D-raf in Drosophila Signal Transduction

本项目的目的是确定在黑腹果蝇中酪氨酸激酶受体介导的信号通路中调节D-raf活性的机制，以建立和完善模式。使用结构/功能方法，我们已经定义了D-raf调控域，这是母体躯干通路中指定胚胎两极细胞命运的信号所必需的。我们的研究表明，末端信号的传递依赖于一个新的调控分支的活性，称为“抑制分支”，它(i)与躯干受体酪氨酸激酶平行作用；（ii）依赖于躯干和fs(1)极孔基因的母体活性；（iii）涉及丝氨酸残基388处D-raf的调控。本项目的具体目的如下：(1)通过在躯干和果蝇EGFR通路中表达D-raf结构域来确定调节机制的异同。(2)开始对影响终端信号通路“抑制分支”的突变进行饱和筛选。(3)利用含丝氨酸388的D-rafCR2模块（保守区2）完成双杂交筛选，鉴定与D-raf相互作用的“抑制分支”因子。由于D-raf是Raf-1原癌基因在果蝇中的同源基因，因此研究其在发育中的作用及其调控机制将为理解高等生物的信号转导过程提供线索。

The goal of this project is to define the mechanisms that regulate the activity of D-raf in receptor tyrosinekinase mediated signaling pathways for the establishment and elaboration of pattern in Drosophila melanogaster. Using a structure/function approach we have defined D-raf regulatory domains that are required for signaling in the maternal Torso pathway to specify cell fates at the embryonic poles. Our studies have revealed that transmission of the terminal signal depends on the activity of a novel regulatory branch, termed the "inhibitory branch", which (i) acts in parallel with the Torso receptor tyrosine kinase: (ii) is dependent on the maternal activities of the trunk and fs(1) pole hole genes; and (iii) involves the regulation of D-raf at serine residue 388. The specific aims of this project are as follows: (1) to define similarities and differences in mechanisms of regulation by expressing D-raf domains in Torso and Drosophila EGFR pathways. (2) to begin a saturation screen for mutations affecting the "inhibitory branch" of the terminal signaling pathway. (3) to complete a two-hybrid screen using the D-rafCR2 module (conserved region 2) containing serine 388 to identify "inhibitory branch" factor(s) that interact with D-raf. Since D-raf is the Drosophila homologue of the Raf-1 proto-oncogene, characterization of its role in development and identification of the mechanisms involved in its regulation will provide clues to understand the process of signal transduction in higher organisms.