STRUCTURE AND FUNCTION OF DSRNA ADENOSINE DEAMINASE

描述：DRDA是一种将多种腺苷 在双螺旋RNA底物中没有明显的内在 序列特异性尽管它在整个动物中是保守的， 王国，DRDA的体内作用一直不清楚，因为它 1987年发现。然而，我们最近的发现和其他人的研究 强烈表明DRDA作为核RNA编辑酶， 调节重要基因转录本的表达，如 中枢神经谷氨酸门控离子通道受体 系统在上一次赠款支持期间，申请人成功地 在分离cDNA克隆DRDA。用这些克隆体来表达 她和她的同事已经获得了重组DRDA蛋白， 初步证据表明DRDA参与GluR RNA 编辑.在本申请中，他们提出解决问题的重点是 DRDA的生物学功能和天然底物RNA。使用 建立体外试验系统，他们将研究 深入研究GluR-B RNA编辑机制，作为模型基因系统， 更好地理解DRDA的行动。DRDA与GluR-B RNA的相互作用 网站选择编辑所需的基本辅助因素将是 研究了建议将辅因子纯化，并且其 cDNA克隆。纯化的辅因子蛋白将用于重构 GluR-B RNA编辑的体外研究。申请人还将 直接确定DRDA的体内靶RNA库。的 通过拟议的实验获得的信息应有助于增强 了解这一生物学功能的全部范围， 有趣的核酶

DESCRIPTION: DRADA is a nuclear enzyme that converts multiple adenosines to isosines in double-helical RNA substrates without apparent intrinsic sequence specificity. In spite of its conservation throughout the animal kingdom, the in vivo role of DRADA has remained obscure since its discovery in 1987. However, our recent findings and studies by others strongly suggest that DRADA acts as a nuclear RNA editing enzyme to regulate the expression of important gene transcripts such as the glutamate-gated ion channel receptors (GluR) of the central nervous system. During the last period of grant support, the applicant succeeded in isolating cDNA clones for DRADA. Using these clones to express recombinant DRADA protein, she and her colleagues have obtained preliminary evidence that demonstrate an involvement of DRADA in GluR RNA editing. In this application, they propose to address questions focussing on the biological functions and natural substrate RNAs of DRADA. Using an established in vitro assay system, they will investigate the mechanism of GluR-B RNA editing in depth, as a model gene system to better understand DRADA action. The interaction of DRADA and GluR-B RNA with an essential co-factor needed for site-selctive editing will be investigated. It is proposed that the co-factor be purified, and its cDNA cloned. The purified co-factor protein will be used to reconstitute the GluR-B RNA editing with DRADA in vitro. The applicant will also determine directly the repertoire of in vivo target RNAs of DRADA. The information gained by the proposed experiments should lead to enhanced understanding of the full range of biological functions of this intriguing nuclear enzyme.