Evolution and consequences of multidrug resistant ribosome

多重耐药核糖体的进化和后果

• 批准号: 10463844
• 负责人: Mee-Ngan F Yap
• 金额: $ 46.34万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-16 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10463844
• 关键词: Address; Adenine; Affect; Antibiotics; Antimicrobial Resistance; Bacterial Infections; Binding; Biochemical; Biochemical Genetics; Biochemistry; Biological Assay; Carotenoids; Catalysis; Cells; Cessation of life; Ciprofloxacin; Clinical; Communities; Complement; Complex; Coupled; Cryoelectron Microscopy; Crystallization; DNA Gyrase; Data; Disulfides; Enzymatic Biochemistry; Enzymes; Evolution; Exhibits; Family; Gene Expression; Generations; Genes; Genetic Polymorphism; Genome; Goals; Gram-Positive Bacteria; Growth; Health Care Costs; Impairment; In Vitro; Infection; Knowledge; Laboratories; Link; Macrolides; Maps; Mass Spectrum Analysis; Measures; Mediating; Medical; Messenger RNA; Methyltransferase; Methyltransferase Gene; Mining; Modeling; Modification; Molecular; Multi-Drug Resistance; Mutation; Nucleotides; Operon; Pathway interactions; Peptides; Population; Protein Biosynthesis; Proteome; Proteomics; RNA; RNA Binding; Regulatory Pathway; Reporter; Resistance; Ribosomal RNA; Ribosome Inactivation; Ribosomes; Role; Site; Site-Directed Mutagenesis; Staphylococcus aureus; Streptogramins; Structure; System; Testing; Therapeutic; Toll-like receptors; Translations; Treatment Failure; Up-Regulation; Virulence; antimicrobial; bacterial genetics; bacterial resistance; base; comparative genomics; cost; crosslink; cytotoxic; design; experimental study; fitness; gel mobility shift assay; genetic approach; genome sequencing; global health; human disease; human pathogen; improved; in vivo; inhibitor; innovation; insight; lincosamide; next generation sequencing; novel strategies; preference; public health relevance; resistance mechanism; ribosome profiling; structural biology; trait; translatome

ABSTRACT Posttranscriptional modifications of bacterial and eukaryotic ribosomes are linked to many human diseases, but the precise role of most modifications remains undefined. Dimethylation of a universally conserved adenine, A2058, in bacterial rRNA causes cross-resistance against all three critically important families of antibiotics (macrolides, lincosamides, and streptogramins (MLS)). A2058 dimethylation occludes MLS from the ribosome, thereby allowing normal protein biosynthesis and bacterial growth. The thirty-five classes of Erm methyltransferases responsible for A2058 dimethylation are invariantly encoded by a two-gene operon preceded by a short ribosome stalling leader sequence. These short stalling peptides considerably vary in size and sequence composition. The functional and evolutionary connections between the stalling sequence and its cognate erm gene are poorly understood. A previous `ribosome stalling' model suggests that macrolide-mediated translational stalling of the leader sequence is required for the upregulation of downstream co-transcribed erm, but clinical surveillance and our data indicate the existence of an alternative pathway. Our unpublished data further show that collateral sensitivity to unrelated antibiotics, reduction in virulence gene expression, accumulation of inactive ribosomes, and loss of in vivo fitness are all part of the trade-offs associated with the A2058 dimethylated ribosome. The exact mechanistic links between these traits are unknown. There is also an unmet need to understand the mechanism by which Erm recognizes and acts on 23S rRNA. This proposal will use a multi-pronged approach consisting of high-precision next-generation sequencing, bacterial genetics, proteomics, comparative genomics, biochemistry and structural biology to address three central questions: What are the underlying mechanisms of the trade-offs conferred by the A2058 dimethylated ribosome? How does the erm operon evolve, and how is the expression of erm regulated? How does Erm find its target substrate RNA? The erm operons are widespread among nosocomial Gram-negative and Gram-positive bacteria, addressing these questions will offer significant mechanistic insight into new antimicrobial strategies tailored to disrupt these biochemical interactions and regulatory pathways.

摘要 细菌和真核生物核糖体的转录后修饰与许多人类 疾病，但大多数修饰的确切作用仍然不确定。一个普遍的二甲基化 细菌rRNA中保守的腺嘌呤A2058引起对所有三种关键的交叉抗性， 重要的抗生素家族（大环内酯类、林可酰胺类和链阳性菌素（MLS））。A2058 二甲基化从核糖体封闭MLS，从而允许正常的蛋白质生物合成， 细菌生长负责A2058二甲基化的35类Erm甲基转移酶 由两个基因操纵子恒定编码，前面是短的核糖体停顿前导序列。 这些短的停滞肽在大小和序列组成上变化很大。的功能和 stalling序列和它的同源基因之间的进化联系知之甚少。 先前的“核糖体停滞”模型表明，大环内酯介导的前导序列的翻译停滞 序列是下游共转录的转录因子上调所必需的，但临床监测和 我们的数据表明存在替代途径。我们未公布的数据进一步表明， 对不相关抗生素的附带敏感性，毒力基因表达减少， 失活的核糖体和体内适应性的丧失都是与A2058相关的权衡的一部分。 二甲基化核糖体这些特征之间的确切机制联系尚不清楚。还有一个 然而，我们需要了解Erm识别和作用于23S rRNA的机制。这 该提案将使用多管齐下的方法，包括高精度的下一代测序， 细菌遗传学、蛋白质组学、比较基因组学、生物化学和结构生物学， 三个核心问题：A2058所赋予的权衡的基本机制是什么 二甲基化核糖体操纵子是如何进化的，以及操纵子的表达是如何调节的？ Erm如何找到它的靶底物RNA？在医院感染中， 革兰氏阴性菌和革兰氏阳性菌，解决这些问题将提供重要的机制 深入了解为破坏这些生化相互作用和调节而定制的新抗菌策略 途径。