Regulation of Stationary Phase in Escherichia coli

大肠杆菌固定相的调节

• 批准号: 8681463
• 负责人: Thomas J. Silhavy
• 金额: $ 30.93万
• 依托单位: PRINCETON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-02-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8681463
• 关键词: 3' Untranslated Regions; Bacteria; Bacteria sigma factor KatF protein; Bacterial Infections; Biochemical; Bioinformatics; Carbon; Cell Survival; Cells; DNA-Directed RNA Polymerase; Data; Ensure; Escherichia coli; Face; Funding; Genes; Genetic; Genetic Transcription; Genotype; Holoenzymes; Mass Spectrum Analysis; Messenger RNA; Mutation; Nitrogen; Noise; Nutrient; Organism; Orphan; Pattern; Peptide Hydrolases; Phase; Phosphorylation; Phosphotransferases; Physiological; Physiology; Polyadenylation; Polyadenylation Pathway; Polynucleotide Adenylyltransferase; Population; Production; Prophages; Protein Dephosphorylation; Proteins; Proteobacteria; Proteolysis; Regulation; Reporting; Sigma Factor; Signal Transduction; Source; Starvation; Stress; Testing; Update; biological adaptation to stress; combat; degradosome; genetic analysis; genetic regulatory protein; insight; mRNA Stability; novel; research study; response; sensor; small molecule; transcription factor

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.

描述（由申请人提供）：替代西格玛因子 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剂。在快速生长的细胞中，会产生 RpoS，但会被 ClpX/P 蛋白酶降解。当缺乏某种特定营养素时，RpoS 合成会增加，降解会停止，活性会增加，或者会发生这些效应的某种组合。两种新型调节蛋白是本提案的重点：SprE (RssB) 和 Crl。 SprE 是一种孤儿反应调节因子，在生长细胞中充当接头，将 RpoS 引导至 ClpX/P 蛋白酶。当碳源耗尽时，这种降解就会停止。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，饥饿是通过 ATP 水平的下降来感知的，而生化数据表明，这种下降是由 ClpX 和 RpoS 本身感知到的。遗传分析进一步揭示了 SprE 在多腺苷酸化和 mRNA 稳定性控制中的第二个功能，并且表明 SprE 的磷酸化对于这种活性很重要。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。使用质谱分析，我们发现 SprE 控制 Poly(A) 聚合酶和 Hfq 与 mRNA 降解体的关联，来自微阵列的数据表明 SprE 具有沉默外源基因的功能。我们将探讨这些新活动的生理意义。我们已经证明，Crl 促进 RpoS 与核心 RNA 聚合酶的结合，并且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 转录增加了 25 倍。然而，Crl 水平没有显着的相应变化。我们证明转录的这种变化允许从嘈杂的 Crl 产生转变为更均匀的 Crl 产生，我们将探讨这两种条件下这些表达模式的生理意义。营养饥饿是细菌面临的最常见的压力，而 RpoS 可以说是大肠杆菌中最重要的全局调节蛋白。更好地了解静止期生理学可能有助于了解如何对抗细菌感染。

项目成果

RpoS proteolysis is regulated by a mechanism that does not require the SprE (RssB) response regulator phosphorylation site.

RpoS 蛋白水解通过不需要 SprE (RssB) 反应调节器磷酸化位点的机制进行调节。

• DOI: 10.1128/jb.186.21.7403-7410.2004
• 发表时间: 2004
• 期刊: Journal of bacteriology.
• 作者: Peterson,CelesteN;Ruiz,Natividad;Silhavy,ThomasJ
• 通讯作者: Silhavy,ThomasJ

Integrating Lys-N proteolysis and N-terminal guanidination for improved fragmentation and relative quantification of singly-charged ions.

• DOI: 10.1016/j.jasms.2010.02.004
• 发表时间: 2010-06
• 期刊: Journal of the American Society for Mass Spectrometry
• 影响因子: 3.2
• 作者: Carabetta VJ;Li T;Shakya A;Greco TM;Cristea IM
• 通讯作者: Cristea IM

Transcriptional occlusion caused by overlapping promoters.

由重叠启动子引起的转录封闭。

• DOI: 10.1073/pnas.1323413111
• 发表时间: 2014
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1
• 作者: Zafar,MAmmar;Carabetta,ValerieJ;Mandel,MarkJ;Silhavy,ThomasJ
• 通讯作者: Silhavy,ThomasJ