PHOSPHORYLATION IN A DEVELOPMENTAL SIGNALING PATHWAY

这项研究的长期目标是了解一个信号 转导途径，调节基因表达的程序， 控制调节蛋白的亚细胞定位。 在 果蝇，细胞内信号转导收到 在胚胎表面促进细胞核转位， 转录因子Dorsal，从而建立了背腹侧 轴线 在苍蝇、植物和哺乳动物中，同源途径直接将核 相关转录因子导入作为免疫或 防御性反应。 除了Dorsal，果蝇背腹信号通路 需要跨膜受体Toll，支架蛋白Tube， 蛋白激酶Pelle和抑制剂Cactus。事实上这些 五个途径组分已在遗传和 分子水平，并可以通过胚胎显微注射进行检测， 特别适合于实验研究的系统。 现在 因此，有可能解决有关的基本问题， 信号转导机制。 本提案的前两个具体目标将侧重于 启动细胞内信号传导。 的势函数 肌动蛋白结合蛋白Filamin作为Toll和Tube之间的联系， 在免疫共沉淀和遗传相互作用研究中进行了研究。 另外的实验将讨论的机制和意义， 响应Toll激活的管重新定位。 第三和 第四个目标将解决信号如何传播的问题。 自磷酸化将被探索为Pelle的潜在机制 体内调节。 分子和生物化学研究也将在 进行以确定哪些途径蛋白经历Pelle介导的 胚胎中的磷酸化。 第五个也是最后一个具体目标将侧重于 关于合胞体环境中信号的空间调节 胚盘胚胎 考虑到信号传导途径的保守性质， 建议的研究不仅应具有实质性意义， 昆虫胚胎中的模式形成， 在广泛的植物和动物中的转导和免疫。

The long term goal of this research is to understand a signal transduction pathway that regulates programs of gene expression by controlling the subcellular localization of regulatory proteins. In the fruit fly Drosophila, intracellular transduction of a signal received at the surface of the embryo promotes nuclear translocation of the transcription factor Dorsal and thereby establishes the dorsoventral axis. In flies, plants, and mammals, homologous pathways direct nuclear import of related transcription factors as a critical step in immune or defensive responses. In addition to Dorsal, the Drosophila dorsoventral signaling pathway requires the transmembrane receptor Toll, the scaffolding protein Tube, the protein kinase Pelle, and the inhibitor Cactus. The fact that these five pathway components have been characterized at both the genetic and molecular level and can be assayed by embryo microinjection makes this system particularly amenable to experimental investigation. It is now possible, therefore, to address fundamental questions about the mechanism for signal transduction. The first two specific aims of this proposal will focus on the initiation of intracellular signaling. The potential function of the actin-binding protein Filamin as a link between Toll and Tube will be investigated in coimmunoprecipitation and genetic interaction studies. Additional experiments will address the mechanism and significance of Tube relocalization in response to Toll activation. The third and fourth aims will address how the signal is propagated. Autophosphorylation will be explored as a potential mechanism for Pelle regulation in vivo. Molecular and biochemical studies will also be carried out to determine which pathway proteins undergo Pelle-mediated phosphorylation in embryos. The fifth and final specific aim will focus on the spatial regulation of signaling within the syncytial environment of the blastoderm embryo. Given the conserved nature of the signaling pathway, the results of the proposed research should be of substantial interest with regard not only to pattern formation in the insect embryo, but also to signal transduction and immunity in a broad range of plants and animals.