Essential roles of CD14 and lipopolysaccharide-binding protein for activation of toll-like receptor (TLR)2 as well as TLR4 - Reconstitution of TLR2-and TLR4-activation by distinguishable ligands in LPS preparations
Essential roles of CD14 and lipopolysaccharide-binding protein for activation of toll-like receptor (TLR)2 as well as TLR4 - Reconstitution of TLR2-and TLR4-activation by distinguishable ligands in LPS preparations
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DOI：
10.1046/j.1432-1327.2001.02385.x
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发表时间：
2001-08-01
期刊：
影响因子：
--
通讯作者：
Takeshige, K
中科院分区：
文献类型：
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作者：
Muta, T;Takeshige, K
尽管遗传学研究表明toll样受体（TLR） 4在脂多糖（LPS）的生物学反应中起着关键作用，但异位表达的TLR4和TLR2的活性仍存在争议。我们发现在适当的转染条件下，TLR2和TLR4均介导lps诱导的人胚胎肾293细胞NF-kappaB活化。我们在这里建立的重组系统允许对每个受体的激活进行直接的生化表征和比较。TLR4对LPS的敏感性大约是TLR2的100倍。与对商业LPS制剂的反应相反，TLR2对再纯化的LPS或合成脂质A无反应，表明需要额外的分子。另一方面，脂质a中和试剂多粘菌素B阻断了LPS制备刺激这两种受体的能力，表明脂质a也参与了TLR2的激活。在细胞质区域有点突变的突变TLRs在受到刺激时不能转导信号，并且作为显性负突变体特异性地抑制相应类型受体的激活，而不抑制其他类型受体的激活。因此，这两个受体被可区分的配体独立激活。然而，两种TLRs对LPS制备的反应强烈依赖于血清和CD14，而LPS结合蛋白对于两种受体的激活是必不可少的。支持其功能意义，这两个受体都被发现与CD14相关。
Although genetic studies have revealed a critical role for the toll-like receptor (TLR) 4 in the biological response to lipopolysaccharide (LPS), the activities of ectopically expressed TLR4 and TLR2 are controversial. We have found that under appropriate transfection conditions, both TLR2 and TLR4 mediate LPS-induced NF-kappaB activation in human embryonic kidney 293 cells. The reconstitution systems we established here allow direct biochemical characterization and comparison of activation of each receptor. TLR4 is approximate to 100-fold more sensitive to LPS than TLR2. In contrast to the response to commercial LPS preparations, TLR2 is unresponsive to repurified LPS or synthetic lipid A, indicating the requirement for an additional molecule(s). On the other hand, a lipid A-neutralizing reagent, polymyxin B, blocks the ability of the LPS preparation to stimulate both receptors, suggesting that lipid A is also involved in the activation of TLR2. Mutant TLRs harboring a point mutation in the cytoplasmic domain is inactive in transducing the signal upon stimulation, and act as dominant-negative mutants specifically inhibiting the activation of corresponding type of the receptor but not the other type. Thus, the two receptors are independently activated by distinguishable ligands. Nevertheless, the responses of both TLRs to the LPS preparation are strongly dependent on serum and CD14 and LPS-binding protein are essential for the activation of both of the two receptors. Supporting its functional significance, both receptors are found to associate with CD14.