Host tRNA-derived small RNAs (tsRNAs) mediate interactions between host and oral microbes

宿主 tRNA 衍生的小 RNA (tsRNA) 介导宿主和口腔微生物之间的相互作用

• 批准号: 10577837
• 负责人: Xuesong He
• 金额: $ 49.61万
• 依托单位: ADA FORSYTH INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-22 至 2027-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10577837
• 关键词: Address; Affect; Bacteria; Bacterial Genes; Bacterial Proteins; Behavior; Biochemical; Biochemistry; Biological Models; Biological Process; Carrier Proteins; Cell Line; Cells; Communication Programs; Complex; Coupled; Data; Defense Mechanisms; Disease; Dissection; Encapsulated; Epithelial Cells; Exhibits; Fluorescence Microscopy; Follow-Up Studies; Fusobacterium nucleatum; Gene Expression; Genes; Goals; Growth; Health; Homeostasis; Host Defense; Human; Image; Infectious Agent; Intake; Knowledge; Maintenance; Mediating; Membrane; Membrane Transport Proteins; Microbe; Mouth Diseases; Mucous Membrane; National Institute of Dental and Craniofacial Research; Natural Immunity; Oral; Oral health; Oral mucous membrane structure; Pb in saliva; Periodontitis; Plants; Play; Porphyromonas; Predisposition; Process; Protein Binding Domain; Protocols documentation; Publishing; RNA; RNA Recognition Motif; Regulatory Element; Regulatory Pathway; Reporting; Research; Resolution; Role; Saliva; Salivary; Small RNA; Strategic vision; Streptococcus mitis; Surface; Testing; Therapeutic; Transfer RNA; United States National Institutes of Health; Work; adaptive immunity; antimicrobial; bacterial genetics; beneficial microorganism; cell immortalization; exosome; extracellular; genetic analysis; gut microbiome; host microbiome; host-microbe interactions; human subject; keratinocyte; microbial; microbial host; microbiome; novel therapeutic intervention; nucleic acid-based therapeutics; opportunistic pathogen; oral bacteria; oral commensal; oral microbiome; pathogen; periodontopathogen; prevent; response; saliva sample; social; trafficking; transcriptome sequencing

Host mucosal surfaces are highly specialized and possess a complex array of innate and adaptive immunity. They provide the first line of protection against infectious agents by initiating protective responses to potential pathogens. Furthermore, the symbiotic relationship of the hundreds of microbial species with the host requires a fine-tuned response at the mucosal surface that prevents overgrowth of opportunistic pathogens, but sparing beneficial microbes. While small RNAs (sRNAs) have been regarded as a class of intracellular regulatory elements, emerging studies in plant-pathogen and host-gut microbiome interactions uncover that hosts can exploit sRNAs, such as transfer RNA (tRNA)-derived sRNAs (termed “tsRNAs”), via encapsulation in exosomes, as a new mode of bacterial gene modulation or defense mechanism. Yet, it remains to be elucidated whether such sRNA-mediated inter-kingdom gene modulation or defense mechanism exist in the context of host-oral microbiome interaction. To address this knowledge gap, the present R01 application will focus on host-derived, exosome-borne, salivary tsRNAs, of which the biological functions have remained largely elusive to date. Speficically, we recently demonstrated that human Normal Oral Keratinocyte- Spontaneously Immortalized (NOKSI) cells released two exosome-borne tsRNAs, tsRNA-000794 and tsRNA- 020498, when challenged with Fusobacterium nucleatum (Fn), a key oral commensal and opportunistic periodontal pathogen. Importantly, these two tsRNAs can be readily detected from saliva in healthy human subjects. Intriguingly, both tsRNAs exhibit highly selective, Fn-targeting antimicrobial activity—directly adding synthetic mimics of these two tsRNAs, but not scramble RNA, to bacterial culture inhibits the growth of Fn, but not that of Porphyromonas ginigivalis (Pg), a gram-negative periodontal pathogen or Streptococcus mitis (Sm), a health-associated oral bacterium. In preliminary work, we further took a multi-facet approach to identify a putative RNA-specific membrane transporter as well as candidate intracellular bacterial protein targets of Fn- targeting tsRNAs. Building on our comprehensive preliminary data, the goal of this application is two-fold: (Part I) achieve mechanistic understanding of the cross-kingdom trafficking of host-derived Fn-targeting tsRNAs and their modulating effect on Fn growth during NOKSI-Fn interaction, through exosome tracking and in-depth dissection of the tsRNAs transporter and intracellular targets in Fn; (Part II) expand our work to profile and compare salivary tsRNAs between healthy and periodontitis subjects, with a focus on demonstrating the broad implication of host-generated tsRNAs as a conserved mechanism to achieve host-microbial homeostasis. The realization of this application will not only address a fundamental question by defining tsRNAs as a new class of host defense molecules in maintaining host-microbiome homeostasis via targeted microbial modulation, but also will pave the way for a new therapeutic strategy against oral diseases, considering the already successful trajectory of nucleic acid therapeutics in recent years.

