Alternative Splicing of the Drosophila Dscam Pre-mRNA

果蝇 Dscam 前 mRNA 的选择性剪接

• 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
• 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; 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

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（由申请人提供）：该提案的长期目标是了解果蝇唐氏综合症细胞粘附分子（DSCAM）基因的替代剪接如何受到调节，并确定DSCAM替代剪接相互排斥的机制。 DSCAM基因编码轴突引导受体，该引导受体在神经发育中起重要作用，并且是迄今已知的最广泛的剪接基因。 DSCAM基因包含115个外显子，其中95个是剪接的。替代外显子分为4个不同的簇，其中包含12、48、33和2个互斥外显子。由于每个群集中的外显子都以相互排斥的方式剪接，因此可以表达38,016个不同的DSCAM同工型。已经提出，每个DSCAM同工型都可能与不同的轴突引导线索相互作用，并且通过细胞表达的DSCAM同工型的集合将直接参与引导神经元以不同地址引导神经元。因此，可能必须以某种方式编程单个神经元以特定方式将DSCAM Pre-MRNA拼写下来。因此，了解调节DSCAM替代剪接的机制将提供对指定神经接线的遗传程序的见解。该建议旨在理解调节DSCAM外显子4群集的替代剪接所涉及的机制，该集群包含12个相互排斥的外显子。首先，我们将确定参与调节DSCAM替代剪接的RNA序列以及与这些元素结合的蛋白质。其次，我们将确定外显子4.2替代剪接的发育调节所涉及的机制。第三，我们将确定SR蛋白B52和一般剪接因子DU2AF调节外显子4.4替代剪接的机制。最后，我们将确定DSCAM外显子4簇的替代剪接相互排斥的机械基础。总之，这些实验将对调节替代剪接的机制，负责相互排斥的替代剪接的机制以及决定果蝇中神经布线的特异性的遗传程序提供重大见解。