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Molecular genetic studies on iron-starvation stress response of Vibrio species and its involvement in pathogenesis

弧菌铁饥饿应激反应及其发病机制的分子遗传学研究

基本信息

批准号：
10670257
负责人：
YAMAMOTO Shigeo
金额：
$ 1.92万
依托单位：
Okayama University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

1. V.parahaemolyticus strains isolated from different sources were assayed for their ability to produce a siderophore, vibrioferrin (VF), under iron-limited conditions. VF production of clinical isolates was significantly higher than those for food and coastal isolates, suggesting that greater producitivity of VF by clinical isolates may be associated with a selective advantage for survival and proliferation under conditions of iron-limitation such as in the intestine.2. V.parahaemolyticus strains carrying mutations in the fur gene were isolated by the Mn selection method to assess the function of Fur in connection with alterations in the coordinate expression of the siderophore VF and iron-repressive outer membrane proteins (IROMPs). Ten of 25 mutants constitutively produce VF and expressed at least two IROMPs. DNA sequencing of the fur genes identified 4 different point mutations causing amino acid change. Western blotting using anti-Fur antiserum revealed that the intracellular abun … More dance of the wild-type Fur was not significantly affected by the iron concentration in the growth medium. Fur regulon was identified for the first time in V.parahaemolyticus.3. Many kinds of iron-regulated gene fragments containing a putative Fur box were isolated from pathogenic vibrios by their nucleotide sequencing. The full-length genes encoding three IROMPs in V.parahaemolyticus were cloned and analyzed. Also, the aerobactin operon and heme receptor were cloned and analyzed from V.mimicus. The genetic organization of the former was unusual in that it contains the genes responsible for the iron transport system (fhuBCD) in addition to the genes encoding biosynthesis enzymes (iucABCD) and an outer membrane receptor (iutA).4. Iron-repressible sodA genes encoding MnSOD were cloned from three vibrio species, and the Fur boxes were identified in their promoters. These MnSODs were produced in response to iron-limitation.5. One of the genes responsible for VF biogenesis was identified by insertion mutagenesis using a suicide vector. The insertion mutagensis was applied to the above-described V.parahaemolyticus genes to clarify their functions.6. The genes encoding ATP-dependent protease were isolated from three vibrio species. It is of great interest to clarify whether the iron-regulated protease expression is associated with decomposition of Fur in response to iron starvation. Less

1.从不同来源分离的副溶血性弧菌菌株测定其在铁限制条件下产生铁载体、弧菌铁蛋白（VF）的能力。临床分离株的VF产量显著高于食品和沿海分离株，这表明临床分离株的VF产量较高可能与铁限制条件下（如在甜菜碱中）存活和增殖的选择性优势有关.通过Mn选择方法分离在Fur基因中携带突变的副溶血性弧菌菌株，以评估Fur与铁载体VF和铁抑制性外膜蛋白（IROMP）的协调表达的改变相关的功能。25个突变体中有10个组成型产生VF并表达至少两种IROMP。皮毛基因的DNA测序鉴定出4种不同的点突变，导致氨基酸变化。使用抗Fur抗血清的Western印迹显示，细胞内的Fur抗体水平与细胞内的Fur抗体水平呈负相关，而Fur抗体水平与细胞内的Fur抗体水平呈负相关。 ...更多信息野生型Fur的舞蹈不受生长培养基中铁浓度的显著影响。首次在副溶血性弧菌中鉴定出皮毛调节子。从致病性弧菌中分离出多种含Fur盒的铁调控基因片段。对副溶血性弧菌三种IROMP基因进行了克隆和序列分析。此外，从拟态弧菌中克隆并分析了需氧肌动蛋白操纵子和血红素受体。前者的遗传组织是不寻常的，因为它除了编码生物合成酶（iucABCD）和外膜受体（iutA）的基因外，还含有负责铁转运系统（fhuBCD）的基因。从三种弧菌中克隆了编码MnSOD的铁抑制性sodA基因，并在其启动子中鉴定了Fur盒。这些MnSOD的产生是对铁限制的响应。通过使用自杀载体的插入诱变来鉴定负责VF生物发生的基因之一。对上述副溶血性弧菌基因进行插入诱变，以阐明其功能.从三种弧菌中分离出编码ATP依赖蛋白酶的基因。这是非常感兴趣的，以澄清是否铁调节蛋白酶的表达与毛皮的分解响应铁饥饿。少