Functional Portraits of tRNA-derived Small Non-coding RNAs

tRNA 衍生的小非编码 RNA 的功能肖像

基本信息

批准号：
8968710
负责人：
Xiaoyong Bao
金额：
$ 23.25万
依托单位：
UNIVERSITY OF TEXAS MED BR GALVESTON
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-06-15 至 2017-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8968710
关键词：
Algorithms Animals Bioinformatics Biological Biological Assay Biological Markers Biological Process Biological Response Modifiers Code Collaborations Communities Consensus Core Facility Cytoplasm Data Databases Development Disease Elements Eukaryota Event Exhibits Family Funding Future Gene Expression Gene Expression Regulation Gene Silencing Genes Genetic Transcription Genetic Translation Genomics Goals Home environment Human Human Genetics Learning Length Mediating Messenger RNA Methods MicroRNAs Molecular Molecular Biology Mutagenesis National Heart, Lung, and Blood Institute Nuclear Nucleotides Organism Plants Play Portraits Prevention Prokaryotic Cells Proteins Proteomics RNA Regulation Regulator Genes Research Resources Respiratory syncytial virus Role Solid System Testing The Sun Transcriptional Regulation Transfer RNA Translations Untranslated RNA Viral Viral Genes Virus Diseases Virus Replication Western Blotting Work base chemokine cytokine experience genome-wide human DICER1 protein improved mRNA Stability mRNA Transcript Degradation novel novel therapeutics public health relevance response

项目摘要

DESCRIPTION (provided by applicant): Recent discoveries about small non-coding RNAs (sncRNAs) have significantly advanced human genetics and molecular biology, largely due to the identification of a fundamental role of microRNAs (miRNAs), the best- characterized sncRNA family, as gene regulators. tRNA-derived RNA Fragments (tRFs) are a recently discovered sncRNA family that is ubiquitously expressed in organisms ranging from prokaryotes to humans, yet their biological functions and the mechanism(s) underlying those functions are largely unknown. Studies by our group have shown that the sncRNAs most highly induced by respiratory syncytial virus (RSV) belong to the tRF family, while the change in miRNA expression is minimal. These induced tRFs are functional and involved in the regulation of RSV replication. Notably, at least three tested tRFs have a gene trans- silencing function that is mechanistically distinct from that of miRNAs. However, more experimental evidence is needed to determine whether there is a widespread impact of mammalian tRFs on the regulation of gene expression, and more importantly, what mechanism(s) tRFs use for their gene regulatory function. In this project we propose the novel hypothesis that functional tRFs induced by RSV have a common gene trans-silencing function. Since RSV-induced tRFs are localized in the cytoplasm (and so are unlikely to play a role in regulating gene transcription, usually a nuclear event), we also hypothesize that the gene-suppression effect of tRFs occurs at post-transcriptional steps, e.g., mRNA stability or translation. In this exploratory project, these hypotheses will be tested in two Specific Aims. In Aim 1, tRFs important for RSV replication, viral gene expression and associated chemokine/cytokine induction will be identified. We will then determine whether these functional tRFs have gene trans-silencing activity. Aim 2 will test our second hypothesis that the gene- suppression effect of tRFs occurs at post-transcriptional steps. High-throughput methods, including microarray or proteomics analyses, will then be used to identify target candidates for a representative tRF. Target candidates will be experimentally confirmed by qRT-PCR and Western blot analysis. A mutagenesis study will also be performed to confirm that the effect of the tRF on its target(s) is direct and specific. This project may hav important translational implications by suggesting new therapeutic opportunities to modulate viral replication, providing experimental evidence to build a database of functional tRFs, and of more importance to identify the consensus features of tRF*target interaction for the development of a tRF target- prediction algorithm, which will undoubtedly facilitate the discovery of new gene regulatory networks and so will broadly benefit the research community. These goals will be explored in a future R01 application based on the results of this exploratory project.

描述（由申请人提供）：关于小非编码 RNA (sncRNA) 的最新发现显着推进了人类遗传学和分子生物学的发展，这主要是由于对 microRNA (miRNA)（最典型的 sncRNA 家族）的基本作用的识别，如tRNA 衍生的 RNA 片段 (tRF) 是最近发现的 sncRNA 家族，在从原核生物到人类的生物体中普遍表达，但它们的我们小组的研究表明，呼吸道合胞病毒 (RSV) 诱导程度最高的 sncRNA 属于 tRF 家族，而 miRNA 表达的变化很小。诱导的 tRF 具有功能并参与 RSV 复制的调节值得注意的是，至少三个测试的 tRF 具有与 miRNA 机制不同的基因反沉默功能。需要确定哺乳动物 tRF 是否对基因表达的调节存在广泛影响，更重要的是，tRF 使用什么机制来发挥其基因调节功能。在本项目中，我们提出了 RSV 诱导功能性 tRF 的新假设。由于 RSV 诱导的 tRF 位于细胞质中（因此不太可能在调节基因转录（通常是核事件）中发挥作用），因此我们还发现了基因抑制效应。 tRF 发生在转录后步骤，例如 mRNA 稳定性或翻译。在这个探索性项目中，这些假设将在两个具体目标中进行测试。在目标 1 中，tRF 对 RSV 复制、病毒基因表达和相关趋化因子/细胞因子诱导非常重要。然后我们将确定这些功能性 tRF 是否具有基因反式沉默活性。目标 2 将检验我们的第二个假设，即 tRF 会产生基因抑制作用。然后，将使用高通量方法（包括微阵列或蛋白质组学分析）来鉴定代表性 tRF 的靶标候选物。还将通过 qRT-PCR 和蛋白质印迹分析对靶标候选物进行实验确认。进行以确认 tRF 对其靶标的影响是直接且特异的。该项目可能通过提出调节病毒复制的新治疗机会、为建立功能数据库提供实验证据而具有重要的转化意义。 tRF，更重要的是确定 tRF* 靶点相互作用的共识特征，以开发 tRF 靶点预测算法，这无疑将促进新基因调控网络的发现，从而广泛惠及研究界。基于该探索性项目的结果，将在未来的 R01 应用中进行探索。