Regulation of programmed -1 ribosomal frameshifting by micro-RNAs

micro-RNA 对程序性 -1 核糖体移码的调节

• 批准号: 9278237
• 负责人: Jonathan D Dinman
• 金额: $ 29.68万
• 依托单位: UNIV OF MARYLAND, COLLEGE PARK
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-25 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9278237
• 关键词: 3-Dimensional; AIDS/HIV problem; Address; Affect; Animals; Area; Base Pairing; Basic Science; Biological Assay; Biological Models; CCR5 gene; Cell Line; Cell physiology; Cells; Cellular biology; Cis-Acting Sequence; Codon Nucleotides; Collaborations; Complex; Computer Simulation; Computing Methodologies; Cytokine Receptors; Data; Databases; Defect; Development; Developmental Biology; Elements; Eukaryota; Feedback; Foundations; Gene Expression; Genetic Code; Genome; Global Change; HIV-1; Health; Human; Immune response; Immune signaling; Immunology; In Vitro; JAK2 gene; Laboratories; Link; Mathematics; Mediating; Messenger RNA; Methods; MicroRNAs; Modeling; Molecular; Molecular Biology; Pathway interactions; Play; Preclinical Drug Evaluation; Protein Tyrosine Kinase; Proteins; Publishing; RNA; Reading Frames; Regulation; Reporter; Research; Resolution; Ribosomal Frameshifting; Ribosomes; Role; Sequence Analysis; Series; Signal Transduction; Single Nucleotide Polymorphism; Structure; Structure-Activity Relationship; T-Lymphocyte; Technology; Telomere Maintenance; Terminator Codon; Testing; Translational Research; Untranslated RNA; Work; Yeasts; base; biophysical techniques; cytokine; design; drug development; frontier; innovation; mRNA Decay; mRNA Transcript Degradation; next generation; novel; premature; protein expression; response; ribosome profiling; small molecule therapeutics; structural biology; therapeutic target; unpublished works; virology

PROJECT SUMMARY/ABSTRACT Since the genetic code was elucidated in the early 1960's, it has been assumed that mRNAs are always decoded in three base codons, that any deviation from this fundamental rule must be erroneous and thus, deleterious. However, over the past decade, we have shown that a significant fraction of cellular mRNAs harbor cis-acting sequence elements that direct elongating ribosomes to shift reading frame by one base in the 5' (-1) direction. In cellular mRNAs, such Programmed -1 Ribosomal Frameshift (-1 PRF) signals direct ribosomes to premature termination codons where they become substrates for rapid degradation through the Nonsense-Mediated mRNA Decay (NMD) pathway, resulting in decreased expression of the proteins encoded by these mRNAs. Importantly, rates of mRNA degradation are proportional to rates of -1 PRF, a relationship that is conserved in eukaryotes from yeast to humans. Observations from yeast to humans that global changes in -1 PRF rates are deleterious to cellular function suggested that regulation of -1 PRF must be sequence- specific. Recently, we discovered that this is achieved through the interactions between -1 PRF signals and miRNAs. These findings have initiated a completely new avenue of research by establishing -1 PRF on cellular mRNAs, and its regulation by miRNAs as a new, fundamental paradigm in gene expression. This proposal seeks to deepen our understanding of the molecular mechanisms underlying regulation of -1 PRF in human cells. The well-defined Jurkat human T-cell line and a focus on -1 PRF signals embedded in mRNAs encoding cytokine receptors and a critical cytokine-responsive tyrosine kinase provides a model system to address a series of questions ranging from basic molecular and structural biology to regulation and control of the acquired immune response. Aim 1 of this proposal seeks to confirm -1 PRF promoted by sequences identified in the mRNAs encoding IL2Rγ, IL7Rα, and JAK2, characterize the effects of SNPs on -1 PRF, and develop the next generation PRF technology. Aim 2 will identify and validate miRNAs that naturally interact with these -1 PRF signals, and will test an autoregulatory feedback loop model of -1 PRF. Aim 3 is oriented towards characterizing the effects of miRNAs on gene expression and RNA structure. By the end of the proposed studies, we will have 1) deepened our understanding of this new paradigm gene expression control, 2) identified specific miRNAs that are used by T-cells to control their responses to important cytokines, and 3) established new rules describing mRNA/miRNA atomic scale structural interactions. These studies will immediately impact many fields including basic molecular and cell biology, and more applied fields including immunology and HIV/AIDS, and will lay the foundation for the design and discovery of small molecule therapeutics targeted to specific -1 PRF signals.

项目总结/摘要 自从20世纪60年代早期遗传密码被阐明以来，人们一直认为mRNA总是与基因组中的蛋白质结合。 以三个碱基密码子解码，任何偏离这一基本规则的行为都是错误的，因此， 有害的然而，在过去的十年里，我们已经表明，细胞mRNA的一个显着的部分， 含有顺式作用序列元件，指导延长核糖体将阅读框移动一个碱基， 5'（-1）方向。在细胞mRNA中，这种程序性-1核糖体移码（-1 PRF）信号直接 核糖体到提前终止密码子，在那里它们成为快速降解的底物， 无义介导的mRNA衰变（NMD）途径，导致编码的蛋白质表达减少 通过这些mRNA。重要的是，mRNA降解速率与-1 PRF的速率成比例， 在从酵母到人类的真核生物中是保守的。从酵母到人类的观察表明全球变化 在-1PRF率是有害的细胞功能表明，调节-1PRF必须是顺序- 特定.最近，我们发现这是通过PRF信号和 miRNAs。这些发现开创了一个全新的研究途径， mRNAs及其通过miRNAs的调控作为基因表达的一种新的基本范式。这项建议 旨在加深我们对人类-1 PRF调节的分子机制的理解， 细胞明确定义的Jurkat人T细胞系和对嵌入在编码人T细胞的mRNA中的-1 PRF信号的关注。 细胞因子受体和一个关键的酪氨酸反应性酪氨酸激酶提供了一个模型系统，以解决 一系列问题，从基础分子和结构生物学，以调节和控制的， 获得性免疫反应本提案的目的1旨在确认由鉴定的序列促进的-1PRF 在编码IL 2 R γ、IL 7 R α和JAK 2的mRNA中，描述SNP对-1 PRF的影响，并开发 下一代PRF技术目标2将鉴定和验证与细胞因子相互作用的miRNAs。 这些-1\f25 PRF-1信号，并将测试-1\f25 PRF-1的自动调节反馈回路模型。目标3是定向 研究miRNAs对基因表达和RNA结构的影响。结束时 建议的研究，我们将有1）加深我们对这种新的范式基因表达控制的理解， 2)鉴定了T细胞用于控制其对重要细胞因子的应答的特异性miRNA，以及3） 建立了描述mRNA/miRNA原子尺度结构相互作用的新规则。这些研究将 直接影响包括基础分子和细胞生物学在内的许多领域，以及更多的应用领域， 免疫学和HIV/AIDS，并将为小分子的设计和发现奠定基础 靶向特定-1 PRF信号的治疗剂。