A novel class of long non-coding RNA in regulation of the type I interferonresponse

项目摘要 哺乳动物基因组被广泛转录以产生复杂的非编码转录组。一本小说 一类被称为狗的长非编码RNA(LncRNA)，其含有包含基因下游的转录本 最近发现的。为了应对各种压力，如病毒感染，狗的RNA通过以下方式表达 在一些蛋白质编码基因末端持续至少5kb的直读转录。值得注意的是， 这种以前未知的RNA物种作为一个群体，占所有基因间转录本的高达30%，但他们的 其功能仍完全未知。我发现对病毒核酸的先天免疫感知和 激活I型干扰素(干扰素)抗病毒反应导致数千只狗的RNA表达 基因，最明显的是来自干扰素基因本身。因此，我假设狗的RNA可能起到 调节其上游起源基因，这是先天抗病毒的一个基本调节过程 回应。从机制上讲，我认为狗的rna重塑了染色质和/或转录。 其上游基因的景观达到了一种平稳的状态。这对生物体是有益的，因为 可以实现对病毒感染的快速反应。在目标1中，我将评估狗RNA的作用 IfNB1基因(IfNB1-DOG RNA)通过基因、转录和后转录调控干扰素-β的表达 控制狗RNA水平的转录方法。在目标2中，我将研究分子基础 Ifnb1-狗RNA的功能是通过表征Ifnb1基因位点的染色质和转录图谱来实现的。 我将测试狗的RNA表达是否会影响转录后加工、表观遗传标记和染色质 建筑。在目标3中，我将确定狗RNA在体内的生理重要性。我将生成Ifnb1- 狗RNA缺陷小鼠，首先产生针对Ifnb1下游的引导RNA敲入小鼠，然后 通过将这些基因与dCas9转基因小鼠杂交。我将研究Ifnb1-Dog RNA对先天性免疫的影响 在动态平衡和病毒感染期间与这些小鼠。I型干扰素应答是 我们的抗病毒防御机制，但会导致人类疾病，如I型干扰素病和全身性 当红斑狼疮调节失调时。本研究旨在加深对I型干扰素调控的认识。 通过对一类新的lncRNAs进行功能表征。从这项研究中获得的见解可能会 用于开发针对感染性疾病和干扰素介导的自身免疫性疾病的改进疗法。

Project Summary The mammalian genome is pervasively transcribed to generate a complex non-coding transcriptome. A novel class of long non-coding RNA (lncRNA) called DoGs for “Downstream-of-Gene”-containing transcripts was recently discovered. In response to various stressors such as viral infections, DoG RNAs are expressed by readthrough transcription that continues past the ends of some protein-coding genes for at least 5kb. Remarkably, this previously unknown RNA species as a group accounts for up to 30% of all intergenic transcripts, yet their function remains completely unknown. I discovered that innate immune sensing of viral nucleic acids and activation of the type I interferon (IFN) antiviral response leads to expression of DoG RNAs from thousands of genes, most notably from the IFN gene itself. I therefore hypothesize that a DoG RNA may function to regulate its upstream gene of origin, and that this is an essential regulatory process in innate antiviral responses. Mechanistically I propose that a DoG RNA remodels the chromatin and/or transcriptional landscape of its upstream gene to achieve a poised state. This would be beneficial for organisms because rapid responses to viral infections can be achieved. In Aim 1, I will evaluate the role of the DoG RNA from the Ifnb1 gene (Ifnb1-DoG RNA) in regulating IFN-β expression using genetic, transcriptional, and post- transcriptional approaches to manipulate DoG RNA levels. In Aim 2, I will investigate the molecular basis of Ifnb1-DoG RNA function by characterizing the chromatin and transcriptional landscape of the Ifnb1 gene locus. I will test whether DoG RNA expression affects post-transcriptional processing, epigenetic marks, and chromatin architecture. In Aim 3, I will determine the physiological importance of DoG RNA in vivo. I will generate Ifnb1- DoG RNA deficient mice by first generating guide RNA knock-in mice that targets downstream of Ifnb1 and then by crossing these with dCas9 transgenic mice. I will study the effect of the Ifnb1-DoG RNA on innate immunity at homeostasis and during viral infections with these mice. The type I IFN response is a critical component of our antiviral defense mechanism, but can lead to human disease such as type I interferonopathies and systemic lupus erythematosus when dysregulated. This study aims to advance our understanding of type I IFN regulation by functionally characterizing a novel class of lncRNAs. Insights gained from this study can potentially be leveraged to develop improved therapeutics for infectious and IFN-mediated autoimmune diseases.