"Ribonomics" of Gene Regulation to predict Innate Immune Responses

描述(申请人提供)：基因的表达涉及一系列的酶事件，如转录、信使核糖核酸加工、信使核糖核酸衰变和翻译，这些都受制于蛋白质-核酸相互作用的基因调控网络(GRN)。众所周知，通过调控增强子和启动子活性的调控网络对转录的控制并不是基因产物产生的唯一决定因素，但外显子跳过是普遍存在的，转录后机制，如mRNA剪接和衰退决定了mRNA诱导和丰度的动力学。事实上，在我们对巨噬细胞对病原体的反应的初步研究中，我们发现大多数诱导的基因表达事件导致的mRNAs与基因组浏览器预测的mRNAs有很大偏离，并且mRNAs的衰退受蛋白质-核酸和miRNA调控网络的共同控制。拟议的基因调控核经济学中心利用并开创了下一代测序和计算建模方法，以开发一个预测模型，用于预测mRNA异构体的表达以及给定的启动子活性和转录起始率处于什么水平。我们将开发普遍适用的工具，结合或深入地对巨噬细胞对病原体相关内毒素的反应进行定量的实验分析，这导致1000多个基因的显著上调。值得注意的是，我们的初步数据发现了900多个外显子跳过事件，以及大量替代5‘或3’剪接位点的使用案例，强调了启动后事件的重要贡献。此外，这些剪接模式依赖于巨噬细胞亚型特定的染色质景观，并在启动或耐受状态下被可诱导的剪接因子改变。因此，我们将利用描述良好的巨噬细胞生物学和相关的实验模型系统，研究基因结构和序列(目标1)、染色质修饰(目标2)和反式作用剪接因子(目标3)在确定成熟mRNA的身份及其相关合成速率方面的作用，然后添加刺激响应调节网络，赋予mRNA半衰期控制，从而确定每个mRNA亚型的丰度(目标4)。

DESCRIPTION (provided by applicant): The expression of genes involves a sequence of enzymatic events, such as transcription, mRNA processing, mRNA decay, and translation, that are subject to gene regulatory networks (GRNs) of protein-nucleic acid interactions. It is well appreciated that the control of transcription via regulatory networks that regulate enhancer and promoter activities are not the sole determinant of what gene products result, but that exon skipping is pervasive and post-transcriptional mechanisms such as mRNA splicing and decay determine the kinetics of mRNA induction and abundance. Indeed, in our preliminary studies of the macrophage response to pathogens, we find that a majority of induced gene expression events result in mRNAs that deviate substantially from those predicted by the genome-browser, and that mRNA decay is controlled by both protein- nucleic acid and miRNA regulatory networks. The proposed Center for the "Ribonomics" of Gene Regulation leverages and pioneers Next Gen Sequencing and computational modeling approaches to develop a predictive model for which mRNA isoforms are expressed and at what level given a given promoter activity and transcription initiation rate. We will develop generally applicable tools in conjunction with or in depth and quantitative experimental analysis of the macrophage response to pathogen-associated endotoxin, which results in the dramatic up regulation of more than 1000 genes. Strikingly, our preliminary data identified more than 900 exon skipping events in addition to numerous cases of alternative 5' or 3' splice site use, emphasizing the essential contribution of post-initiation events. Further, these splice patterns are dependent on the macrophage subtype-specific chromatin landscape and are altered by inducible splice factors in primed or tolerated states. Thus we will leverage the well-described macrophage biology and associated experimental model systems, to examine the role of gene structure and sequence (Aim 1), the role of chromatin modifications (Aim 2), and of trans-acting splicing factors (Aim 3) in determining the identity of mature mRNAs and their associated synthesis rates, before adding the stimulus-responsive regulatory networks that confer mRNA half-life control and thus determine the abundance of each mRNA isoform (Aim 4).