Functional analysis of Mycobacterium tuberculosis VapC toxins

结核分枝杆菌VapC毒素的功能分析

• 批准号: 9108982
• 负责人: Melvili Cintron
• 金额: $ 3.28万
• 依托单位: RBHS-ROBERT WOOD JOHNSON MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9108982
• 关键词: Address; Amino Acid Sequence; Antibiotics; Anticodon; Antitoxins; Bacteria; Bacterial Toxins; Biochemical; Biochemical Genetics; Cells; Characteristics; Cleaved cell; Complex; Consensus Sequence; Coupled; Data; Employee Strikes; Environment; Enzymes; Escherichia coli; Family; Family member; Fellowship; Genome; Growth; Health; Human; Immune response; Infection; Ions; Laboratories; Learning; Literature; Messenger RNA; Methods; Molecular; Mono-S; Mycobacterium tuberculosis; N-terminal; Organism; Peptide Sequence Determination; Phenotype; Physiological; Physiology; Process; Property; Protein Family; RNA; RNA Sequences; Refractory; Regulation; Ribonucleases; Ribosomal RNA; Role; Serine; Site; Specificity; Structure; System; Techniques; Testing; Therapeutic; Threonine; Time; Toxin; Training; Transfer RNA; Translations; Tuberculosis; Virulence; Work; antimicrobial; assault; base; cell growth; cellular targeting; genetic approach; genome-wide; inorganic phosphate; latent infection; member; novel; research study; stem; transcriptome; transcriptome sequencing

DESCRIPTION (provided by applicant): Mycobacterium tuberculosis (Mtb) has adapted to survive a wide range of assaults-from our immune response to antimicrobial therapeutics-intended to eradicate the organism. This unique ability to persist for long periods of time in its human host as a latent tuberculosis infection dominates the infection landscape relative to active tuberculosis cases. Despite ongoing efforts to understand latent tuberculosis infection, the molecular switches that enable Mtb to slow or stop replication and become dormant are poorly understood. However, the convergence of studies on bacterial toxin-antitoxin (TA) systems/modules demonstrating their general growth-regulating properties, coupled with the identification of a striking abundance of TA systems in the Mtb genome, suggest that their expression may facilitate the non-replicating persistent state of Mtb during latent tuberculosis infection. This proposal seeks to fill the gap in our understanding of the molecular basis of laten tuberculosis infection by investigating the structure and function of the large, uncharacterized family of Mtb VapC toxins through detailed study of three family members. The studies in this fellowship proposal integrate into the broader focus of the Woychik laboratory and address our hypothesis that the VapC toxin family in Mtb represents a new class of enzymes that recognize both RNA sequence and tertiary structure to specifically cleave tRNA and/or rRNA(s) and inactivate the translation machinery. The studies proposed in Aim 1 enlist a specialized genome scale method, 5' RNA-seq, to identify the RNA targets for three Mtb VapC toxins using an Escherichia coli host, those in Aim 2 validate the RNA targets and cleavage sites identified by 5' RNA-seq.

描述（由申请人提供）：结核分枝杆菌（MTB）已经适应了我们对抗菌治疗药的免疫反应，以消除生物体。作为潜在的结核病感染，这种独特的能力长期存在于其人类宿主中，相对于活跃的结核病病例，该局势主导了感染景观。尽管持续不断地了解潜在的结核病感染，但使MTB能够减慢或停止复制和休眠的分子开关知之甚少。然而，对细菌毒素 - 抗毒素（TA）系统/模块的研究的收敛性，证明了它们的一般生长调节特性，并鉴定出MTB基因组中TA系统的惊人丰度，这表明它们的表达可能促进非复制的MTB持续性MTB持续性持续性。该提议试图通过研究三个家庭成员的详细研究，通过研究大型，未表征的MTB VAPC毒素家族的结构和功能来填补我们对拉链结核感染分子基础的差距。该奖学金提案中的研究融入了Woychik实验室的更广泛的重点，并解决了我们的假设，即MTB中的VAPC毒素家族代表了一种新的酶，这些酶识别RNA序列和第三级结构，以特异性地切除TRNA和/或RRNA和/或RRNA（s），并使得翻译机器。 AIM 1中提出的研究邀请了一种专业的基因组量表方法5'RNA-Seq，以使用Escherichia Coli宿主识别三种MTB VAPC毒素的RNA靶标，AIM 2中的研究验证了RNA靶标和切割位点，该靶标由5'RNA-RNA-rna-seq鉴定出RNA靶标。