COORDINATE REGULATION OF BACTERIAL VIRULENCE FACTORS

我们将研究协调调节的遗传基础。 利用三种实验方法确定细菌毒力决定因素 生物-霍乱弧菌、百日咳波氏杆菌和埃希氏菌 Coli.我们的目标是定义分子机制， 每种生物体中的毒力基因组都伴随着 表达的或压抑的。实现这一目标的试验性方法 对于每个被检查的遗传系统都是相似的，并且将 包括以下几个步骤：1.营养与生理分析 毒力调控参数；2.TnPhoA基因的分离 受这些参数控制的融合；3.确认 预期的毒力调节基因控制着 这些融合基因的表达；4.鉴定 通过以下方式选择TnPhoA融合的启动子和控制点 标准方法(即DNA测序、S1和引物延伸 信使核糖核酸图谱、缺失、化学和寡核苷酸导向 突变和蛋白质-DNA结合研究)。霍乱弧菌 我们将扩大我们对ToxR协调调控的分析 系统将包括其他可能的ToxR调节的毒力 性质(即菌毛、OMPS、神经氨酸酶的表达， 蛋白水解酶、溶血和动力)。在百日咳杆菌中，我们将 继续我们对VIR正负坐标的分析 包括启动子结构和基因分析的监管 调制响应。对于霍乱弧菌和大肠杆菌，我们将 研究志贺基因对铁的调节反应 如毒素1型(SltAB)以及其他毒力因子(例如， 溶血素、侵袭性和黏附因子)可能 由它们各自的铁反应毛皮基因产物控制 有机体。在可能的情况下，我们将把我们收集的数据应用于 霍乱、百日咳疫苗的研制与改进。

We will study the genetic basis for the coordinate regulation of bacterial virulence determinants utilizing three experimental organisms-Vibrio cholerae, Bordetella pertussia and Escherchia coli. Our goal is to define the molecular mechanism by which groups of virulence genes in each organism are concomitantly expressed or repressed. The experimental approach to this goal will be similar for each of the genetic systems examined and will include the following steps: 1. Analysis of nutritional and physical parameters that regulate virulence; 2. Isolation of TnphoA gene fusions that are regulated by these parameters; 3. Confirmation that the expected virulence regulatory gene controls the expression of these TnphoA gene fusions; 4. Identification of the promoter and control sites for selected TnphoA fusions by standard methods (i.e., DNA sequencing, S1 and primer extention mRNA mapping, deletion, chemical, and oligonucleotide-directed mutagenesis, and protein-DNA binding studies). For V. cholerae we will expand our analysis of the toxR coordinate regulatory system to include other possible ToxR-regulated virulence properties (i.e., expression of pili, OMPs, neuraminadase, protease, hemolysis, and motility). In B. pertussis we will continue our analysis of vir positive and negative coordinate regulation to include promoter structure and genetic analysis of the modulation response. For V. cholerae and E. coli we will study iron-regulatory responses focusing on the genes for Shiga- like toxin type 1 (sltAB) as well as other virulence factors (e.g., hemolysin, invasive properties, and adherence factors) potentially controlled by the iron-responsive fur gene products of each organism. Where possible, we will apply data we collect to the development and improvement of cholera and pertussis vaccines.