Regulation of Stationary Phase in Escherichia coli

描述（由申请人提供）：替代σ因子RpoS是大肠杆菌和许多其他变形菌中稳定期和一般应激反应的主要调节因子。在快速生长的细胞中，产生RpoS，但它被ClpX/P蛋白酶降解。当饥饿的特定营养素，RpoS合成增加，降解停止，活动增加，或这些影响的一些组合发生。两种新的调节蛋白是该提议的焦点，SprE（RssB）和Crl。SprE是一种孤儿反应调节剂，其在生长的细胞中充当将RpoS引导至ClpX/P蛋白酶的衔接子。当碳源耗尽时，这种降解停止。我们知道SprE磷酸化/去磷酸化不参与饥饿信号传导。遗传分析表明，饥饿感测为ATP水平的降低，生物化学数据表明，这种降低是由ClpX和RpoS本身感测的。遗传分析进一步揭示了SprE在多聚腺苷酸化和mRNA稳定性控制中的第二种功能，并且它表明SprE的磷酸化对于这种活性是重要的。我们将鉴定分离SprE两种功能的突变，并鉴定相关的小分子或激酶。使用质谱法，我们已经发现SprE控制Poly（A）聚合酶和Hfq与mRNA降解体的缔合，并且来自微阵列的数据表明SprE的功能是沉默外源基因。我们将探讨这些新活动的生理意义。我们已经表明，Crl促进RpoS与核心RNA聚合酶的关联，我们知道这种活性在氮饥饿条件下尤其重要。在这些条件下，crl转录增加25倍。然而，Crl水平没有显著的相应变化。我们证明，这种变化的转录允许开关从嘈杂的更均匀的Crl生产，我们将探测这些表达模式在这两种条件下的生理意义。营养饥饿是细菌面临的最常见的胁迫，RpoS可以说是大肠杆菌中最重要的全局调节蛋白。杆菌更好地了解稳定期生理学可能会为如何对抗细菌感染提供见解。

DESCRIPTION (provided by applicant): The alternate sigma factor RpoS is the master regulator of stationary phase and the general stress response in Escherichia coli and many other proteobacteria. In rapidly growing cells, RpoS is made but it is degraded by the ClpX/P protease. When starved for a particular nutrient, RpoS synthesis increases, degradation ceases, activity is increased, or some combination of these effects occurs. Two novel regulatory proteins are the focus of this proposal, SprE (RssB) and Crl. SprE is an orphan response regulator that functions as an adaptor to direct RpoS to the ClpX/P protease in growing cells. When carbon sources are depleted, this degradation stops. We know that SprE phosphorylation/dephosphorylation is not involved in starvation signaling. Genetic analysis suggests that starvation is sensed as a decrease in ATP levels, and biochemical data suggest that this decrease is sensed by ClpX and RpoS itself. Genetic analysis has further revealed a second function for SprE in polyadenylation and the control of mRNA stability, and it suggests that phosphorylation of SprE is important for this activity. We will identify mutations that separate the two functions of SprE and identify the relevant small molecule or kinase. Using mass spectrometry we have discovered that SprE controls the association of Poly(A) polymerase and Hfq with the mRNA degradosome and data from microarrays suggest that SprE functions to silence foreign genes. We will probe the physiological significance of these novel activities. We have shown that Crl facilitates the association of RpoS with core RNA polymerase, and we know that this activity is especially important under nitrogen starvation conditions. Under these conditions crl transcription increases 25 fold. However, there is no significant corresponding change in Crl levels. We demonstrate that this change in transcription allows a switch from noisy to more uniform Crl production and we will probe the physiological significance of these expression patterns under both conditions. Nutrient starvation is the most common stress that bacteria face and RpoS is arguably the most important global regulatory protein in E. coli. A better understanding of stationary phase physiology may provide insights into how to combat bacterial infections.