MOLECULAR ANALYSIS OF THE BOSS TRANSMEMBRANE LIGAND

细胞间通讯在细胞分化过程中起着关键作用 流程. 果蝇R7感光细胞的诱导 复眼由邻近的R8细胞构成，已成为经典之一 两个细胞之间的感应相互作用的示例。 R7被诱导 通过sevenless（sev）受体酪氨酸激酶 在R7前体细胞和七无新娘（老板）的表面上， 在相邻的R8细胞表面表达。 长期目标 这项研究的目的是了解R7中的生化步骤， 诱导途径 该建议集中于诱导配体凸台的分析。 boss蛋白由一个大的胞外结构域（498个氨基酸）组成， 酸），具有七个跨膜区段的跨膜区（7 TM 该区域是激活sevenless和诱导R7所必需的 细胞命运 大细胞外进入邻近的R7细胞作为一个 这是它与SEV受体相互作用的结果。 这些发现 提出了一些重要的问题：老板是如何作为一个配体， SEV受体？ 7 TM域是否构成约束的一部分 SEV受体的位置或者，7 TM区域是否需要 与第二种蛋白相互作用形成同源二聚体或异源二聚体？什么 是将boss转移到R7的机制？ 这些问题将通过以下具体目标加以解决： 1. 定义凸台7 TM区域在激活sev受体中的作用 酪氨酸激酶 嵌合蛋白将用于定义最小的 R7诱导所需的7 TM区域的一部分，并推断其 功能意义。 2. 对boss蛋白进行纯化和功能重组。 如果 重组的boss蛋白不足以激活SEV 受体酪氨酸激酶，我们将确定boss的中间步骤 内化 许多（如果不是全部）信号关键成分的突变 诱导R7发育的转导途径将很快可用。 这些突变提供了一个完美的分析环境， 通路 生物化学和遗传学方法的结合， 培养系统和转基因动物将使我们能够深入了解 这条路。 这些知识将为 了解许多信号通路调节这些不同的 过程如神经元分化、生长控制和瘤形成。 一 在特定蛋白质-蛋白质水平上的信号转导知识 相互作用将是朝着 合理设计调节这些过程的药物。

Cell-cell communication plays a key role in may cellular differentiation processes. Induction of the R7 photoreceptor cell in the Drosophila compound eye by the neighboring R8 cell has become one of the classic examples for an inductive interaction between two cells. R7 is induced by an interaction between the sevenless (sev) receptor tyrosine kinase on the surface of the R7 precursor cell and bride of sevenless (boss), expressed on the surface of the neighboring R8 cell. The long term goal of this research is to understand the biochemical steps in the R7 inductive pathway. This proposal concentrates on an analysis of the inducing ligand boss. The boss protein is composed of a large extracellular domain (498 amino acids), a transmembrane region with seven membrane spanning segments (7TM region is necessary for the activation of sevenless and induction of R7 cell fate. The large extracellular into the neighboring R7 cell as a consequence of its interaction with the sev receptor. These findings raise a number of important questions: How does boss act as a ligand for the sev receptor? Does the 7TM domain constitute part of the binding site for the sev receptor? Alternatively, is the 7TM region necessary to interact with a second protein to form a homodimer or heterodimer? What is the mechanism of transfer of boss into R7? These questions will be addressed by the following specific aims: 1. Define the role of the boss 7TM region in activating the sev receptor tyrosine kinase. Chimeric proteins will be used to define the minimal part of the 7TM region required for R7 induction and to infer its functional significance. 2. Purify and functionally reconstitute the boss protein. If reconstituted boss protein is not sufficient for activation of the sev receptor tyrosine kinase we will identify intermediate steps of boss internalization. Mutations in many -if not all- of the critical components of the signal transduction pathway inducing R7 development will soon be available. These mutations provide a perfect setting for an analysis of this pathway. A combination of biochemical and genetic methods using cell culture systems and transgenic animals will allow an intimate knowledge of this pathway. This knowledge will lay the foundation for understanding numerous signaling pathways regulating such diverse processes as neuronal differentiation, growth control and neoplasia. A knowledge of signal transduction at the level of specific protein-protein interactions will be an invaluable step forward in the direction of the rational design of drugs regulating these processes.