CAREER: Effects of Reductive Evolution on Amino Acid Stress Responses in Bacteria

职业：还原进化对细菌氨基酸应激反应的影响

• 批准号: 1810599
• 负责人: Scot Ouellette
• 金额: $ 101.99万
• 依托单位: University of Nebraska Medical Center
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-10-26 至 2023-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1810599&HistoricalAwards=false
• 关键词: CAREER; Effects; Reductive; Evolution; Amino

Most bacteria cope with starvation by launching a global change in gene expression known as the stringent response. The stringent response causes growth-related genes to be turned off and genes that are needed for survival under nutrient-limiting conditions to be activated. This response is absent in obligate intracellular human pathogens such as Chlamydia, which only grow inside a host cell. The aim of this project to understand how these bacterial pathogens cope with starvation, since they lack the stringent response and yet they are often able to survive amino acid starvation, which is induced by the human host to combat the infection. This project will address fundamental questions about the evolution of pathogens and the unique manner in which they respond to starvation. More practically, it will result in the identification of potential drug and diagnostic targets to treat and identify diseases caused by pathogens such as Chlamydia. In an effort to broaden participation, several strategies will be used to increase participation of American Indians in college-level research. Some of these students will present their research to their alma mater high schools to help encourage the students to pursue college degrees in STEM. In addition, various science-themed activities will be used to increase interest/awareness/participation of American Indian high school and college students in STEM. This study aims to understand the adaptations and their consequences in evolving to obligate intracellular growth. The obligate intracellular bacterium Chlamydia has minimized its genome by eliminating the stringent response and amino acid biosynthesis pathways. The stringent response is activated when an uncharged tRNA binds in the A site of the ribosome, leading to (p)ppGpp synthesis, a global regulator that, in part, regenerates amino acid pools. Chlamydia does not make tryptophan (trp) or (p)ppGpp thus how it survives trp starvation is not well understood. Prior work by the PI has shown that Chlamydia globally increases its transcription in response to trp starvation while globally decreasing its translation. This observation is unusual for a bacterium, and therefore, represents a unique opportunity to advance knowledge. How the amino acid content of a transcript affects gene expression during starvation has not been fully characterized. The hypothesis is that, during trp limitation, chlamydial gene expression is determined by the trp content of the encoded protein. The goals of this project are the following: 1) to determine if the trp content of open reading frames controls transcription during trp starvation; 2) to determine if trp-free proteins are preferentially translated during trp starvation; 3) to compare and contrast the chlamydial response to trp starvation (1&2) with that of other trp auxotrophs, Streptococcus pyogenes (wild-type and stringent mutant) and Coxiella. The major goal of the broader impacts of this project is to increase participation of American Indians in STEM research. In conjunction with the Native Studies Program, the PI also plans to develop a course on science in the context of the American Indian worldview.

大多数细菌通过引发基因表达的全球变化来应对饥饿，这种变化被称为“严格反应”。这种严格的反应导致生长相关基因被关闭，而在营养限制条件下生存所需的基因被激活。这种反应在专性细胞内的人类病原体中是不存在的，比如衣原体，它只在宿主细胞内生长。该项目的目的是了解这些细菌病原体如何应对饥饿，因为它们缺乏严格的反应，但它们通常能够在人类宿主诱导的氨基酸饥饿中存活下来，以对抗感染。该项目将解决有关病原体进化的基本问题以及它们对饥饿作出反应的独特方式。更实际的是，它将导致识别潜在的药物和诊断靶标，以治疗和识别由衣原体等病原体引起的疾病。为了扩大参与，将采用若干策略来增加美国印第安人对大学水平研究的参与。其中一些学生将向母校高中展示他们的研究成果，以鼓励学生们攻读STEM专业的大学学位。此外，还将举办各种以科学为主题的活动，以提高美国印第安高中生和大学生对STEM的兴趣/意识/参与。本研究旨在了解细胞内生长的适应性及其影响。专性细胞内细菌衣原体通过消除严格的反应和氨基酸生物合成途径使其基因组最小化。当一个未带电的tRNA结合在核糖体的A位点上，导致(p)ppGpp合成时，严格的反应被激活，ppGpp是一种全局调节剂，在一定程度上再生氨基酸库。衣原体不产生色氨酸（trp）或(p)ppGpp，因此它如何在缺乏色氨酸的情况下存活尚不清楚。PI先前的工作表明，衣原体在全球范围内增加其转录以响应trp饥饿，同时在全球范围内减少其翻译。这种观察结果对细菌来说是不寻常的，因此，它代表了一个独特的机会来推进知识。转录物的氨基酸含量如何影响饥饿期间的基因表达尚未得到充分表征。假设是，在色氨酸限制期间，衣原体基因的表达由编码蛋白的色氨酸含量决定。该项目的目标如下：1)确定开放阅读框的trp内容是否在trp缺乏期间控制转录；2)确定不含TRP的蛋白在TRP饥饿时是否优先翻译；3)比较和对比衣原体对trp饥饿（1&2）的反应与其他trp营养不良菌、化脓性链球菌（野生型和严格突变型）和科希氏菌的反应。该项目更广泛影响的主要目标是增加美国印第安人对STEM研究的参与。PI还计划与土著研究项目一起，开设一门关于美国印第安人世界观背景下的科学课程。

项目成果

The iron-dependent repressor YtgR is a tryptophan-dependent attenuator of the trpRBA operon in Chlamydia trachomatis.

• DOI: 10.1038/s41467-020-20181-5
• 发表时间: 2020-12-22
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Pokorzynski ND;Hatch ND;Ouellette SP;Carabeo RA
• 通讯作者: Carabeo RA

The ClpX and ClpP2 Orthologs of Chlamydia trachomatis Perform Discrete and Essential Functions in Organism Growth and Development

• DOI: 10.1128/mbio.02016-20
• 发表时间: 2020-09-01
• 期刊: MBIO
• 影响因子: 6.4
• 作者: Wood, Nicholas A.;Blocker, Amanda M.;Ouellette, Scot P.
• 通讯作者: Ouellette, Scot P.

Initial Characterization of the Two ClpP Paralogs of Chlamydia trachomatis Suggests Unique Functionality for Each

• DOI: 10.1128/jb.00635-18
• 发表时间: 2019-01-01
• 期刊: JOURNAL OF BACTERIOLOGY
• 影响因子: 3.2
• 作者: Wood, Nicholas A.;Chung, Krystal Y.;Ouellette, Scot P.
• 通讯作者: Ouellette, Scot P.

Inhibition of tRNA Synthetases Induces Persistence in Chlamydia

• DOI: 10.1128/iai.00943-19
• 发表时间: 2020-04-01
• 期刊: INFECTION AND IMMUNITY
• 影响因子: 3.1
• 作者: Hatch, Nathan D.;Ouellette, Scot P.
• 通讯作者: Ouellette, Scot P.

Characterization of Chlamydial Rho and the Role of Rho-Mediated Transcriptional Polarity during Interferon Gamma-Mediated Tryptophan Limitation

衣原体 Rho 的表征以及 Rho 介导的转录极性在干扰素 Gamma 介导的色氨酸限制过程中的作用

• DOI: 10.1128/iai.00240-18
• 发表时间: 2018
• 期刊: Infection and Immunity
• 影响因子: 3.1
• 作者: Ouellette, Scot P.;Messerli, Parker R.;Wood, Nicholas A.;Hajovsky, Heather;Roy, Craig R.
• 通讯作者: Roy, Craig R.