Campylobacter jejuni DNA methylation and gene regulation

描述（由申请方提供）：空肠弯曲杆菌是NIH分类的B类生物恐怖主义制剂。它是美国细菌性胃肠炎的主要原因，并且是散发性疾病以及食源性和水源性爆发的原因。至少有240万例C。空肠胃肠炎的发病率超过沙门氏菌和志贺氏菌的总和（1）。C.空肠感染也是发生格林-巴利综合征的最常见的前因事件，格林-巴利综合征是一种明显由针对C.空肠表面抗原尽管弯曲杆菌病的高患病率和超过20年的研究，C。空肠引起的疾病仍不清楚。几个C。空肠毒力因子已被鉴定，但它们的调节机制在很大程度上是未知的。 最近的证据表明，DNA甲基化是控制沙门氏菌和其他细菌毒力基因表达的重要信号。鉴于此，我们对预测的C.空肠DNA甲基化酶Cj 1461显示cj 1461在37 ℃（人体内部温度）下被诱导，这与致病性生长适应中的作用一致。进一步的实验表明，cj 1461的突变导致ca的表达显著改变。50-60种蛋白质，包括已知的毒力因子和其他预测的调节蛋白，如孤儿反应调节因子Cj 0355。因此，Cj 1461似乎参与了一个复杂的调控回路，控制许多C。jejuni基因;生长温度是信号之一。 我们建议进一步研究C. jejuni，重点是那些蛋白质，预测的DNA甲基化酶Cj 1461和孤儿反应调节Cj 0355的调节。我们将使用微阵列，蛋白质组学和生物化学方法的组合来实现这三个具体目标中概述的目标： 具体目标1。确定负责调节毒力基因表达的Cj 1461和Cj 0355的功能特性。 具体目标2。确定Cj 1461和Cj 0355的下游靶点，以确定它们对基因调控的组合影响。 具体目标3。阐明控制Cj 1461和Cj 0355表达的上游因子和信号，以揭示整个调控回路。

DESCRIPTION (provided by applicant): Campylobacter jejuni is a Category B Bioterrorism Agent as classified by the NIH. It is the leading cause of bacterial gastroenteritis in the United States, and is responsible for sporadic disease as well as food-borne and water-borne outbreaks. There are at least 2.4 million cases of C. jejuni gastroenteritis in the U.S. annually, with an incidence exceeding that of Salmonella and Shigella combined (1). C. jejuni infection is also the most common antecedent event to development of Guillain-Barr Syndrome, an acute motor paralysis apparently resulting from an autoimmune response directed at C. jejuni surface antigens. Despite the high prevalence of Campylobacter disease and more than 20 years of study, the mechanisms by which C. jejuni causes disease remain obscure. Several C. jejuni virulence factors have been identified, yet their regulatory mechanisms are largely unknown. Recent evidence implicates DNA methylation as an important signal controlling virulence gene expression in Salmonella and other bacteria. In light of this, our studies on the predicted C. jejuni DNA methylase Cj1461 showed that cj1461 was induced at 37C, the internal temperature of humans, consistent with a role in pathogenic growth adaptation. Further experiments revealed that mutation of cj1461 resulted in the dramatically altered expression of ca. 50-60 proteins, including known virulence factors and additional predicted regulatory proteins such as the orphan response regulator Cj0355. Consequently, Cj1461 appears to be involved in a complex regulatory circuit controlling expression of numerous C. jejuni genes; growth temperature is one of the signals. We now propose further study of gene regulation in C. jejuni, focusing on those proteins that are regulated by the predicted DNA methylase Cj1461 and the orphan response regulator Cj0355. We will use a combination of microarray, proteomics, and biochemical approaches to achieve the goals outlined in these three specific aims: Specific Aim 1. Determine the functional properties of Cj1461 and Cj0355 responsible for modulating virulence gene expression. Specific Aim 2. Identify the downstream targets of Cj1461 and Cj0355 to define their combinatorial impact on gene regulation. Specific Aim 3. Elucidate the upstream factors and signals controlling Cj1461 and Cj0355 expression to reveal the entire regulatory circuit.