Post-Exponential Phase Gene Expression in Escherichia coli

许多对适应和长期生存至关重要的细菌基因在指数期条件下表达不良。我的实验室已经帮助表征了许多在营养限制条件下表达的适应性基因，特别是那些由大肠杆菌中RpoS替代sigma因子控制的基因。我们已经证明，非必需的rpoS基因虽然对长期生存很重要，但可能在选择过程中丢失，因为rpoS突变体在非首选碳源上具有增强的生长代谢能力。这些研究，结合我们的基因表达研究，将继续是我的实验室的主要重点。我们最近发现，除了RpoS之外，其他鲜为人知的调控过程对长期基因表达也很重要，这些也将被研究。我们将构建缺乏rpos控制功能的靶向组合突变体，利用微阵列、RNA Seq和下一代测序技术来研究它们在细菌代谢中的作用。除了表征rpos依赖基因外，我们还将研究从我们之前的研究中产生的关于rpos依赖类似物表达的具体问题，包括平衡碳利用的关键系统，即转醛醇酶/转酮醇酶。这些固定特异性基因中的每一个在帮助细胞适应应激条件方面可能都很重要，因为我们发现它们只在细胞处于饥饿状态时的固定阶段表达，而在正常条件下它们的表达很差。为了表征在长期（bbb10 - 1天）培养中高度表达的非rpos基因，我们将构建先前鉴定（未发表）的基因的适当缺失突变体，并将使用标准平板试验检查细菌存活率。为了进一步扩展我们的研究，我们还将研究环境细菌分离物的指数后生理学。我们的长期目标是加强我们对细菌如何适应次优生长条件的理解，并验证指数后基因表达的调控模型，这些模型已经使用大肠杆菌实验室菌株开发，用于环境中的天然大肠杆菌群体。

Many bacterial genes that are important for adaptation and long-term survival are poorly-expressed under the exponential phase conditions. My laboratory has helped to characterize many adaptive genes that ares expressed under nutrient limiting conditions, particularly those that are controlled by the RpoS alternative sigma factor in Escherichia coli. We have shown that the non-essential rpoS gene, while important for long-term survival, may be lost under selection as rpoS mutants have enhanced metabolic capability for growth on non-preferred carbon sources. These studies, combined with our gene expression studies, continue to be a major focus for my laboratory. We have recently found that other poorly-understood regulatory processes, in addition to and independent of RpoS, are important for long-term gene expression and these will also be examined. We will construct targeted combinatorial mutants deficient in RpoS-controlled functions to examine the role in bacterial metabolism using microarray, RNA Seq and next generation sequencing. In addition to characterizing RpoS-dependent genes, we will investigate specific questions that have arisen from our previous studies regarding expression RpoS-dependent paralogs including a key system for balance carbon utilization namely Transaldolase/Transketolase. Each of these stationary-specific genes are likely important in helping the cell adapt to stress conditions since we have found that they are expressed only in stationary phase when cells are in a starved state--they are poorly-expressed under normal conditions. To characterize non-RpoS genes that are highly expressed in long-term (> 1 day) cultures, we will construct appropriate deletion mutants of genes that we have previously identified (unpublished) and will examine bacterial survival using standard plate assays. To extend our studies further, we will also examine post exponential physiology in environmental bacterial isolates. Our long term goal is to enhance our understanding of how bacteria adapt to sub-optimal growth conditions and validate regulatory models for post-exponential gene expression, which have been developed using E. coli laboratory strains of , to natural E. coli populations in the environment.  Our short term objectives are to: 1. Examine how expression of the RpoS regulon is controlled by other sigma factors and by the Crl regulatory protein. 2. Identify and characterize the Post-Stationary Phase (PSP) regulon. 3. Examine the physiological role of key RpoS-dependent paralogs in E. coli.