BIOCHEMISTRY OF REGULATED PRE-MRNA SPLICING/DROSOPHILA

选择性的前mRNA剪接是调节后生动物基因表达的一种常见机制。 的确，RNA处理是基因组信息传递到蛋白质组信息的管道。 事实上，现在人们已经认识到，已知的人类和老鼠疾病基因突变中有40%-50%会影响 拼接过程。因此，了解内含子是如何识别的，以及选择性剪接模式是如何 可以允许治疗性干预。这项拨款建议的研究的总体目标是 阐明调控信使RNA前体剪接的生化机制 果蝇在发育过程中，黑腹果蝇。我们的努力将集中在理解 详细介绍了两个选择性剪接的例子，以及更一般的RNA结合蛋白在其中所起的作用 控制拼接模式。为了实现我们的总体目标，我们将： 1.提纯和分析含有果蝇hnRNP蛋白、PSI、hrp48、hrp40、 Hrp38和hrp36。 2.用生化方法研究P元件与Squid/hrp40前-mRNAs的剪接规律 学习。 3.分析去除特定基因后全基因组范围内对不同的前-mRNA剪接模式的影响 利用果蝇剪接连接微阵列的RNA结合剪接因子。 4.分析PSI、果蝇U2AF和U2AF相关蛋白的结构和功能。

Alternative pre-mRNA splicing is a common mechanism for regulating gene expression in metazoans. Indeed, RNA processing is a conduit by which genomic information is transferred to proteomic information. In fact, it is now recognized that 40-50% of the known human and mouse disease gene mutations affect the splicing process. Thus, understanding how introns are recognized and how patterns of alternative splicing are set up may allow therapeutic intervention. The overall objective of the research proposed in this grant is to elucidate biochemical mechanisms used to regulate splicing of messenger RNA precursors (pre-mRNA) during development in the fruit fly,Drosophila melanogaster. Our efforts will be focused on understanding two examples of alternative splicing in detail and, more generally, the role that RNA binding proteins play in controlling splicing patterns. In order to accomplish our overall objectives, we will: 1. Purify and analyze nuclear pre-mRNPs that contain the Drosophila hnRNP proteins, PSI, hrp48, hrp40, hrp38 and hrp36. 2. Investigate the splicing regulation of the P element and squid/hrp40 pre-mRNAs using biochemical studies. 3. Analyze the genome-wide effects on alternative pre-mRNA splicing patterns after removal of specific RNA binding splicing factors using Drosophila splice junction microarrays. 4. Analyze the structure and function of PSI, Drosophila U2AF and U2AF-related proteins.