Alternative Splicing of the Drosophila Dscam Pre-mRNA

果蝇 Dscam 前 mRNA 的选择性剪接

基本信息

批准号：
6878632
负责人：
Brenton R. Graveley
金额：
$ 27.55万
依托单位：
UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
依托单位国家：
美国
项目类别：
财政年份：
2003
资助国家：
美国
起止时间：
2003-05-01 至 2007-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6878632
关键词：
Drosophilidae RNA binding protein RNA splicing SDS polyacrylamide gel electrophoresis arthropod genetics autoradiography cell line confocal scanning microscopy genetic regulation neural cell adhesion molecules neuroanatomy neuronal guidance nucleic acid sequence polymerase chain reaction precursor mRNA protein binding protein structure function transfection

项目摘要

DESCRIPTION (provided by applicant): The long-term goals of this proposal are to understand how the alternative splicing of the Drosophila Down syndrome cell adhesion molecule (Dscam) gene is regulated and to determine the mechanism by which Dscam alternative splicing is mutually exclusive. The Dscam gene encodes an axon guidance receptor that plays an important role in neural development and is the most extensively alternatively spliced gene known to date. The Dscam gene contains 115 exons, 95 of which are alternatively spliced. The alternative exons are organized into 4 distinct clusters containing 12, 48, 33 and 2 mutually exclusive exons each. Because the exons within each cluster are alternatively spliced in a mutually exclusive manner, it is possible that 38,016 different Dscam isoforms can be expressed. It has been proposed that each Dscam isoform might interact with a different set of axon guidance cues and that the collection of Dscam isoforms expressed by a cell will be directly involved in guiding neurons to different addresses. It is therefore likely that individual neurons must in some way be programmed to splice the Dscam pre-mRNA in specific ways. Thus understanding the mechanisms regulating Dscam alternative splicing will provide insight into the genetic program that specifies neural wiring. This proposal is aimed at understanding the mechanisms involved in regulating the alternative splicing of the Dscam exon 4 cluster which contains 12 mutually exclusive exons. First, we will identify RNA sequences involved in the regulation of Dscam alternative splicing and the proteins that bind to these elements. Second, we will determine the mechanism involved in the developmental regulation of exon 4.2 alternative splicing. Third, we will determine the mechanism by which the SR protein B52 and the general splicing factor dU2AF modulate exon 4.4 alternative splicing. Finally, we will determine the mechanistic basis by which alternative splicing of the Dscam exon 4 cluster is mutually exclusive. Together, these experiments will provide significant insight into the mechanisms involved in regulating alternative splicing, the mechanism responsible for mutually exclusive alternative splicing, and the genetic program that determines the specificity of neural wiring in Drosophila.

描述（申请人提供）：本提案的长期目标是了解果蝇唐氏综合症细胞粘附分子（Dscam）基因的选择性剪接是如何调控的，并确定Dscam选择性剪接相互排斥的机制。 Dscam 基因编码轴突引导受体，在神经发育中发挥重要作用，是迄今为止已知最广泛的选择性剪接基因。 Dscam基因包含115个外显子，其中95个是选择性剪接的。替代外显子被组织成 4 个不同的簇，每个簇包含 12、48、33 和 2 个互斥的外显子。由于每个簇内的外显子以互斥方式交替剪接，因此可能可以表达 38,016 种不同的 Dscam 同工型。有人提出，每个 Dscam 同工型可能与一组不同的轴突引导线索相互作用，并且细胞表达的 Dscam 同工型集合将直接参与引导神经元到不同的地址。因此，单个神经元很可能必须以某种方式进行编程，以特定的方式剪接 Dscam 前 mRNA。因此，了解调节 Dscam 选择性剪接的机制将有助于深入了解指定神经线路的遗传程序。该提案旨在了解 Dscam 外显子 4 簇（包含 12 个互斥外显子）的选择性剪接调节机制。首先，我们将鉴定参与 Dscam 选择性剪接调节的 RNA 序列以及与这些元件结合的蛋白质。其次，我们将确定外显子 4.2 选择性剪接发育调控所涉及的机制。第三，我们将确定SR蛋白B52和通用剪接因子dU2AF调节外显子4.4选择性剪接的机制。最后，我们将确定 Dscam 外显子 4 簇的选择性剪接互斥的机制基础。总之，这些实验将提供对参与调节选择性剪接的机制、负责相互排斥的选择性剪接的机制以及决定果蝇神经线路特异性的遗传程序的重要见解。