Maeshima Nina, Evans-Atkinson Tara, Hajjar Adeline M, Fernandez Rachel C
From the Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3 and.
Department of Comparative Medicine, University of Washington, Seattle, Washington 98195.
J Biol Chem. 2015 May 22;290(21):13440-53. doi: 10.1074/jbc.M115.653881. Epub 2015 Apr 2.
Lipid A in LPS activates innate immunity through the Toll-like receptor 4 (TLR4)-MD-2 complex on host cells. Variation in lipid A has significant consequences for TLR4 activation and thus may be a means by which Gram-negative bacteria modulate host immunity. However, although even minor changes in lipid A structure have been shown to affect downstream immune responses, the mechanism by which the TLR4-MD-2 receptor complex recognizes these changes is not well understood. We previously showed that strain BP338 of the human pathogen Bordetella pertussis, the causative agent of whooping cough, modifies its lipid A by the addition of glucosamine moieties that promote TLR4 activation in human, but not mouse, macrophages. Using site-directed mutagenesis and an NFκB reporter assay screen, we have identified several charged amino acid residues in TLR4 and MD-2 that are important for these species-specific responses; some of these are novel for responses to penta-acyl B. pertussis LPS, and their mutation does not affect the response to hexa-acylated Escherichia coli LPS or tetra-acylated lipid IVA. We additionally show evidence that suggests that recognition of penta-acylated B. pertussis lipid A is dependent on uncharged amino acids in TLR4 and MD-2 and that this is true for both human and mouse TLR4-MD-2 receptors. Taken together, we have demonstrated that the TLR4-MD-2 receptor complex recognizes variation in lipid A molecules using multiple sites for receptor-ligand interaction and propose that host-specific immunity to a particular Gram-negative bacterium is, at least in part, mediated by very subtle tuning of one of the earliest interactions at the host-pathogen interface.
脂多糖中的脂质A通过宿主细胞上的Toll样受体4（TLR4）-MD-2复合物激活天然免疫。脂质A的变异对TLR4激活具有重大影响，因此可能是革兰氏阴性菌调节宿主免疫的一种方式。然而，尽管脂质A结构的微小变化已被证明会影响下游免疫反应，但TLR4-MD-2受体复合物识别这些变化的机制尚不清楚。我们之前发现，人类病原体百日咳博德特氏菌（百日咳的病原体）的BP338菌株通过添加促进人类而非小鼠巨噬细胞中TLR4激活的葡糖胺部分来修饰其脂质A。通过定点诱变和NFκB报告基因检测筛选，我们在TLR4和MD-2中鉴定出几个对这些物种特异性反应很重要的带电荷氨基酸残基；其中一些对于对五酰化百日咳博德特氏菌脂多糖的反应是新发现的，并且它们的突变不影响对六酰化大肠杆菌脂多糖或四酰化脂质IVA的反应。我们还提供了证据表明，对五酰化百日咳博德特氏菌脂质A的识别依赖于TLR4和MD-2中的不带电荷氨基酸，并且人类和小鼠的TLR4-MD-2受体都是如此。综上所述，我们证明了TLR4-MD-2受体复合物利用多个受体-配体相互作用位点识别脂质A分子的变异，并提出宿主对特定革兰氏阴性菌的特异性免疫至少部分是由宿主-病原体界面最早相互作用之一的非常细微的调节介导的。
