Molecular mechanism of biological activity expressed by hemolysins derived from Listeriae inside macrophages.

巨噬细胞内李斯特菌溶血素表达的生物活性的分子机制。

基本信息

批准号：
14370092
负责人：
MITSUYAMA Masao
金额：
$ 8.06万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

On the molecular mechanisms of biological activity expressed by hemolysins derived from various Listeria spp., the following results have been obtained by using recombinant proteins and mutant strains. (1)All the hemolysin proteins, LLO, ILO and LSO, consist of 4 domains and the known hemolytic activity is highly dependent on domain 4, while the newly found cytokine-inducing activity is exclusively dependent on domain 1-3 at he N-terminus. (2)The very low cytokine-inducing activity observed in ILO appeared to be due to the absence of PEST-like sequence at the N-terminus portion. (3)Upon stimulation with LLO, macrophage first produce IL-12 and IL-18, then these IFN-γ-inducing cytokines stimulate NK cells to produce a massive amount of IFN-γ. (4)TLRs, especially TLR2 and TLR4 are both required for the cytokine production by macrophages in response to LLO. (5)The cytokine response in epithelial cells that do not express surface TLRs appeared to be due to the cytokine induction triggered by the Ca-influx caused by sublethal level of membrane damage by LLO. (6)The in-flame deletion mutant of hly (a gene encoding LLO) has been established. By using recombinant strains, it was confirmed that both escape of bacteria from phagosome into cytosolic space in macrophages and the presence of PEST-like sequence are required for cytokine response. (7)Upon infection of macrophages by Listeria monocytogenes, a significant level of upregulation was induced in hly and orfA gene expression. Oxygen radicals produced by infected macrophages appeared to be responsible for this upregulation of virulence gene expression.

关于由源自各种李斯特氏菌的血素表达的生物学活性的分子机制，通过使用重组蛋白和突变菌株获得了以下结果。（1）所有溶血素蛋白，LLO，ILO和LSO，由4个结构域组成，已知的溶血活性高度依赖于结构域4，而新发现的细胞因子诱导的活性仅取决于N-末端的域1-3。（2）在ILO中观察到的非常低的细胞因子诱导的活性似乎是由于N末端部分没有害虫样序列。（3）用LLO刺激，巨噬细胞首次产生IL-12和IL-18，这些IFN-γ诱导的细胞因子会刺激NK细胞产生大量的IFN-γ。（4）TLR，尤其是TLR2和TLR4，巨噬细胞对LLO的细胞因子产生都需要。（5）不表达表面TLR的上皮细胞中的细胞因子反应似乎是由于由LLO通过llo造成的膜损伤造成的CA-炎症引起的细胞因子诱导。（6）已经建立了HLY的Flame内缺失突变体（编码LLO的基因）。通过使用重组菌株，可以证实细菌在巨噬细胞中从吞噬体逃脱到胞质空间，并且需要细胞因子反应的存在类似害虫样序列。（7）在通过单核细胞增生李斯特菌感染巨噬细胞后，在HLY和ORFA基因表达中诱导了显着的上调。被感染的巨噬细胞产生的氧自由基似乎是病毒基因表达的上调。