS, but not by TLR2 agonists, yet failed to affect phosphorylation of Akt by phospboinositol-3-kinase induced by either TLR2- or TLR4-mediated signaling. These data confirm the importance of heterotrimeric G proteins in TLR4-mediated responses in cells that use either soluble or membrane-associated CD14 and reveal a level of TLR and signaling pathway specificity not previously appreciated.