Analysis of the regulation mechanisms for the expression of virulence factors in gram-positive cocci.

革兰阳性球菌毒力因子表达调控机制分析。

基本信息

批准号：
14370090
负责人：
OHTA Michio
金额：
$ 7.17万
依托单位：
Nagoya 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

We analyzed the mechanisms of the expression of exoproteins such as toxins and enzymes in Staphylococcus aureus and Streptococcus pyogenes. We performed comprehensive analyses of exoproteins of these bacteria by two dimensional gel electrophoresis (2-DE) and LC/MS/MS and constructed 2-DE maps. We detected putative genes that affect the expression of exoproteins in genome databases and constructed knockout mutants of these genes and analyzed the functions of these genes. The results were as follows.Molecular epidemiological study for 23 exotoxin genes revealed that most Japanese MRSA strains were TSST-1,SEC3 and egc locus-positive, suggesting that these MRSA are very clonal. The differences of the expressed amount of each toxin were analyzed by 2-DE. We found that the expression of TSST-1 was very high in the strains that caused neonatal toxic shock-like exanthematous disease. The production of TSST-1 was suppressed by glucose-mediated catabolite repression. The difference of agr locus … More was not observed in theses trains.We found that the expression of Sic protein was regulated by mga, a global regulator of S.pyogenes, since the production of Sic was reduced in an mga-knockout mutant, and that other several exoproteins were also regulated by mga.It is very likely that the specific secretion machineries are involved in the expression of exoproteins in S.aureus and S.pyogenes, respectively. We found a tat-like gene similar to tat gene of Escherichia coli in the genome of S.aureus. We constructed tat-knockout mutants and found that the expression of several exoproteins were reduced in these mutants, suggesting that this tat-like gene is functional in S.aureus. Interestingly, fat-like gene was found in S.intermedius but not in other coagulase-negative staphylococci. These results suggested that Tat was involved in the virulence of staphylococci.We detected four signal peptidase I (SPaseI)-like genes in the genome of S.pyogenes and constructed knockout mutants of sipC and spi genes. A sipC mutant reduced the expression of 54 exoproteins and a spi mutant reduced the expression of 36 exoproteins. We found preferential expression of some exoproteins in each mutant. Less

我们分析了金黄色葡萄球菌和化脓性链球菌中异蛋白（例如毒素和酶）表达的机制。我们通过二维凝胶电泳（2-DE）和LC/MS/MS进行了对这些细菌的异蛋白的全面分析，并构建了2-DE地图。我们检测到了推定的基因，这些基因影响了基因组数据库中脱甲蛋白的表达，并构建了这些基因的基因敲除突变体，并分析了这些基因的功能。结果如下。对23个外毒素基因的分子流行病学研究表明，大多数日本MRSA菌株都是TSST-1，SEC3和EGC基因座阳性，这表明这些MRSA非常基因。通过2-DE分析了每种毒素的表达量的差异。我们发现，TSST-1的表达在引起新生儿有毒休克样疾病的菌株中非常高。葡萄糖介导的分解代谢物表达抑制TSST-1的产生。在这些火车中没有观察到AGR基因座的差异。我们发现，SIC蛋白的表达受到s.pyogenes的全球调节剂MGA的调节，因为SIC的产生在MGA-KNOCKOUT突变体中降低了，其他几种量蛋白也很可能受到特定的分泌机器的调节。我们发现了一个类似Tat的基因，类似于S.Aureus基因组中大肠杆菌的Tat基因。我们构建了tat-knockout突变体，发现这些突变体中几种脱蛋白的表达降低，这表明该TAT样基因在s. aureus中起作用。有趣的是，在链球菌中发现了类似脂肪的基因，但在其他凝血酶阴性葡萄球菌中未发现。这些结果表明，TAT参与了葡萄球菌病毒。我们检测到S.pyogenes基因组中的四个信号肽I（SPASEI）样基因以及SIPC和SPI基因的构造敲除突变体。 SIPC突变体降低了54种54个脱甲蛋白的表达，而SPI突变体降低了36个Opopotein的表达。我们发现每个突变体中某些脱蛋白的首选表达。较少的