The Function analysis of MD-2 molecule about LPS recognition

MD-2分子对LPS识别的功能分析

• 批准号: 13670460
• 负责人: AKASHI Sachiko
• 金额: $ 2.62万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13670460/
• 关键词: MD-2; TLR4; LPS; CD14; LipdIVa; E5531; RP105; MD-1

Lipopolysaccharide (LPS), a membrane constituent of Gram-negative bacteria, is the best-studied ligand that potently activates the immune system and induces endotoxin shock. Recently we discovered MD-2 molecule, which is associated with TLR4 extracellular domain, and we showed that TLR4/MD-2 complex is indispensable for LPS recognition and response, but it is not clear how TLR4/MD-2 recognizes LPS. Firstly to address the role played by MD-2 in vivo, we generated MD-2-/- mice. B cells, Macrophages, and Dendritic cells of MD-2-/- mice did not respond to LPS. And We showed that MD-2-/- mice did survive endotoxic shock but were susceptible to Salmonella typhimurium infection. We found that in MD-2-/- embryonic fibroblasts, TLR4 was not able to reach the plasma membrane and predominantly resided in the Golgi apparatus, whereas TLR4 was distributed at the leading edge surface of cells in wild-type embryonic fibroblasts. Next we showed that MD-2 directly regulates LPS recognition by TLR4. To address the issue, we took advantage of species-specific pharmacology of lipidIVa, an analogue of lipidA. LipidIVa acted agonistically on mouse(m)TLR4/MD-2 but not on human(h)TLR4/MD-2. LipidIVa antagonized the agonistic effect of lipidA on hTLR4/MD-2. We examined the chimeric complex consisting of mTLR4 and hMD-2 to ask whether species specificity is coinferred by TLR4 or MD-2. hMD-2 conferred on mTLR4 responsiveness to lipidA but not to lipidIVa. Moreover, lipidIVa acted as a lipidA antagonist on mTLR4 that is associated with hMD-2. Collectively, MD-2 directly influences the fine specificity of TLR4.

脂多糖（LPS）是革兰氏阴性细菌的一种膜成分，是研究最佳的配体，可有效激活免疫系统并诱导内毒素休克。最近，我们发现了与TLR4细胞外结构域相关的MD-2分子，我们表明TLR4/MD-2复合物对于LPS识别和响应是必不可少的，但是尚不清楚TLR4/MD-2如何识别LPS。首先，要解决MD-2在体内扮演的角色，我们生成了MD-2 - / - 小鼠。 MD-2 - / - 小鼠的B细胞，巨噬细胞和树突状细胞对LPS无反应。我们表明，MD-2 - / - 小鼠确实在内毒性休克中存活，但易受伤寒沙门氏菌感染的影响。我们发现，在MD-2 - / - 胚胎成纤维细胞中，TLR4无法到达质膜，并且主要驻留在高尔基体中，而TLR4则分布在野生型胚胎成纤维纤维细胞中细胞的前沿表面。接下来，我们表明MD-2直接调节TLR4的LPS识别。为了解决这个问题，我们利用了Lipidiva的物种特异性药理学，这是Lipida的类似物。 Lipidiva对小鼠（M）TLR4/MD-2的作用作用，而不是人（H）TLR4/MD-2。 Lipidiva拮抗了Lipida对HTLR4/MD-2的激动作用。我们检查了由MTLR4和HMD-2组成的嵌合复合物，以询问物种特异性是否由TLR4或MD-2共同授予。 HMD-2赋予MTLR4对Lipida的响应性，但对Lipidiva不反应。此外，Lipidiva在MTLR4上充当了与HMD-2相关的Lipida拮抗剂。总体而言，MD-2直接影响TLR4的良好特异性。

项目成果

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