Study the roles of RNA editing enzyme ADAR1 in the regulation of cell death

描述（由申请人提供）：ADAR1（作用于RNA -1的腺苷脱氨酶）是细胞存活和胚胎发育所必需的蛋白质。在小鼠中敲除编码该蛋白的基因会导致细胞迅速和广泛的死亡，从而导致胚胎死亡。ADAR1在细胞存活中的关键作用表明，在生理条件下，ADAR1在调节细胞存活和细胞死亡之间的平衡方面发挥着强有力的作用，这表明ADAR1的失调可能导致病理。研究ADAR1的长期目标是明确其功能和机制，以便在病理过程中靶向治疗ADAR1。我们的初步数据表明，在哺乳动物细胞中，ADAR1直接与NF-kB通路相互作用，并通过IKK蛋白复合物上调NF-kB活性。本应用的目的是确定ADAR1在调节NF-kB信号传导中的机制。核心假设是，ADAR1通过蛋白-蛋白相互作用调节NF-kB介导的基因表达，有助于调节细胞死亡和细胞存活之间的平衡。将讨论两个具体目标。目的1：确定NF-kB通路中ADAR1调控NF-kB靶基因表达的机制。将定义ADAR1和IKK2亚基在细胞因子刺激下的动态相互作用。蛋白质-蛋白质相互作用的生化和信号转导结果将被确定。目标2：确定ADAR1中与IKK2结合并调节NF-kB活性的功能域。通过在哺乳动物细胞中突变和截断的ADAR1的表达来确定ADAR1的蛋白结合域，并检测其结合IKK2和调节NF-kB靶基因转录的能力。阳性或阴性功能的蛋白片段也将被检测是否能挽救ADAR1缺乏症的细胞死亡表型。这些研究完成后，有望揭示ADAR1在NF-kB通路中的新功能及其作用机制。ADAR1中与NF-kB信号通路相互作用的功能域将被确定。这个蛋白质结构域可能会导致一种肽的发现，这种肽将用于治疗由于NF-kB信号不平衡和细胞死亡引起的疾病。这项研究有望在RNA编辑、信号转导和涉及细胞死亡机制的疾病领域产生重要影响，从而有助于改善人类健康。公共卫生相关性：本研究涉及与NFkB信号通路相关的疾病，如炎症、关节炎、感染性和恶性疾病。

DESCRIPTION (provided by applicant): ADAR1 (Adenosine Deaminase Acting on RNA -1) is an essential protein required for cell survival and embryonic development. Knockout of the gene encoding for this protein in mice gives rise to rapid and wide-spread cell death resulting in embryo death. The critical role of ADAR1 in cell survival indicates a potent role for ADAR1 in regulation of the balance between cell survival and cell death under physiologic conditions, and suggests that dysregulation of ADAR1 could lead to pathology. The long-term goal of the research of ADAR1 is to define its function and mechanism in order to therapeutically target ADAR1 in pathologic processes. Our preliminary data has shown that ADAR1 directly interacts with the NF-kB pathway and up-regulates NF-kB activities through IKK protein complex in mammalian cells. The objective in this application is to identify the mechanism of ADAR1 in the regulation of NF-kB signaling. The central hypothesis is that ADAR1 modulates NF-kB mediated gene expression through a protein-protein interaction that contributes to regulation of the balance between cell death and cell survival. Two specific aims will be addressed. Aim #1: Identify the mechanism in NF-kB pathway by which ADAR1 regulates NF-kB target gene expression. The dynamic interaction of ADAR1 and IKK2 subunit upon cytokine stimulation will be defined. The biochemical and signaling consequence of the protein- protein interaction will be determined. Aim #2: Determine the functional domain in ADAR1 that binds to IKK2 and regulates NF-kB activities. The protein binding domain of ADAR1 will be determined through expressions of mutated and truncated ADAR1 in mammalian cells and detect their capabilities to bind IKK2 and regulate NF-kB target gene transcriptions. The positively or negatively functioning protein fragments will also be tested whether they rescue the cell death phenotype of ADAR1 deficiency. At the completion of these studies, it is expected that a new function of ADAR1 in NF-kB pathway and the related mechanism will be revealed. A functional domain in ADAR1 interacts with NF-kB signal pathway will be identified. This protein domain may lead to a discovery of a peptide which will be used to treat diseases due to unbalance NF-kB signaling and cell death. This study is expected to have an important impact in the fields of RNA editing, signal transduction, and diseases that involve cell death mechanisms and therefore contribute to improve human health. PUBLIC HEALTH RELEVANCE: This research has relevance to diseases related to NFkB signaling pathway, such as inflammation, arthritis, infectious and malignant diseases.