Evolution and consequences of multidrug resistant ribosome

多重耐药核糖体的进化和后果

基本信息

批准号：
10264066
负责人：
Mee-Ngan F Yap
金额：
$ 45.33万
依托单位：
NORTHWESTERN UNIVERSITY AT CHICAGO
依托单位国家：
美国
项目类别：
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/10264066
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/antagonist innovation insight lincosamide next generation sequencing novel strategies preference public health relevance resistance mechanism ribosome profiling structural biology trait translatome

项目摘要

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.

抽象的细菌和真核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不清楚。普遍存在的二甲基化细菌 rRNA 中的保守腺嘌呤 A2058 会导致对所有三种药物的交叉耐药性重要的抗生素家族（大环内酯类、林可酰胺类和链阳菌素 (MLS)）。 A2058 二甲基化阻断了核糖体的 MLS，从而允许正常的蛋白质生物合成细菌生长。负责 A2058 二甲基化的三十五类 Erm 甲基转移酶不变地由前面有短核糖体停滞前导序列的双基因操纵子编码。这些短的停滞肽在大小和序列组成上有很大差异。功能性和人们对停滞序列与其同源 erm 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分子翻译停滞下游共转录 erm 的上调需要序列，但临床监测和我们的数据表明存在替代途径。我们未发表的数据进一步表明对不相关抗生素的附带敏感性、毒力基因表达的减少、无活性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。还有一个未满足的需求是了解 Erm 识别和作用于 23S rRNA 的机制。这该提案将采用多管齐下的方法，包括高精度下一代测序，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来解决三个核心问题：A2058 所赋予的权衡的根本机制是什么二甲基化核糖体？ erm操纵子是如何进化的，erm的表达是如何受到调控的？ Erm 如何找到其靶底物 RNA？ erm 操纵子在医院内广泛存在革兰氏阴性和革兰氏阳性细菌，解决这些问题将提供重要的机制深入了解专为破坏这些生化相互作用和监管而定制的新抗菌策略途径。