Dorsoventral Patterning Through Transcriptional Control

描述(由申请人提供)：拟议的研究旨在了解在动物发育过程中，单个形态因子如何启动导致轴规范的复杂事件层次。研究中的形态原，DULSAL，是一种转录因子，在果蝇胚泡胚中以背腹浓度梯度分布。该因子既是转录的激活因子，又是转录的抑制因子，可以沿着背腹轴建立多个基因活性区域。背部的不同功能是由背部Rel同源结构域(RHD)、背部CTr终端域(CTD)和许多额外的调节因子之间的相互作用网络介导的。一种背部相互作用的蛋白质对于背部介导的抑制是必不可少的，它是辅阻遏物Groucho。这一建议的第一个具体目的是通过表征最近在背部系统突变中发现的两组突变来阐明背部介导的激活机制。这两类突变在RHD上定义了两个不同的表面。亲和层析将被用来鉴定新的共激活子，它们介导由II类突变定义的表面的功能。遗传和生化方法都将被用来检验这一假设，即由I类突变定义的表面调节CTD的活性。第二个具体目标是确定格罗乔镇压的机制。以前的研究已经得出了Groucho介导的远程抑制的假说，涉及Groucho通过背部招募Groucho，随后Groucho招募组蛋白脱乙酰酶，最后Groucho沿着去乙酰化模板扩散。遗传分析、结构研究和染色质免疫沉淀实验将结合起来严格检验这一假设。 调节果蝇背腹模式的途径与调节脊椎动物免疫反应和脊椎动物模式形成的途径是同源的。因此，这些研究可能会导致对人类免疫和发育障碍的更好理解。

DESCRIPTION (provided by applicant): The proposed studies are aimed at understanding how a single morphogen initiates the complex hierarchy of events leading to axis specification during animal development. The morphogen under investigation, Dorsal, is a transcription factor that is distributed in a dorsoventral concentration gradient in the Drosophila blastoderm embryo. This factor functions as both an activator and a repressor of transcription to establish multiple domains of gene activity along the dorsoventral axis. The diverse functions of Dorsal are mediated by a network of interactions between the Dorsal rel homology domain (RHD), the Dorsal Cterminal domain (CTD), and many additional regulatory factors. One Dorsal-interacting protein that is essential for Dorsal-mediated repression is the corepressor Groucho. The first specific aim of this proposal is to illuminate the mechanism of Dorsal-mediated activation by characterizing two sets of mutations discovered in a recent systematic mutagensis of Dorsal. These two classes of mutations define two distinct surfaces on the RHD. Affinity chromatography will be employed to identify novel coactivators that mediate the function of the surface defined by the class II mutations. Both genetic and biochemical approaches will be employed to test the hypothesis that the surface defined by the class I mutations modulates the activity of the CTD. The second specific aim is to determine the mechanism of repression by Groucho. Previous studies have led to a hypothesis for Groucho-mediated long-range repression that involves the recruitment of Groucho by Dorsal, the subsequent recruitment of a histone deacetylase by Groucho, and finally the spreading of Groucho along the deacetylated template. Genetic analysis, structural studies, and chromatin immunoprecipitation experiments will be combined to test this hypothesis rigorously. The pathway regulating dorsoventral patterning in Drosophila is homologous to pathways regulating the vertebrate immune response and vertebrate pattern formation. Thus, these studies may lead to a better understanding of human immunological and developmental disorders.