Clostridium difficile Toxin Gene Regulation

艰难梭菌毒素基因调控

• 批准号: 7154110
• 负责人: ABRAHAM Lincoln SONENSHEIN
• 金额: $ 27.05万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-12-15 至 2009-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7154110
• 关键词: Affect; Affinity Chromatography; Amino Acids; Antibiotic-Associated Colitis; Antibiotics; Bacillus subtilis; Binding; Biochemistry; Biological Assay; Biotin; Carbon; Cells; Clostridium difficile; Clostridium perfringens; Co-Immunoprecipitations; Collection; Complex; Condition; DNA Microarray Chip; DNA Microarray format; DNA Sequence; DNA-Directed RNA Polymerase; Disease; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Genomics; Glucose; Growth; Homologous Gene; In Vitro; Knock-out; Laboratories; Measures; Mediating; Mediator of activation protein; Microarray Analysis; Molecular; Molecular Genetics; Mutation; Pathogenicity; Pathogenicity Islets; Phase; Physiology; Point Mutation; Promoter Regions; Protein Overexpression; Proteins; Pseudomembranous Colitis; Regulation; Repression; Research; Research Personnel; Role; Scheme; Sigma Factor; Signal Transduction; Site; Source; Standards of Weights and Measures; System; Temperature; Testing; Time; Toxin; Virulence; Virulence Factors; Vitamins; Work; base; comparative; crosslink; genetic analysis; genetic regulatory protein; in vivo; mutant; novel; null mutation; nutrition; prevent; programs; promoter; protein protein interaction; research study; response; tool

DESCRIPTION (provided by applicant): Clostridium difficile is the principal causative agent of antibiotic-associated colitis and the only known cause of pseudomembranous colitis, a potentially lethal disease. Two large toxin proteins, encoded by the toxA and toxB genes, which lie within a 19 kb pathogenicity islet, are the primary virulence factors. Previous work from the applicants' laboratories has shown that transcription of the tox genes depends entirely on TxeR, a novel RNA polymerase sigma factor encoded within the same pathogenicity locus. The level of expression is strongly modulated, however, by the growth state of the cells and by environmental conditions. For instance, for cells growing in broth medium, tox gene transcription is restricted to stationary phase cells and is repressed by rapidly metabolizable carbon sources, such as glucose. Other factors that influence toxin synthesis are the availability of biotin and certain amino acids and the temperature of cultivation. The present proposal seeks to determine the molecular mechanisms that control toxin synthesis in response to environmental signals. Candidate regulatory proteins, based on comparative genomics, will be specifically tested for their participation in such regulation. These proteins include CodY, CcpA and VirR, whose homologs in Bacillus subtilis or Clostridium perfringens have been shown to mediate the types of regulatory effects in question. In addition, general, unbiased searches for the relevant regulatory proteins will be carried out based on affinity chromatography. A fourth protein, TcdC, will be tested for its potential activity as an antagonist of TxeR. The project makes use of the expertise in C. difficile biochemistry, genetics and physiology of the three collaborating research groups and recent advances made by each of the groups in making this experimental system amenable to detailed molecular genetic analysis.

描述（由申请人提供）：艰难梭菌是抗生素相关性结肠炎的主要病因，也是假膜结肠炎的唯一已知原因，伪膜性结肠炎是一种潜在的致命性疾病。主要的毒力因素是两个大的毒素蛋白，该蛋白质由毒素和毒素基因编码，分别为19 kb的致病性胰岛，是主要的毒力因素。申请人实验室的先前工作表明，托克斯基因的转录完全取决于TXER，这是一种新型的RNA聚合酶Sigma因子，它在相同的致病基因座中编码。但是，通过细胞的生长状态和环境条件，表达水平得到了强烈的调节。例如，对于在肉汤培养基中生长的细胞，托克斯基因转录仅限于固定相细胞，并被迅速代谢的碳源（例如葡萄糖）抑制。影响毒素合成的其他因素是生物素和某些氨基酸的可用性以及培养温度。本提案旨在确定根据环境信号控制毒素合成的分子机制。基于比较基因组学的候选调节蛋白将被专门测试其参与这种调节。这些蛋白质包括Cody，CCPA和Virr，其在枯草芽孢杆菌或灌注梭状芽胞杆菌中的同源物已显示可介导所讨论的调节作用的类型。此外，将根据亲和力色谱法进行一般，公正的对相关调节蛋白的搜索。第四个蛋白TCDC将以TXER的拮抗剂的潜在活性进行测试。该项目利用了三个合作研究小组的艰难梭菌生物化学，遗传学和生理学的专业知识，以及每个小组在使这个实验性系统可容纳详细的分子遗传分析方面取得的最新进展。