Transcriptional control by the global regulator Spx

全球监管机构 Spx 的转录控制

基本信息

批准号：
8528615
负责人：
PETER ZUBER
金额：
$ 39.65万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1992
资助国家：
美国
起止时间：
1992-02-01 至 2015-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8528615
关键词：
ATP-Dependent Proteases Adaptor Signaling Protein Affect Affinity Ala-Trp-Arg-His-Pro-Gln-Phe-Gly-Gly Bacillus anthracis Bacillus subtilis Bacteria Base Sequence Binding Binding Proteins Binding Sites Biological Assay C-terminal Cells Chimeric Proteins Co-Immunoprecipitations Code Communicable Diseases Complex Cysteine DNA DNA Binding DNA Sequence DNA Shuffling DNA-Directed RNA Polymerase DNA-protein crosslink DNase-I Footprinting Defect Disease Disulfides Doctor of Medicine Drug Design Electrophoretic Mobility Shift Assay Elements Energy Transfer Enzymes Exhibits Family FeBABE Fluorescence Fluorescence Anisotropy Gel Genes Genetic Transcription Goals Gram-Positive Bacteria Holoenzymes Immune Immunoprecipitation In Vitro Infection Label LacZ Genes Learning Listeria monocytogenes Mass Spectrum Analysis Measures Mediating Methods Modeling Molecular Conformation Mutation N-terminal Nature Orthologous Gene Oxidants Oxidation-Reduction Oxidative Stress Peptide Hydrolases Peptides Positioning Attribute Process Promoter Regions Protein Family Proteins Proteolysis Reaction Recovery Reducing Agents Regulation Reproduction spores Research Resistance Resolution Response Elements Specific qualifier value Staphylococcus aureus Streptococcus pneumoniae Stress Structure Sulfhydryl Compounds Surface System Technology Testing Transcription Initiation Transcriptional Activation Transcriptional Regulation Ursidae Family Virulence arsenate reductase cell envelope cis acting element combat gene function genetic regulatory protein in vitro Assay in vivo member microorganism mutant oxidation pathogen pathogenic bacteria prevent promoter protein protein interaction public health relevance research study response screening stoichiometry thioredoxin reductase transcription factor zinc-binding protein

项目摘要

DESCRIPTION (provided by applicant): In pathogenic bacteria, complex systems of control have evolved that activate defenses against the various toxic agents used by immune cells to combat infection. These systems regulate important virulence determinants, which are targets of drugs designed to alleviate infectious disease. The low GC Gram-positive bacteria have a unique system of transcriptional control, mediated by the protein Spx, that functions in the response to toxic oxidants, including those produced by phagocytic immune cells. Spx is a transcriptional regulator that we discovered in the spore-forming bacterium, Bacillus subtilis, but is also found in Staphylococcus aureus, Streptococcus pneumoniae, Listeria monocytogenes, and Bacillus anthracis. Spx has been shown to control the expression of virulence-associated activities in S. aureus, and is produced to high levels in Listeria monocytogenes and B. anthracis during host cell invasion. Spx defines a family of regulators that structurally resemble the enzyme arsenate reductase. It bears an N-terminal CXXC redox disulfide center that regulates its activity. Its mechanism of control is unique in that free Spx does not exhibit DNA-binding activity, but when bound to RNA polymerase (RNAP) in its oxidized disulfide form, it directs the enzyme to transcribe specific genes that function in oxidative stress defense. Spx concentration is reduced to low levels after recovery from stress by the protease ClpXP and the Spx-binding protein YjbH, which accelerates Spx proteolysis. The goal of the proposed research is to learn how B. subtilis Spx activates transcription initiation and how YjbH functions to mediate Spx proteolysis. Specific nucleotide sequence elements have been identified in the promoter regions of genes controlled by Spx. These sequences bind a complex of Spx and the C-terminal domain of RNAP subunit (CTD). The promoter DNA sequence and the structural requirements of CTD/Spx for promoter DNA binding will be determined. Furthermore, the dynamics of subunit positioning in RNAP holoenzyme within the Spx-activated transcription complex will be explored. Efficient proteolysis requires YjbH, a zinc-binding protein that interacts with Spx and accelerates ClpXP-catalyzed Spx proteolysis. Studies are proposed to investigate the oxidant-induced stabilization of Spx and redox control of YjbH activity. This study will focus on the Zn-binding site of the adaptor to test the model that oxidation of Zn-coordinating cysteine residues causes inactivation of YjbH and release of Spx. Differential thiol labeling and mass spectrometry will be performed to investigate the extent and nature of thiol oxidation of Spx and YjbH in vivo. A small protein factor, YirB that interacts with YjbH, will be studied to determine if it is a member of a growing list of small proteins that prevent protease-catalyzed degradation by inhibiting protease adaptor proteins. PUBLIC HEALTH RELEVANCE: Complex systems of control have evolved in disease-causing bacteria to mobilize defenses that counter the toxic oxidizing agents produced by phagocytic immune cells. The regulatory protein Spx controls important components of the bacterial response to oxidative attack, and the study of its mechanism of action will uncover targets for neutralizing infectious microorganisms.

