Regulatory and Mechanistic Understanding of ADAR-Mediated RNA Editing

项目摘要/摘要 先天免疫得到很好的控制，以及时和敏感的方式对病原体感染做出反应，而 耐受细胞中的“自我”分子。先天免疫调节的缺陷会导致各种紊乱， 例如自身免疫性疾病或感染的易感性增加。我们和其他人之前的研究重新审视了 小牛ADAR1，一种RNA编辑酶，催化dsRNA上的腺苷到肌苷(A-to-I)编辑，作为一种 在调节对双链RNA(DsRNAs)的先天免疫反应中起关键作用。ADAR1 RNA编辑 结合活性已被证明可以阻止内源(自身)dsRNAs激活胞浆 DsRNA是MDA5和PKR的传感器，但其潜在的分子机制尚不清楚。此外, ADAR1 P150亚型对至少部分dsRNA的细胞质编辑对于抑制dsRNA- 介导的自身免疫，尽管细胞核中的ADAR1 p110亚型通常要丰富得多。多么 这些dsRNA是在细胞质中编辑的，而不是在细胞核中编辑的吗？ 在这个Mira应用程序中，我们重点关注两个项目，以弥补这些知识差距。首先，我们将澄清 在dsRNA先天免疫途径中，ADAR1和其他参与者之间的相互作用。具体地说，我们将投资- 报道ADAR1调节dsRNA传感的MDA5和PKR通路的机制。我们建议 ADAR1在RNA编辑依赖和非依赖FASH中调节dsRNA介导的先天免疫 离子。我们将研究这两种作用模式在体外如何运作，并在小鼠身上测试它们的体内意义。 模特们。其次，我们将揭示细胞质编辑与核编辑的调控机制。具体来说，我们 将进行遗传筛查和生化分析，以确定和表征导致 编辑中的空间差异。综上所述，这些创新的研究将提供对 先天免疫中dsRNA编辑和dsRNA传感界面的分子机制。

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.