宿主粘膜表面是高度特化的，具有复杂的先天性和适应性免疫。 它们通过启动对潜在病原体的保护性反应，提供对抗传染性病原体的第一线保护。 病原体此外，数百种微生物物种与宿主的共生关系需要 粘膜表面的微调反应，可防止机会性病原体的过度生长， 有益微生物虽然小RNA（sRNA）已被认为是一类细胞内调节蛋白， 元素，植物-病原体和宿主-肠道微生物组相互作用的新兴研究揭示，宿主可以 利用sRNA，如转移RNA（tRNA）衍生的sRNA（称为“tsRNA”），通过包封在 外泌体作为细菌基因调控或防御机制的一种新模式。然而，它仍然是 阐明了这种sRNA介导的界间基因调节或防御机制是否存在于 宿主-口腔微生物组相互作用的背景。为了解决这一知识差距，目前的R 01应用程序将 重点是宿主来源的，外泌体携带的，唾液的tsRNA，其生物学功能仍然存在 迄今为止基本上难以捉摸。特别是，我们最近证明，人正常口腔角质形成细胞- 自发永生化（NOKSI）细胞释放两种外泌体携带的tsRNA，tsRNA-000794和tsRNA-000794。 020498，当用具核梭杆菌（Fn）攻击时， 牙周病原体重要的是，这两种tsRNA可以容易地从健康人的唾液中检测到 科目有趣的是，两种tsRNA都表现出高度选择性的Fn靶向抗微生物活性-直接加入 这两种tsRNA的合成模拟物，而不是scramble RNA，对细菌培养物抑制Fn的生长， 而不是革兰氏阴性牙周病原体牙龈卟啉单胞菌（Pg）或缓症链球菌（Sm）， 一种与健康相关的口腔细菌。在初步工作中，我们进一步采取了多方面的方法来确定一个 假定的RNA特异性膜转运蛋白以及Fn-的候选细胞内细菌蛋白靶点， 靶向tsRNA。基于我们全面的初步数据，本申请的目标有两个：（第一部分） I）实现对宿主来源的Fn靶向tsRNA的跨界运输的机制理解，以及 在NOKSI-Fn相互作用期间，它们对Fn生长的调节作用，通过外泌体跟踪和深入研究 tsRNAs转运蛋白和Fn细胞内靶点的解剖;（第二部分）扩展我们的工作， 比较健康和牙周炎受试者之间的唾液tsRNA，重点是证明广泛的 宿主产生tsRNA作为实现宿主-微生物稳态的保守机制的意义。的 该应用的实现将不仅通过将tsRNA定义为新的类别来解决基本问题， 宿主防御分子通过靶向微生物调节维持宿主微生物组稳态， 考虑到已经取得的成功， 近年来核酸疗法的发展轨迹。