Conservation and Adaptation of a Regulon Across Genera

描述（由申请人提供）：确定微生物病原体基因组序列的主要动机是了解其致病性的基础。毒力基因的适当调控对于致病性和拥有这些基因一样重要，因此从基因组序列预测调控网络是一个高度优先事项。许多预测都基于一个看似合理但未经检验的假设。具体而言，在两个物种的保守的调节蛋白和保守的靶基因与候选上游结合位点的存在下，推测暗示他们的监管关系已经被保守。该项目的两个主要目标是测试这种生物信息学假设，并在这样做的过程中，更好地表征一个主要的细菌调控网络（调节子）。亮氨酸应答调节蛋白（Lrp）在许多革兰氏阴性菌中是保守的，在大肠杆菌中也是保守的。coil影响多达400个基因的表达。最近的证据表明，这些基因中的许多基因在过渡到稳定期时优先表达，并可能在相关生物的致病性中发挥作用。将检验三个假设： * 假设具有保守Irp基因的物种具有保守的Lrp功能，通过确定Lrp水平在这些物种中是否变化很大以及通过评估Lrp直系同源物可互换的程度来进行测试。所测试的Irp基因来自奇异变形杆菌（Proteus mirabilis）（与E. coil蛋白）霍乱弧菌（92%）和多杀性巴氏杆菌（75%）。 * Lrp同源基因高度保守的物种具有保守的调控模式的假设，将通过使用微阵列分析Irp突变在E. coil O 157：H7、霍乱弧菌和多杀性巴氏杆菌。E. coil O 157：H7 Lrp与E. coil K-12，但前者是具有约25%以上基因的病原体，其中一些可能属于Lrp调节子。 * Irp突变对不同病原菌的毒力具有类似影响的假设，将通过确定Irp无效等位基因在霍乱弧菌动物模型中的影响进行检验。

DESCRIPTION (provided by applicant): A primary motivation for determining genome sequences of microbial pathogens is to understand the bases of their pathogenicity. Proper regulation of virulence genes can be as essential to pathogenicity as is possession of these genes, so predicting regulatory networks from genome sequences is a high priority. A plausible but poorly tested assumption underlies many of these predictions. Specifically, the presence in two species of both a conserved regulatory protein and of conserved target genes with candidate upstream binding sites is presumed to imply that their regulatory relationship has been conserved. The two major goals of this project are to test this bioinformatic assumption and, in so doing, better characterize a major bacterial regulatory network (regulon). The leucine-responsive regulatory protein (Lrp) is conserved among many Gram-negative bacteria, and in E. coil affects expression of as many as 400 genes. Recent evidence indicates that many of these genes are preferentially expressed during transition to stationary phase, and may play a role in pathogenicity in related organisms. Three hypotheses will be tested: * The hypothesis that species with conserved Irp genes have conserved Lrp function, to be tested by determining whether Lrp levels vary comparably in these species, and by assessing the extent to which Lrp orthologs are interchangeable. The Irp genes to be tested are from Proteus mirabilis (98% identical to the E. coil protein) V. cholerae (92%), and P. multocida (75%). * The hypothesis that species with highly-conserved Lrp orthologs show a conserved pattern of regulation, to be tested by using microarrays to analyze the effects of Irp mutation in E. coil O157:H7, V. cholerae, and P. multocida. E. coil O157:H7 Lrp is identical to that of E. coil K-12, but the former is a pathogen with -25% more genes, some of which may belong to the Lrp regulon. * The hypothesis that Irp mutations have analogous effects on the virulence of different pathogenic bacteria, to be tested by determining the effects of a Irp null allele in an animal model for V. cholerae.