MtrA and Mycobacterium tuberculosis proliferation

MtrA 和结核分枝杆菌增殖

• 批准号: 7230008
• 负责人: MALINI RAJAGOPALAN
• 金额: $ 16.69万
• 依托单位: UNIVERSITY OF TEXAS HLTH CTR AT TYLER
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7230008
• 关键词: Affect; Amidohydrolases; Animals; Attenuated; Binding; Biochemical; Biochemical Genetics; Biological; Biological Models; Calmette-Guerin Bacillus; Cell Cycle; Cell Cycle Progression; Cell division; Cells; Chromosomes; Condition; DNA; DNA biosynthesis; Defect; Genes; Genome; Goals; Grant; Growth; HIV; Immune; In Vitro; Infection; Infectious Agent; Lead; Lung; Mediating; Membrane; Modeling; Molecular Genetics; Morbidity - disease rate; Mus; Mycobacterium tuberculosis; Phagocytosis; Phagosomes; Phenotype; Phosphorylation; Phosphotransferases; Play; Process; Proliferating; Protein Binding; Protein Overexpression; Proteins; Recombinants; Regulation; Relative (related person); Replication Initiation; Replication Origin; Replication-Associated Process; Rest; Role; Role playing therapy; Signal Transduction; Site; Surface Plasmon Resonance; System; Techniques; Testing; Tuberculosis; Vaccines; Virulence; amidase; attenuation; base; environmental change; genetic element; improved; in vivo; macrophage; mortality; pathogen; promoter; research study; response; secondary infection; sensor; sensor histidine kinase; success

DESCRIPTION (provided by applicant): Infections due to Mycobacterium tuberculosis (Mtb) are the leading causes of morbidity and mortality in both HIV infected and immune competent people. Following initial infections Mtb frequently enters a latent or dormant state for extended periods and subsequently, under appropriate conditions or following immune suppression, revives, multiplies and causes a secondary infection. The genetic elements responsible for regulation of Mtb replication and multiplication or the factors that promote latent state in vivo are largely unknown. The two component response regulatory signal transduction systems (2CR) consist of a membrane bound sensor kinase and an intracellular regulator, and may play critical roles in processes responsible for Mtb survival and persistence in vivo. The MtrA-MtrB system is an essential 2CR of Mtb, but the roles, if any, of this system in regulating Mtb multiplication in vivo are unknown. Replication of chromosomal DNA and subsequent cell division are essential for multiplication, and interactions of DnaA, the initiator protein with origin of replication or oriC, result in replication initiation. The genetic elements that regulate initiation of chromosomal DNA replication are unknown. Our proposal focuses on connecting the MtrA-MtrB 2CR system to oriC replication and centers on a hypothesis that MtrA has a role in DNA replication, and that both oriC and DnaA are MtrA targets in vivo. This proposal aims to test the hypothesis that the phosphorylation state of MtrA is affected by growth of Mtb in vivo and this in turn affects the replication and subsequent proliferation of Mtb. Using genetic and molecular biological approaches we will characterize the recombinant strains of Mtb expressing elevated levels of wild type and phosphorylation defective MtrA protein and will evaluate the phosphorylation state of MtrA in Mtb isolated from phagosomes. Furthermore, we will test the hypothesis that Mtb strains expressing elevated levels of MtrA are attenuated for growth in vivo in murine infection model system. Biochemical interactions of recombinant wild type and phosphorylated MtrA with oriC will be examined by surface plasmon resonance technique. These experiments will establish whether oriC replication is a target of MtrA system and will improve our understanding of regulation of Mtb multiplication in vivo.

描述（由申请方提供）：结核分枝杆菌（Mtb）感染是HIV感染者和免疫功能正常者发病和死亡的主要原因。初次感染后，结核分枝杆菌经常进入潜伏或休眠状态，持续较长时间，随后在适当条件下或免疫抑制后，恢复、繁殖并引起继发感染。负责调节结核分枝杆菌复制和增殖的遗传元件或促进体内潜伏状态的因子在很大程度上是未知的。双组分反应调节信号转导系统（2CR）由膜结合传感器激酶和细胞内调节器组成，并且可能在负责Mtb在体内存活和持久性的过程中发挥关键作用。MtrA-MtrB系统是结核分枝杆菌的重要2CR系统，但该系统在体内调节结核分枝杆菌增殖中的作用（如果有的话）尚不清楚。染色体DNA的复制和随后的细胞分裂对于增殖是必不可少的，并且DnaA（起始蛋白）与复制起点或oriC的相互作用导致复制起始。调节染色体DNA复制起始的遗传元件是未知的。我们的建议侧重于连接的MtrA-MtrB 2CR系统oriC复制和中心的假设，MtrA在DNA复制中的作用，和oriC和DnaA的MtrA目标在体内。该提议旨在检验MtrA的磷酸化状态受体内Mtb生长的影响并且这进而影响Mtb的复制和随后的增殖的假设。使用遗传和分子生物学方法，我们将表征表达水平升高的野生型和磷酸化缺陷型MtrA蛋白的Mtb的重组菌株，并将评估从吞噬体分离的Mtb中MtrA的磷酸化状态。此外，我们将检验表达升高水平的MtrA的Mtb菌株在鼠感染模型系统中的体内生长被减毒的假设。将通过表面等离子体共振技术检测重组野生型和磷酸化MtrA与oriC的生化相互作用。这些实验将确定oriC复制是否是MtrA系统的靶点，并将提高我们对体内Mtb增殖调控的理解。