Genome-wide Identification of Coordinated Post-transcriptional Gene Regulation by RNA Binding Proteins

项目摘要/摘要 RNA结合蛋白(RNAbindingProteins，RBPs)是基因表达的主要调控因子。 转录基因调节(PTGR)与许多 疾病，包括神经退行性疾病和癌症进展；然而， 在全基因组范围内对PTGR的协调控制仍然知之甚少。根据 最近对RNA相互作用组的研究，估计人类基因组编码~1500 限制性商业惯例，其中约600个能与mRNAs特异性结合。新开发的下一代网络的应用 测序工具，如PAR-CLIP，能够在全基因组上绘制限制性商业惯例的图谱 研究表明，单个RBP可以结合高达50%的表达的mRNAs。给定 限制性商业惯例的多样性及其针对转录组重要部分的倾向，a 单个转录本通常受多个限制性商业惯例的约束。因此，PTGR的最终结果是 依赖于与转录本绑定的限制性商业惯例的组合控制。有趣的是，限制性商业惯例 以相同的成绩单为目标并不总是彼此独立地行动。几个 合作或竞争结合mRNAs以控制PTGR的限制性商业惯例的实例 在文献中都有报道。然而，这些研究发现，协调的PTGR仅通过一个 限制性商业惯例很少，因此是对其活动的高度关注的观察。它仍然是未知的 限制性商业惯例如何在全基因组范围内协调PTGR的组合控制。考虑到他们的 与许多疾病的临床相关性，迫切需要确定监管 限制性商业惯例在基因组水平上的相互作用。如果没有这样的信息，科学地理解 PTGR的疾病状态和发病机制之间的联系将是不完整的。在目标1中 将使用每个公开可用的和内部生成的PAR-CLIP数据集来识别RBP 它们共同靶向相似的mRNAs组，或沿着mRNAs结合共同定位的位点。数据分析将 使用已建立的PAR-CLIP分析流水线结合客户 分析脚本。在目标2中，协调组合PTGR的机制将是 通过rna-seq、免疫共沉淀和原位检测候选限制性商业惯例对 共同本地化实验。第一对待表征的限制性商业惯例，METTL1和CDK1， 已经通过我们对公开的PAR-CLIP数据的初步分析确定了。 这些结果将为通过协调行动分析PTGR提供一个框架 限制性商业惯例，从而为研究机制或 临床重要基因的调控系统。

PROJECT SUMMARY/ABSTRACT RNA binding proteins (RBPs) are major controllers of gene expression through post- transcriptional gene regulation (PTGR) and have been implicated in the onset of numerous disorders, including neurodegenerative diseases and cancer progression; however, the coordinated control of PTGR on a genome-wide scale remains poorly understood. According to recent surveys of the RNA interactome, the human genome is estimated to encode ~1500 RBPs, of which ~600 specifically bind mRNAs. Application of newly developed next-generation sequencing tools, such as PAR-CLIP, that enable the mapping of RBPs on a genome-wide scale, revealed that a single RBP can bind up to 50% of expressed mRNAs. Given the multiplicity of RBPs and their propensity to target significant fractions of the transcriptome, a single transcript is often bound by multiple RBPs. As such, the final outcome for PTGR is dependent on the combinatorial control of the RBPs bound to a transcript. Interestingly, RBPs targeting the same transcript do not always act independently from each other. Several instances of RBPs either cooperatively or competitively binding mRNAs to control PTGR have been reported in the literature. However, these studies identified coordinated PTGR by only a few RBPs and is therefore a highly focused observation of their activity. It remains unknown how RBPs coordinate combinatorial control of PTGR on a genome-wide scale. Considering their clinical relevance to numerous diseases, there is a critical need to identify regulatory interactions of RBPs on a genomic scale. Without such information, scientific understanding of the link between the disease states and mechanisms of PTGR will be incomplete. In Aim 1 every publicly available and in-house generated PAR-CLIP dataset will be used to identify RBPs that co-target similar sets of mRNAs or bind co-localized sites along mRNAs. Data analysis will be carried out using an established PAR-CLIP analysis pipeline in combination with custom analytical scripts. In Aim 2 the mechanism of coordinated combinatorial PTGR will be determined for candidate pairs of RBPs through RNA-seq, co-immunoprecipitation and in situ co-localization experiments. The first pair of RBPs to be characterized, METTL1 and CDK1, were already identified through our preliminary analysis of publicly available PAR-CLIP data. These results will provide a framework for the analysis of PTGR by the coordinated action of RBPs, thereby providing critical knowledge to researchers studying the mechanism or regulatory system of clinically important genes.