RNA-Protein interactions regulating mRNA stability

描述(申请人提供)：珠蛋白基因表达保持不变 由于珠蛋白mRNAs的不同寻常的稳定性而加重 在分化为转录沉默的去核红细胞过程中。 对调节珠蛋白mRNA的成分知之甚少 稳定性。MRNA稳定性的不规则性可能会产生深远的后果 表现为临床表型，包括地中海贫血，例如 体力衰竭，一个不变的春天(A)。携带a型S变种的患者严重 由于突变导致的循环网织红细胞中几乎没有mRNA的贫血 这导致了ACSm RNA的不稳定。然而，分子机制 参与特定的mRNAs周转的人很少，而且很少 参与核糖核酸降解的核酸酶已经被确定。我们有 设计了一种重述受调控的mRNA的体外信使核糖核酸衰退试验 A-珠蛋白mRNA的周转，并确定了一个特定的序列 内切核糖核酸酶，EREN，它参与了该mRNA的降解。二连 是一种红系特异性内切核糖核酸酶，它特异性地切割 在体外和细胞内的A-珠蛋白mRNA，从而证明了这一点 核酸酶活性对于α-珠蛋白mRNA的正常生物发生是必不可少的。这个 这项建议的长期目标是了解决定因素 调节珠蛋白信使核糖核酸的稳定和衰退。这项建议的重点是：(AIM 1)生化分离和分子克隆Eren；(AIM 2)在功能上 内切核糖核酸酶活性及其表达谱的研究 (AIM 3)启动对异源mRNA的靶向调控 和(目的4)鉴定受Eren调控的额外的红系mRNAs。一个 深入了解珠蛋白基因表达的所有分子调控 是有效改善血红蛋白疾病和人类遗传疾病所必需的 一般情况下都会有障碍。这项工作将提供对mrna的基本见解。 周转，这是基因中一个重要但相对未被探索的组成部分 表达，并提供了调控基因表达的新方法。 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.