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。对毒力基因的适当调控与拥有这些基因一样，对致病性至关重要，因此从基因组序列预测调控网络是一个高度优先考虑的问题。一个看似合理但未经验证的假设构成了许多预测的基础。具体来说，在两个物种中同时存在一个保守的调控蛋白和具有候选上游结合位点的保守靶基因，这意味着它们的调控关系是保守的。该项目的两个主要目标是测试这种生物信息学假设，并在此过程中更好地表征主要的细菌调节网络（regulon）。亮氨酸反应调节蛋白（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，这些基因中的许多在向静止期过渡时优先表达，并可能在相关生物的致病性中发挥作用。将测试三个假设：

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.