GENETIC ANALYSIS OF RNA BINDING PROTEINS

描述(摘自申请者的摘要)：对RNA结合的兴趣 蛋白质(RNA-Bps)是近年来随着人们对蛋白质认识的不断提高而迅速发展起来的 这些蛋白质在各种各样的细胞过程中起着关键作用。 然而，人们对它们的结构特征知之甚少 使他们能够识别他们的RNA靶标的蛋白质 专一性。这些调查将侧重于确定和 描述了RNA-BPS的一些重要特性，这些特性对其 能够绑定到特定的RNA。我们将特别关注 代表三个不同RNA-Bps家族的蛋白质范例： 哺乳动物剪接体蛋白U1a和U2b“(RRM家族)，HIV-1蛋白 REV(富含精氨酸的家族)和酵母调节蛋白Mer1(KH 家庭)。实验策略将强调基因 随机突变的选择/筛选以识别重要的RNA-BP 残留物，然后进行定点突变和生化分析 更好地了解这些残基在RNA结合中的作用。结果是 这些研究将为三个成员提供重要的见解 不同的RNA-BP家族识别它们的RNA靶标，从而脱落 对分子生物学的一个关键方面的了解，目前这方面还很差 明白了。这些信息将有助于理解的目标 一般情况下，RNA-Bps是如何以高亲和力和 专一性。鉴于RNA结合蛋白在 HIV-1感染中基因表达的转录后控制，以及 在脆性X精神发育迟滞患者中，这一知识应该对 了解发病机制，并最终可能形成基础 用于新的基因疗法。

DESCRIPTION (Adapted from applicant's abstract): Interest in RNA-binding proteins (RNA-BPs) has burgeoned in recent years with the growing awareness that these proteins play a key role in a wide variety of cellular processes. However, very little is known about the structural features of these proteins that enable them to recognize their RNA targets with such high specificity. These investigations will focus on identifying and characterizing some of the features of RNA-BPs that are important for their ability to bind to specific RNAs. Particular attention will be devoted to protein paradigms representing three different families of RNA-BPs: the mammalian spliceosomal proteins U1A and U2B" (RRM family), the HIV-1 protein Rev (arginine-rich family), and the yeast regulatory protein Mer1 (KH family). The experimental strategy will emphasize genetic selection/screening of random mutations to identify important RNA-BP residues, followed by site-directed mutagenesis and biochemical analysis to better understand the role of these residues in RNA binding. The results of these studies will provide important insights into how members of three different RNA-BP families recognize their RNA targets, thereby shedding light on a key aspect of molecular biology that presently is poorly understood. This information will contribute to the goal of understanding how RNA-BPs in general are able to bind their targets with high affinity and specificity. In view of the central role that RNA-binding proteins play in the post-transcriptional control of gene expression, in HIV-1 infection, and in fragile X mental retardation, this knowledge should be of value for understanding mechanisms of pathogenesis and could eventually form the basis for novel gene therapies.