Contribution of catabolite control protein A to group A streptococcal virulence

分解代谢物控制蛋白 A 对 A 组链球菌毒力的贡献

• 批准号: 8300803
• 负责人: SAMUEL A SHELBURNE
• 金额: $ 31.6万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-15 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8300803
• 关键词: A-Form DNA; Address; Affect; Affinity; Bacillus (bacterium); Bacteria; Bacterial Genes; Bacterial Infections; Bacterial Physiology; Binding; Binding Sites; Bioinformatics; Carbon; Cessation of life; Consensus Sequence; DNA; DNA Binding; DNA Sequence; Data; Development; Disease; Eating; Environment; Future; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Genes; Goals; Gram-Positive Bacteria; Growth; Histidine; Human; In Vitro; Infection; Investigation; Laboratories; Metabolic; Methods; Modeling; Molecular Profiling; Mus; Muscle; Mutation; Myositis; Nature; Necrotizing fasciitis; Pathogenesis; Pathway interactions; Pharyngeal structure; Pharyngitis; Phosphoproteins; Phosphorylase Kinase; Physiological; Preventive Intervention; Production; Proteins; Recombinants; Research; Response Elements; Role; Serine; Serotyping; Site; Source; Stimulus; Streptococcal Infections; Streptococcus pyogenes; Testing; Transcript; United States; Virulence; Virulence Factors; base; defined contribution; design; genetic regulatory protein; in vivo; innovation; insight; mouse model; multiple myeloma M Protein; mutant; novel; novel therapeutic intervention; pathogen; pathogenic bacteria; response

DESCRIPTION (provided by applicant): Infections by Gram-positive bacteria cause tens of thousands of deaths in the United States each year. The transcriptional regulator catabolite control protein A (CcpA) has recently been found to influence the virulence gene expression profile during growth in standard laboratory medium of numerous Gram-positive pathogens including group A Streptococcus (GAS). CcpA was originally identified in Bacillus species where it binds DNA sequences termed catabolite response element (cre) sites in conjunction with its co-factor histidine-containing phosphoprotein phosphorylated at serine residue 46 (HPr-Ser46-P). Orthologues of CcpA, HPr, and the kinase/phosphorylase that regulates HPr phosphorlyation status (HPrK/P) are present in all major Gram- positive human pathogens, and there are accumulating data suggesting that the CcpA-(HPr-Ser46-P)-HPrK/P axis contributes to the infectivity of diverse bacterial species. The long term goals of this project are to clearly define how the CcpA-(HPr-Ser46-P)-HPrK/P axis contributes to GAS virulence and to ascertain mechanisms by which CcpA influences gene expression during infection. The specific aims of this proposal have been designed to test hypotheses arising from preliminary data demonstrating that: 1) Inactivation of CcpA altered GAS gene expression during infection in mouse muscle; 2) Inactivation of CcpA decreased GAS virulence in a mouse myositis model; 3) Mutation of the central CpG of a GAS cre site decreased CcpA-(HPr-Ser46-P) DNA binding affinity; and 4) CcpA influenced gene expression under conditions in which HPr-Ser46-P levels are low or absent. In aim 1, gene expression analysis of wild-type and CcpA-inactivated GAS strains will be performed in multiple mouse models of infection to test the hypothesis that CcpA is a key virulence factor regulator during GAS infection. In aim 2, the virulence of CcpA isogenic mutant strains derived from different M protein serotype parental strains will be compared to their parental wild-type strains in various mouse models of infection to test the hypothesis that CcpA positively contributes to the infectivity of GAS strains of multiple M protein serotypes. In aim 3, a combination of in vitro and in vivo methods will be used to test the hypothesis that the central cre site CpG is critical for the effect of CcpA on GAS gene expression. In aim 4, the gene expression profiles of CcpA and HPr-Ser46-P inactivated strains will be compared under different metabolic conditions to test the hypothesis that CcpA can affect GAS gene expression in the absence of HPr-Ser46-P. The highly conserved nature of the CcpA-(HPr-Ser46-P)-HPrK/P axis amongst major Gram-positive bacterial pathogens means that the successful completion of the proposed research may generate new insights into disease pathogenesis that are relevant to a broad array of pathogenic bacteria. Such information could form the basis for the development of novel therapeutic or preventive interventions.

描述（由申请人提供）：革兰氏阳性细菌的感染每年在美国导致成千上万的死亡。最近发现，转录调节剂分解代谢物对照蛋白A（CCPA）会影响众多革兰氏阳性病原体的标准实验室培养基在包括A组链球菌（GAS）的标准实验室培养基中的毒力基因表达谱。 CCPA最初是在杆菌物种中鉴定出来的，其中结合了称为分解代谢物反应元件（CRE）位点的DNA序列与其含丝氨酸残基46（HPR-SER46-P）的含有组氨酸的磷酸化磷酸化磷酸化的含磷酸化的磷酸化磷酸蛋白。 CCPA，HPR和调节HPR磷酸化状态（HPRK/P）的激酶/磷酸化酶的直系同源物都存在于所有主要的革兰氏阴性人类病原体中，并且存在积累的数据，表明CCPA-（HPR-SER46-P）-hprk/P轴 - hprk/p axis对多样的细菌造成了多样的细菌。该项目的长期目标是清楚地定义CCPA-（HPR-SER46-P）-HPRK/P轴如何有助于气体毒力和确定CCPA在感染过程中影响基因表达的机制。该提案的具体目的旨在测试由初步数据引起的假设，表明：1）CCPA失活在小鼠肌肉中感染期间的气体基因表达改变； 2）CCPA失活减少了小鼠肌炎模型中的气体毒力； 3）气体CRE位点中央CpG的突变降低了CCPA-（HPR-SER46-P）DNA结合亲和力； 4）CCPA在HPR-SER46-P水平低或不存在的条件下影响基因表达。在AIM 1中，将在多种感染小鼠模型中对野生型和CCPA灭活气体菌株进行基因表达分析，以检验CCPA是气体感染过程中CCPA是关键的毒力因子调节剂的假设。在AIM 2中，将将来自不同M蛋白血清型亲代亲代菌株的CCPA等生突变菌株的毒力与其在各种感染小鼠感染模型中的父母野生型菌株进行比较，以检验CCPA的假设，即CCPA积极地有助于多种M蛋白质血清的气体菌株的感染性。在AIM 3中，将使用体外和体内方法的组合来检验以下假设：CCPA对CCPA对气体基因表达的影响至关重要。在AIM 4中，将在不同的代谢条件下比较CCPA和HPR-SER46-P灭活菌株的基因表达谱，以检验CCPA在没有HPR-SER46-P的情况下会影响气体基因表达的假设。 CCPA-（HPR-SER46-P）-HPRK/P轴的高度保守性质在主要的革兰氏阳性细菌病原体中意味着，所提出的研究的成功完成可能会引起对疾病发病机理的新见解，这与广泛的致病性细菌有关。这些信息可能构成开发新的治疗或预防性干预措施的基础。