Decoding the signature of sperm RNA & RNA modification of environmental stressors on the intergenerational transmission of metabolic phenotypes

解码精子 RNA 的特征

• 批准号: 10250396
• 负责人: Qi Chen
• 金额: $ 43.49万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10250396
• 关键词: Arsenic; Arsenites; Biogenesis; Bioinformatics; Biological; Cells; Code; Comparative Study; Complex; Computer software; Data; Diet; Embryo; Embryonic Development; Endocrine Disruptors; Environment; Environmental Exposure; Enzymes; Exposure to; Future; Genetic Transcription; Genets; Health; Heterogeneity; High Fat Diet; Individual; Inherited; Injections; Knowledge; Lead; Ligation; Liquid Chromatography; Mammals; Masks; Mediating; Messenger RNA; Metabolic; Metabolic Diseases; Methods; Methyltransferase; Modification; Molecular; Nature; Obesity; Oxidative Stress; Pathologic; Pathologic Processes; Performance; Phenotype; Physiological; Property; Protocols documentation; RNA; RNA-Directed DNA Polymerase; Ribosomes; Science; Site; Small RNA; Specificity; Testing; Therapeutic Intervention; Transfer RNA; Translating; Untranslated RNA; comparative; environmental stressor; glucose metabolism; improved; interest; intergenerational; lipid metabolism; male; metabolic phenotype; mouse model; novel; obesogenic; offspring; precision medicine; programs; ribosome profiling; sperm cell; tandem mass spectrometry; transcriptome sequencing; transcriptomics; transmission process; tributyltin

Project Summary Emerging evidence has shown that small non-coding RNAs (sRNAs) harbor a diversity of RNA modifications. RNA modifications have the potential to store a secondary layer of labile biological information that is responsive to various environmental exposures and can modulate RNA properties such as stability and interaction potential, thus contributing to complex physiological/pathological processes. In our previous mouse model of paternal high- fat diet (HFD)-induced intergenerational inheritance, we found that tRNA-derived small RNAs (tsRNAs) and RNA methylatranserase (Dnmt2)-mediated site-specific RNA modification established a “sperm RNA code” that is required for intergenerational transmission of paternally acquired metabolic disorders (Science 2016; Nat Cell Biol 2018). These data, along with others, support an emerging concept that RNA modifications in sperm small RNAs serve as an additional layer of paternal hereditary information that can be modulated by environmental input, and is essential for regulating offspring phenotype via embryo development. These advances have set the stage to further examine whether a wider range of paternal environmental exposures, such as tributyltin (TBT) and arsenite (both are known to associate with obesity and metabolic disorders) will similarly alter sperm RNAs to confer offspring phenotype. This concerns the nature of the core sperm RNA code (i.e. a group of modified tsRNAs) shared by different exposure that is responsible for the intergenerational phenotype transmission; and also the molecular mechanism by which the modified sperm tsRNAs regulate embryo development to dictate offspring’s metabolic performance. In present project, we aim to first decipher the essential sperm tsRNAs & associated RNA modifications that responsible for programming offspring metabolic health, by comparatively studying different paternal environmental stressors (HFD, TBT & arenite exposure) with improved small RNA- seq protocol, which reduces sequencing bias by enzymatically removing RNA modifications that block reverse transcriptase and terminal adaptor ligation; we also explore the upstream regulators of the altered sperm tsRNAs, with a focus on RNA modifications enzymes (Aim 1). We will further isolate individual tsRNAs followed by RNA modification quantification using Liquid Chromatography-tandem Mass Spectrometry (LC-MS/MS), and test their function in conferring offspring phenotype by zygotic RNA injection and offspring phenotype tracking (Aim 2). Mechanistically, we will test the hypothesis whether modified sperm tsRNAs can program the metabolic state by regulating ribosome heterogeneity that control distinct translational pool of mRNAs (Aim3). In other words, we propose that environmental stressor-induced “sperm RNA code” is transformed into an “embryonic ribosome code”, which generates translational specificity to define the metabolic phenotype of offspring. Data from the proposed study may not only reveal the nature and mechanism of metabolic disorder related sperm RNA code, but also generate fundamental knowledge for future therapeutic intervention facing the obesogenic environment.

项目摘要 新出现的证据表明，小的非编码RNA（sRNA）具有多种RNA修饰。 RNA修饰有可能储存第二层不稳定的生物信息， 并能调节RNA的性质，如稳定性和相互作用潜力， 从而导致复杂的生理/病理过程。在我们之前的小鼠模型中- 高脂饮食（HFD）诱导的代际遗传，我们发现，tRNA衍生的小RNA（tsRNA）和RNA 甲基转移酶（Dnmt 2）介导的位点特异性RNA修饰建立了一个“精子RNA密码”， 父亲获得性代谢障碍的代际传播所需的（Science 2016; Nat Cell Biol 2018）。这些数据，沿着其他数据，支持了一个新兴的概念，即精子中的RNA修饰小 RNA作为父系遗传信息的额外一层，可以通过环境调节 输入，并通过胚胎发育调节后代表型是必不可少的。这些进步使 进一步研究是否有更广泛的父亲环境接触，如三丁基锡（TBT） 和亚砷酸盐（两者都与肥胖和代谢紊乱有关）同样会改变精子RNA 以赋予后代表型。这涉及核心精子RNA密码的性质（即一组修饰的 由负责代际表型传递的不同暴露所共享的（tsRNA）;以及 修饰的精子tsRNA调节胚胎发育的分子机制也决定了 后代的代谢表现。在本项目中，我们的目标是首先破译精子必需的tsRNA & 相关的RNA修饰，负责规划后代的代谢健康，通过比较 研究不同的父亲环境压力（HFD，TBT和砂暴露）与改进的小RNA- seq方案，通过酶促去除阻断逆转录酶的RNA修饰来降低测序偏倚。 转录酶和末端接头连接;我们还探索了改变的精子tsRNA的上游调节因子， 重点是RNA修饰酶（Aim 1）。我们将进一步分离单个的tsRNA， 使用液相色谱-串联质谱法（LC-MS/MS）进行修饰定量，并测试其 通过合子RNA注射和后代表型跟踪赋予后代表型的功能（目的2）。 从机制上讲，我们将测试修饰的精子tsRNA是否可以通过以下方式编程代谢状态的假设： 调节核糖体异质性，控制不同的mRNA翻译库（Aim 3）。换句话说我们 提出环境应激诱导的“精子RNA密码”转化为“胚胎核糖体 编码”，其产生翻译特异性以定义后代的代谢表型。的数据 这项研究不仅可以揭示与精子RNA编码相关的代谢紊乱的本质和机制， 而且还为未来面对致肥胖环境的治疗干预提供了基础知识。