Regulatory and Mechanistic Understanding of ADAR-Mediated RNA Editing

项目概要/摘要 先天免疫受到良好的控制，能够及时、灵敏地对病原体感染作出反应， 容忍细胞中的"自我"分子。先天免疫调节的缺陷导致各种疾病， 例如自身免疫性疾病或对感染的高度脆弱性。我们和其他人以前的研究重新- vealed ADAR 1，一种RNA编辑酶，催化dsRNA上的腺苷到肌苷（A-to-I）编辑， 在调节对双链RNA（dsRNA）的先天免疫应答中起关键作用。ADAR1 RNA编辑 并且结合活性已经显示出防止内源性（"自身"）dsRNA激活胞质溶质 dsRNA传感器MDA5和PKR，但其潜在的分子机制还不清楚。此外，本发明还提供了一种方法， ADAR 1 p150同种型对至少一些dsRNA的细胞质编辑对于抑制dsRNA至关重要。 介导的自身免疫，尽管细胞核中的ADAR 1 p110亚型通常更丰富。如何 这些双链RNA是在细胞质中编辑而不是在细胞核中编辑的吗？ 在这个MIRA应用程序中，我们专注于两个项目来解决这些知识差距。首先，我们将阐明 ADAR1与dsRNA先天免疫途径中的其他参与者之间的相互作用。具体来说，我们将投资- 拮抗ADAR 1调节dsRNA传感的MDA5和PKR途径的机制。我们提出 ADAR1在RNA编辑依赖性和非依赖性两种模式下调节dsRNA介导的先天免疫， 离子。我们将研究这两种作用模式如何在体外运作，并测试它们在小鼠体内的影响 模型其次，我们将揭示细胞质与核编辑的调控机制。我们特别 将进行遗传筛选和生化分析，以确定和表征负责的因素， 编辑的空间差异。总之，这些创新的研究将提供一个深刻的理解 在先天免疫中dsRNA编辑和dsRNA传感的界面处操作的分子机制。

PROJECT SUMMARY/ABSTRACT The innate immunity is well-controlled to respond to pathogenic infection in a timely and sensitive manner, while tolerating “self” molecules in the cell. Defects in the regulation of innate immunity result in various disorders, such as autoimmune diseases or heightened vulnerability to infections. Previous studies by us and others re- vealed ADAR1, an RNA editing enzyme that catalyzes Adenosine to Inosine (A-to-I) editing on dsRNAs, as a key player in the regulation of innate immune response to double-stranded RNAs (dsRNAs). ADAR1 RNA editing and binding activities have been shown to prevent endogenous (“self”) dsRNAs from activating the cytosolic dsRNA sensors MDA5 and PKR, but the underlying molecular mechanisms are not well understood. In addition, the cytoplasmic editing of at least some dsRNAs by the ADAR1 p150 isoform is crucial to suppress the dsRNA- mediated autoimmunity, although the ADAR1 p110 isoform in the nucleus is generally a lot more abundant. How are these dsRNAs edited in the cytoplasm but not in the nucleus? In this MIRA application, we focus on two projects to address these knowledge gaps. First, we will elucidate the interplay between ADAR1 and other players in the dsRNA innate immunity pathways. Specifically, we will inves- tigate the mechanisms by which ADAR1 regulates the MDA5 and PKR pathways of dsRNA sensing. We propose that ADAR1 regulates dsRNA-mediated innate immunity in both RNA editing-dependent and -independent fash- ion. We will study how these two modes of action operate in vitro and test their in vivo implications in mouse models. Second, we will uncover the regulatory mechanisms for cytoplasmic vs. nuclear editing. Specifically, we will perform genetic screens and biochemical assays to identify and characterize the factors responsible for spatial differences in editing. Taken together, these innovative studies will provide a deep understanding of the molecular mechanisms operating at the interface of dsRNA editing and dsRNA sensing in innate immunity.