Physiological consequences of CodY: a master regulator in gram-positive bacteria.

CodY 的生理后果：革兰氏阳性菌的主要调节因子。

基本信息

批准号：
7540673
负责人：
Shaun R Brinsmade
金额：
$ 4.48万
依托单位：
TUFTS UNIVERSITY BOSTON
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-08-01 至 2011-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7540673
关键词：
Address Alleles Amino Acid Substitution Amino Acids Anabolism Antibiotics Bacillus subtilis Bacteria Bacterial Physiology Binding Binding Sites Biochemistry Biological Biological Assay Branched-Chain Amino Acids Carbon Cells Collaborations Companions Complement Complex Condition DNA Binding DNA Binding Domain DNase-I Footprinting Data Development Disease Energy-Generating Resources Famines Gel Gene Expression Genes Genetic Genetic Transcription Genome Glucose Glutamine Goals Gram-Positive Bacteria Growth Guanosine Triphosphate Homeostasis Housing Human In Vitro Isoleucine Laboratories LacZ Genes Lead Leucine Logic Metabolic Metabolism Methods Molecular Molecular Biology Molecular Profiling Monitor Mutation Nitrogen Numbers Nutrient Nutritional status Operon Pathway interactions Phase Physiological Process Production Proteins Proteolysis Regulation Regulon Reporter Repression Research Resources Reverse Transcriptase Polymerase Chain Reaction Signal Transduction Source Structure System Techniques Testing Universities Valine Variant Virulence Virulence Factors Work X-Ray Crystallography analog antimicrobial antitermination cytotoxic gel mobility shift assay genetic regulatory protein in vivo interdisciplinary approach member microbial mimicry pathogen post-doctoral training prevent professor programs research study response sensor structural biology three dimensional structure transcription factor transcription termination ward

项目摘要

DESCRIPTION (provided by applicant): Bacteria in the wild seldom find themselves in conditions that promote efficient and robust growth. Short periods of nutrient surplus give rise to even longer periods of nutrient limitation. When deprived of nutrients or otherwise favorable conditions, many bacteria attempt to adapt by deploying strategies to acquire alternative carbon and energy sources either by activating secondary catabolic pathways and transporters or by unleashing cytotoxic compounds, including antimicrobials, to ward off competition and scavenge for resources. Many of these adaptive responses are coincidently harmful to humans. The CodY protein is the master nutrient sensor and transcriptional regulator responsible for controlling the expression of this adaptive program in low G+C gram-positive bacteria, including a number of recalcitrant human pathogens. This application aims to determine how CodY processes multiple input signals for controlling transcription on a global level to alter important aspects of central metabolism. In addition, research proposed in this application will define the physiological importance of altering CodY activity under different growth conditions, and will identify additional coeffectors of CodY. The broad goal is to elucidate the molecular mechanism by which CodY functions to activate and repress transcription at numerous loci. The projects described herein take a multidisciplinary approach using genetics, biochemistry, molecular biology and structural biology to address biological questions from a variety of directions. Reporter fusions, quantitative RT-PCR and microarrays will provide local and global expression data in vivo. In vitro techniques including gel mobility shift assays, DNase I footprinting, partial proteolysis, and in vitro transcription will confirm and complement in vivo data. Collaborations with experts in X-ray crystallography (Professor Anthony Wilkinson - University of York) and bacterial physiology (Uwe Sauer - ETH, Zurich) have been established to increase the breadth of research and discovery. Dissecting this complex regulatory network and studying the physiological consequences of disrupting genetic and metabolic circuitry underlying CodY activity will reveal how bacteria decide to activate developmental and adaptive programs. Recent work has documented genes encoding virulence determinants as direct targets of CodY-dependent repression. As such, methods and bioactive compounds that can increase the capacity of CodY to maintain repression of virulence determinants can lead to new ways to control and prevent illnesses of microbial origin.

描述（由申请人提供）：野生细菌很少发现自己处于促进有效和强劲生长的条件下。短期的营养过剩会导致更长时间的营养限制。当缺乏营养或其他有利条件时，许多细菌试图通过部署策略来获取替代碳和能源来适应，或者通过激活次级分解代谢途径和转运蛋白，或者通过释放细胞毒性化合物（包括抗菌剂）来避免竞争和资源掠夺。许多适应性反应同时对人类有害。 CodY 蛋白是主要的营养传感器和转录调节因子，负责控制低 G+C 革兰氏阳性细菌（包括许多顽固的人类病原体）中这种适应性程序的表达。该应用旨在确定 CodY 如何处理多个输入信号以在全局水平上控制转录，从而改变中枢代谢的重要方面。此外，本申请中提出的研究将定义在不同生长条件下改变 CodY 活性的生理重要性，并将鉴定 CodY 的其他协同效应子。总体目标是阐明 CodY 在多个位点激活和抑制转录的分子机制。本文描述的项目采用多学科方法，利用遗传学、生物化学、分子生物学和结构生物学从各个方向解决生物学问题。报告基因融合、定量 RT-PCR 和微阵列将提供体内局部和整体表达数据。包括凝胶迁移率变化测定、DNase I 足迹、部分蛋白水解和体外转录在内的体外技术将证实和补充体内数据。与 X 射线晶体学专家（约克大学 Anthony Wilkinson 教授）和细菌生理学专家（苏黎世联邦理工学院 Uwe Sauer）建立了合作，以扩大研究和发现的广度。剖析这个复杂的调控网络并研究破坏 CodY 活性的遗传和代谢回路的生理后果将揭示细菌如何决定激活发育和适应性程序。最近的工作记录了编码毒力决定子的基因作为 CodY 依赖性抑制的直接目标。因此，可以增加 CodY 维持毒力决定簇抑制能力的方法和生物活性化合物可以带来控制和预防微生物源性疾病的新方法。