Post-Exponential Phase Gene Expression in Escherichia coli

许多对适应和长期生存很重要的细菌基因在指数期条件下表达很差。我的实验室帮助确定了许多在营养限制条件下表达的适应性基因的特征，特别是那些由大肠杆菌中的rpos替代西格玛因子控制的基因。我们已经证明，非必需的rpos基因，虽然对长期生存很重要，但在选择下可能会丢失，因为rpos突变体增强了在非首选碳源上生长的代谢能力。这些研究，结合我们的基因表达研究，仍然是我的实验室的主要关注点。我们最近发现，除了rpos之外，其他知之甚少的调控过程对于长期的基因表达也很重要，这些也将被研究。我们将构建缺乏rpos调控功能的靶向组合突变体，利用微阵列、RNA序列和下一代测序来研究其在细菌代谢中的作用。除了研究依赖于rpos的基因外，我们还将研究以前关于依赖于rpos的基因表达的研究中出现的一些具体问题，其中包括一个平衡碳利用的关键系统，即转醛醇酶/转酮醇酶。这些稳定的特定基因中的每一个在帮助细胞适应压力条件方面都可能是重要的，因为我们已经发现，它们只在细胞处于饥饿状态时的静止阶段表达--它们在正常条件下表达很低。为了确定在长期(1天)培养中高表达的非rpos基因的特征，我们将构建我们之前已确定(未发表)的基因的适当缺失突变体，并将使用标准平板分析来检查细菌的存活率。为了进一步扩展我们的研究，我们还将检查环境细菌分离株的指数后生理学。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，这些模型是使用大肠杆菌和实验室菌株开发的，以适应环境中的自然大肠杆菌种群。*我们的短期目标是：*1.研究rpos调节子的表达如何受其他sigma因子和CRL调节蛋白的控制。*2.识别和表征后稳定期(PSP)调节子。*3.研究关键的rpos依赖蛋白在大肠杆菌中的生理作用。

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.  *** **