The roles of glutathione metabolism in growth and virulence of Listeria monocytogenes

谷胱甘肽代谢在单核细胞增生李斯特菌生长和毒力中的作用

• 批准号: 10526637
• 负责人: BORIS R BELITSKY
• 金额: $ 20.63万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-25 至 2024-06-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10526637
• 关键词: Address; Affect; Amino Acids; Animals; Bacteria; Biochemical; Cells; Cysteine; Cytosol; Degradation Pathway; Dependence; Disease; Drug Metabolic Detoxication; Elderly; Environment; Enzymes; Essential Amino Acids; Eukaryotic Cell; Gene Expression; Gene Expression Profile; Gene Silencing; Genes; Genetic; Genetic Screening; Genetic Transcription; Genome; Glutathione; Glutathione Metabolism Pathway; Gram-Positive Bacteria; Growth; Human Development; Immunocompromised Host; In Vitro; Individual; Infection; Knowledge; Listeria monocytogenes; Maintenance; Metabolic; Metabolic Pathway; Metabolism; Modeling; Mus; Nature; Newborn Infant; Oral; Organism; Oxidation-Reduction; Pathogenesis; Pathogenicity; Pathway interactions; Peptides; Physiological; Pregnant Women; Proteins; Regulation; Research; Role; Signal Transduction; Source; Testing; Therapeutic Intervention; Toxic effect; United States; Virulence; Virulence Factors; Virulent; cysteinylglycine; extracellular; foodborne; foodborne illness; foodborne infection; foodborne pathogen; mouse model; novel; novel therapeutics; pathogen; pathogenic bacteria; response; transcription factor; transposon sequencing; uptake

ABSTRACT Listeria monocytogenes is a low G+C Gram-positive bacterium that can cause severe disease in immunocompromised individuals, pregnant women, newborns, and the elderly. Pathogenic listerial strains use a master transcriptional regulator, PrfA, to induce its most important virulence genes. In turn, the PrfA protein is directly activated by glutathione, a cysteine-containing tripeptide, which also performs important antioxidation and detoxification functions in bacterial and eukaryotic cells. To reach the intracellular level of glutathione required for sufficient PrfA activation and virulence gene expression during infection, the listerial cells need either to synthesize glutathione or import it from the cytosol of host cells, which is rich in glutathione. Surprisingly, listerial glutathione synthesis appears to contribute more strongly to virulence than its uptake from the eukaryotic cytosol. Glutathione uptake pathways have been identified only in several bacterial species; the L. monocytogenes glutathione importer(s) and the reasons for their apparent low activity during infection are not known. To synthesize glutathione and just to grow, L. monocytogenes cells must obtain cysteine, an essential amino acid, or a related compound from the environment. Interestingly, listerial cells can convert efficiently exogeneous glutathione to cysteine. This glutathione degradation, depending on the yet unknown extracellular or intracellular localization of this pathway, can reduce the metabolite level in bacterial or host cells or both. Therefore, a tight regulation of glutathione degradation may be a critical step in listerial virulence during infection. The glutathione-to-cysteine degradation pathway in L. monocytogenes has not been identified, and none of the genes encoding known enzymes of glutathione cleavage are present in the listerial genome. Thus, a novel enzyme of glutathione degradation with an unknown cellular localization is present in listerial cells. The complete lack of information on the nature of the glutathione uptake and degradation pathways, their regulation, and contributions to the glutathione level and PrfA activation impedes our understanding of how virulence genes are induced under various conditions of listerial growth. We propose to fill this important gap in our knowledge and determine the L. monocytogenes genes that are involved in the uptake and degradation of glutathione. Expression patterns of these genes under various growth conditions will be determined. The impact of the corresponding pathways on the glutathione listerial pool, expression of PrfA- dependent virulence genes, and virulence in a mouse model of infection will be major targets of our research. L. monocytogenes is one of the deadliest foodborne pathogens in the United States. This project will allow us to identify metabolic steps required for the accumulation of glutathione at levels that are needed for listerial virulence. In doing so, we may uncover novel pathways for potential therapeutic intervention. Similar pathways are likely to exist in other pathogenic bacteria.

抽象的 单核细胞增生李斯特菌是一种低的G+C革兰氏阳性细菌，可能导致严重的疾病 免疫功能低下的人，孕妇，新生儿和老年人。病原裂菌株的使用 主要的转录调节剂PRFA诱导其最重要的毒力基因。反过来，PRFA蛋白是 直接被含有半胱氨酸的三肽谷胱甘肽直接激活，也执行重要的抗氧化 和排毒功能在细菌和真核细胞中。 要达到足够PRFA激活和毒力基因所需的细胞内谷胱甘肽水平 感染期间的表达，李斯特细胞需要合成谷胱甘肽或从细胞质中导入它 宿主细胞，富含谷胱甘肽。令人惊讶的是，李斯特氏谷胱甘肽合成似乎有助于更多 强烈的毒力比其来自真核细胞质的摄取。谷胱甘肽的吸收途径已经 仅在几种细菌物种中鉴定； L.单核细胞增生谷胱甘肽进口商（S）和原因 它们在感染期间的表观低活动尚不清楚。 要合成谷胱甘肽并仅生长，单核细胞增生李斯特菌细胞必须获得半胱氨酸，这是必不可少的 氨基酸或来自环境的相关化合物。有趣的是，锂细胞可以有效地转换 外止的谷胱甘肽到半胱氨酸。这种谷胱甘肽降解，具体取决于尚未知道的细胞外 或该途径的细胞内定位，可以降低细菌或宿主细胞中的代谢物水平，或两者兼而有之。 因此，严格调节谷胱甘肽降解可能是李斯特式毒力的关键步骤 感染。尚未鉴定出单核细胞增生李斯特氏菌中的谷胱甘肽到半胱氨酸降解途径，并且 李斯特氏基因组中没有一个编码已知谷胱甘肽裂解酶的基因。因此， 李斯特氏细胞中存在一种具有未知细胞定位的谷胱甘肽降解的新型酶。 完全缺乏有关谷胱甘肽摄取和降解途径性质的信息， 它们的监管以及对谷胱甘肽水平和PRFA激活的贡献阻碍了我们对 在层裂生长的各种条件下，如何诱导毒力基因。我们建议填补这个重要的 在我们的知识上差距，并确定参与吸收和涉及的单核细胞增生基因基因 谷胱甘肽的降解。这些基因在各种生长条件下的表达模式将是 决定。相应途径对谷胱甘肽层池的影响，prfa-的表达 依赖性的毒力基因以及感染小鼠模型中的毒力将是我们研究的主要靶标。 单核细胞增生李斯特氏菌是美国最致命的食源性病原体之一。这个项目将 允许我们确定谷胱甘肽在所需的水平上积累所需的代谢步骤 李斯特式毒力。这样，我们可能会发现潜在治疗干预的新途径。相似的 途径可能存在于其他致病细菌中。