RNA-Protein interactions regulating mRNA stability

描述（由申请方提供）：珠蛋白基因表达得以维持， 由于珠蛋白mRNA的异常稳定性， 在分化成转录沉默的去核红细胞的过程中。 对调节珠蛋白mRNA的成分知之甚少 稳定mRNA稳定性的不稳定性会产生深远的后果， 表现为临床表型，包括地中海贫血， 地中海贫血，一个恒定弹簧（a）。携带α-S变异的患者 贫血，由于突变，循环网织红细胞中几乎没有mRNA 导致aCS mRNA不稳定。然而，分子机制 参与mRNA的特异性周转的研究知之甚少， 已经鉴定了参与mRNA降解的核酸酶。我们有 设计了一种体外mRNA衰变测定法， 周转的α-珠蛋白mRNA，并确定了一个序列特异性 核糖核酸内切酶，ErEN，其参与该mRNA的降解。Eren 是红细胞特异性核糖核酸内切酶，其特异性切割 体外和细胞中的a-球蛋白mRNA，从而证明这一点 核酸酶活性对于α-珠蛋白mRNA的正常生物合成是必需的。的 本建议的长期目标是了解 调节珠蛋白mRNA的稳定性和衰变。本提案的重点是：（目的 1)生物化学分离和分子克隆ErEN;（AIM 2）功能性 表征内切核糖核酸酶活性及其在 红细胞生成;（AIM 3）启动异源mRNA的靶向调节 和（AIM 4）鉴定由ErEN调节的另外的红系mRNAs。一 全面了解珠蛋白基因表达的所有分子控制 是有效改善血红蛋白病和人类遗传病所必需的 一般的混乱。这项工作将提供对mRNA的基本见解 周转，这是一个重要的，但相对未开发的组成部分，基因 表达，并提供新的方法来调节基因表达， 治疗策略

DESCRIPTION (provided by applicant): Globin gene expression is maintained and accentuated as a consequence of the unusual stability of the globin mRNAs during differentiation into a transcriptionally silent enucleated erythrocyte. Relatively little is known about the components that regulate globin mRNA stability. Irregularities in mRNA stability can have profound consequences that manifest as clinical phenotypes including thalassemias as exemplified by the athalassemia, a Constant Spring (a). Patients with the a S variant are severely anemic with virtually no mRNA in circulating reticulocytes due to a mutation that causes instability of the aCS mRNA. However, the molecular mechanisms involved in specific turnover of mRNAs are poorly understood and very few nucleases involved in the degradation of mRNA have been identified. We have devised an in vitro mRNA decay assay which recapitulates regulated mRNA turnover of the a-globin mRNA and have identified a sequence specific endoribonuclease, ErEN, which is involved in the degradation of this mRNA. ErEN is an erythroid specific endoribonuclease which specifically cleaves the a-globin mRNA both in vitro and in cells thereby demonstrating that this nuclease activity is essential for the normal biogenesis of a-globin mRNA. The long term objective of this proposal is to understand the determinants that regulate globin mRNA stability and decay. The focus of this proposal is: (AIM 1) to biochemically isolate and molecularly clone ErEN; (AIM 2) to functionally characterize the endoribonuclease activity and its expression profile in erythropoiesis; (AIM 3) to initiate targeted regulation of heterologous mRNA and (AIM 4) to identify additional erythroid mRNAs regulated by ErEN. A thorough understanding of all the molecular controls of globin gene expression are necessary to efficiently ameliorate hemoglobinopathies and human genetic disorders in general. This work will provide fundamental insights into mRNA turnover, which is an important yet relatively unexplored component of gene expression, and afford novel approaches to regulate gene expression in therapeutic strategies.