GLOBAL CONSEQUENCES OF INTERRUPTION OF MICROBIAL AUTOINDUCER SIGNALING

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 细菌分泌多种信号分子，使它们能够协调基因表达并发挥多细胞生物的作用。 为了响应这些“群体感应”或“自诱导剂”信号，毒力因子表达、生物膜形成和耐药性等医学上重要的表型以群体范围的方式受到调节。 5' 甲硫腺苷/S-腺苷高半胱氨酸核苷酶（MTA/SAH 核苷酶，MTN）在导致自诱导剂 I (AI-1) 和自诱导剂 II (AI-2) 形成的生物合成途径中占据中心位置。 此外，MTN 控制着 S-腺苷甲硫氨酸依赖性多胺合成和甲基化反应过程中消耗的甲硫氨酸和腺嘌呤回收的关键步骤。 MTN 的药理学或遗传抑制应阻止蛋氨酸和嘌呤的回收，通过抑制性 MTA 和 SAH 核苷的积累导致生长延迟，并干扰自诱导剂合成和下游信号依赖性过程。 为了研究这种酶在营养物回收和信号通路中的作用，将创建大肠杆菌 (O157:H7) 和肺炎克雷伯菌的 MTN 敲除菌株，并研究其生长变化（速率、碳利用率、生物膜形成）、体外感染模型以及体内定植和毒力的小鼠模型中哺乳动物细胞侵袭的减弱。 还将通过 LC/MS 和 NMR 检查蛋白质组学和代谢组学对 MTN 基因缺失的适应，以进一步表征酶中断的分子后果，并解释观察到的表型改变的基础。 最终，这些实验应该强调细菌中细胞信号传导作为新型抗生素开发目标的重要性。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Bacteria secrete a variety of signaling molecules that allow them to coordinate gene expression and behave as multicellular organisms. In response to these 'quorum sensing' or 'autoinducer' signals, such medically important phenotypes as virulence factor expression, biofilm formation, and drug resistance are modulated in a population wide manner. The enzyme 5' Methylthioadenosine / S-adenosylhomocysteine nucleosidase (MTA/SAH nucleosidase, MTN) occupies a central place in the biosynthetic pathways that lead to both autoinducer I (AI-1) and autoinducer II (AI-2) formation. In addition, MTN governs a crucial step in the recycling of methionine and adenine consumed during S-adenosylmethionine dependent polyamine synthesis and methylation reactions. Pharmacologic or genetic inhibition of MTN should block methionine and purine salvage, cause growth delays through the accumulation of inhibitory MTA and SAH nucleosides, and interfere with autoinducer synthesis and downstream signal dependent processes. To examine the role of this enzyme in nutrient salvage and signaling pathways, MTN knock-out strains of E. coli (O157:H7) and Klebsiella pneumoniae will be created and studied for alterations in growth (rate, carbon utilization, biofilm formation), attenuation of mammalian cell invasion in in vitro models of infection, and in murine models of in vivo colonization and virulence. Proteomic and metabolomic adaptations to MTN gene deletion will also be examined by LC/MS and NMR to further characterize the molecular consequences of enzyme interruption and explain the basis for observed alteration in phenotype. Ultimately, these experiments should underscore the importance of cellular signaling in bacteria as a target for novel antibiotic development.