AXON GIUDANCE AND TARGETING IN DROSOPHILA

描述(申请人提供)：DSCAM是一种轴突引导受体， 果蝇神经中神经元连接的形成所必需的 系统。DSCAM功能丧失的表型已经被详细地表征为 一种胚胎神经，称为博尔维希神经(BN)。DSCAM的表型研究 突变体提示Dscam蛋白在调节短程 BN与中间目标之间的相互作用。DSCAM也是必需的 发育中胚胎中其他连接的规范。DSCAM蛋白 包含10个Ig重复序列，6个纤维连接蛋白III型重复序列，单个跨膜 Dock适配器的片段和具有多个结合位点的细胞质尾巴 连接Dscam和肌动蛋白细胞骨架的信号元件--蛋白质 监管者。DSCAM的替代剪接可能导致超过38,000种形式的 感受器。这些受体具有相同的结构域结构，但在 4个不同结构域的氨基酸序列。三个免疫球蛋白结构域和 跨膜结构域是可变的。这种可变性反映了 另一种外显子。 在本提案中，将确定以BN表示的DSCAM的具体形式 使用RT-PCR法。转基因救援实验将被用来验证这一点 在国阵中，形式是起作用的。含有不同可变区的转基因将 通过在BN中进行靶向表达来测试其功能。额外的实验 将针对确定DSCAM的表达模式 发育中胚胎中的异构体和发育中的眼表象盘 原位杂交和RT-PCR技术。这些研究将形成 确定不同的DSCAM亚型是否有助于 轴突引导和靶向的分子代码。像果蝇Dscam一样 这些研究在结构上与唐氏综合症中涉及的人类基因有关 这项资助中所描述的可能提供了对大脑异常的洞察力 唐氏综合征患者的发展状况。

DESCRIPTION (provided by applicant): Dscam is an axon guidance receptor that is required for the formation of neuronal connections in the Drosophila nervous system. Dscam loss-of-function phenotypes have been characterized in detail for an embryonic nerve called Bolwig's nerve (BN). Phenotypic studies of Dscam mutants suggest that Dscam protein is critical for mediating a short-range interaction between BN and an intermediate target. Dscam also is required for the specification of other connections in the developing embryo. Dscam protein contains 10 Ig repeats, 6 fibronectin type III repeats, a single transmembrane segment and a cytoplasmic tail with multiple binding sites for the Dock adapter protein, a signaling component connecting Dscam to actin cytoskeletal regulators. Alternative splicing of Dscam may lead to more than 38,000 forms of the receptor. These receptors share the same domain structure, but differ in amino acid sequence in 4 different domains. Three of the Ig domains and the transmembrane domain are variable. This variability reflects the use of alternative exons. In this proposal, the specific form of Dscam expressed in BN will be determined using RT-PCR. Transgene rescue experiments will be used to verify that this form is functional in BN. Transgenes containing different variable domains will be tested for function by targeted expression in BN. Additional experiments will be directed towards determining the pattern of expression of Dscam isoforms in the developing embryo and the developing eye imaginal disc using both in situ hybridization and RT-PCR techniques. These studies will form the foundation for determining whether different Dscam isoforms contribute to a molecular code for axon guidance and targeting. As Drosophila Dscam is structurally related to a human gene implicated in Down Syndrome, the studies described in this grant may provide insights into the abnormalities in brain development in Down Syndrome patients.