描述（由申请人提供）：在病原细菌中，复杂的控制系统已经进化，可以激活对免疫细胞用来对抗感染的各种有毒物质的防御。这些系统调节重要的毒力决定因素，它们是旨在减轻传染病的药物的目标。低 GC 革兰氏阳性细菌具有独特的转录控制系统，由蛋白质 Spx 介导，该系统在对有毒氧化剂（包括吞噬免疫细胞产生的氧化剂）做出反应时起作用。 Spx 是一种转录调节因子，我们在产芽孢细菌枯草芽孢杆菌中发现，但也在金黄色葡萄球菌、肺炎链球菌、单核细胞增生李斯特菌和炭疽芽孢杆菌中发现。 Spx 已被证明可以控制金黄色葡萄球菌中毒力相关活性的表达，并且在宿主细胞侵袭期间在单核细胞增生李斯特菌和炭疽芽孢杆菌中产生高水平。 Spx 定义了一个结构类似于砷酸还原酶的调节剂家族。它具有调节其活性的 N 末端 CXXC 氧化还原二硫化物中心。其控制机制的独特之处在于，游离的 Spx 不表现出 DNA 结合活性，但当以氧化二硫键形式与 RNA 聚合酶 (RNAP) 结合时，它会指导该酶转录在氧化应激防御中起作用的特定基因。从应激状态恢复后，蛋白酶 ClpXP 和 Spx 结合蛋白 YjbH 将 Spx 浓度降低至较低水平，从而加速 Spx 蛋白水解。本研究的目的是了解枯草芽孢杆菌 Spx 如何激活转录起始以及 YjbH 如何发挥作用来介导 Spx 蛋白水解。已在 Spx 控制的基因的启动子区域中鉴定出特定的核苷酸序列元件。这些序列结合 Spx 和 RNAP 亚基 (CTD) C 端结构域的复合物。将确定启动子 DNA 序列和 CTD/Spx 对启动子 DNA 结合的结构要求。此外，还将探索 Spx 激活转录复合物内 RNAP 全酶亚基定位的动态。有效的蛋白水解需要 YjbH，这是一种锌结合蛋白，可与 Spx 相互作用并加速 ClpXP 催化的 Spx 蛋白水解。建议研究氧化剂诱导的 Spx 稳定性和 YjbH 活性的氧化还原控制。本研究将重点关注接头的 Zn 结合位点，以测试 Zn 配位半胱氨酸残基氧化导致 YjbH 失活和 Spx 释放的模型。将进行差异硫醇标记和质谱分析来研究 Spx 和 YjbH 体内硫醇氧化的程度和性质。将研究与 YjbH 相互作用的小蛋白因子 YirB，以确定它是否是越来越多的小蛋白中的一员，这些小蛋白通过抑制蛋白酶接头蛋白来防止蛋白酶催化的降解。公共健康相关性：致病细菌进化出复杂的控制系统，以动员防御系统对抗吞噬免疫细胞产生的有毒氧化剂。调节蛋白 Spx 控制细菌对氧化攻击反应的重要组成部分，对其作用机制的研究将揭示中和传染性微生物的靶点。

项目成果

期刊论文数量（43）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Plasmid-amplified comS enhances genetic competence and suppresses sinR in Bacillus subtilis.

质粒扩增的 comS 可增强枯草芽孢杆菌的遗传能力并抑制 sinR。

DOI：
发表时间：
1996-09
期刊：
影响因子：
3.2
作者：
Liu, L;Nakano, M M;Lee, O H;Zuber, P
通讯作者：
Zuber, P

Dual control of sbo-alb operon expression by the Spo0 and ResDE systems of signal transduction under anaerobic conditions in Bacillus subtilis.

枯草芽孢杆菌厌氧条件下信号转导的 Spo0 和 ResDE 系统对 sbo-alb 操纵子表达的双重控制。

DOI：
发表时间：
2000-06
期刊：
影响因子：
3.2
作者：
Nakano, M M;Zheng, G;Zuber, P
通讯作者：
Zuber, P

The MarR-type repressor MhqR (YkvE) regulates multiple dioxygenases/glyoxalases and an azoreductase which confer resistance to 2-methylhydroquinone and catechol in Bacillus subtilis.

MarR 型阻遏物 MhqR (YkvE) 调节多种双加氧酶/乙二醛酶和偶氮还原酶，赋予枯草芽孢杆菌对 2-甲基氢醌和儿茶酚的抗性。

DOI：
发表时间：
2007-10
期刊：
影响因子：
3.6
作者：
Töwe, Stefanie;Leelakriangsak, Montira;Kobayashi, Kazuo;Van Duy, Nguyen;Hecker, Michael;Zuber, Peter;Antelmann, Haike
通讯作者：
Antelmann, Haike

Amino-acylation site mutations in amino acid-activating domains of surfactin synthetase: effects on surfactin production and competence development in Bacillus subtilis.

表面活性素合成酶氨基酸激活域的氨基酰化位点突变：对枯草芽孢杆菌中表面活性素产生和能力发展的影响。