GENE REGULATION IN THE ASCIDIAN CIONA INTESTINALIS

拟议的研究计划代表着我们继续努力揭开复杂的基因调控网络，该网络负责指定和分化内胚层谱系在海鞘中。特别的努力将集中在两个这样的谱系：尾部肌肉和脊索。研究计划包括三个具体目标。首先，将使用生物信息学方法和实验分析相结合的方法来确定新的增强子，这些增强子在电穿孔Ciona胚胎中介导脊索特异或肌肉特异的基因表达。保守的序列基序将在协调调控的增强子中确定，并将努力确定与这些新确定的基序相互作用的特定转录因子。其次，将使用多种方法来确定在海胆中定义的优雅内胚层基因调控网络是否在Ciona中保守。我们的大部分工作将集中在两个关键的内胚层模式基因的顺式和反式调节上：Krox1和GataE。第三，新建立的GFP可视化方法将用于表征过表达内胚层模式基因(如Krox1、FoxA和brachyury)所产生的突变表型。这项拟议的研究计划的成功实施将首次对两种相关但形态截然不同的动物的明确定义的基因调控网络进行系统比较。

The proposed research plan represents a continuation of our efforts to unravel the complex gene regulation networks responsible for the specification and differentiation of endomesoderm lineages in the ascidian, Ciona intestinalis. Particular efforts will focus on two such lineages: the tail muscles and notochord. The research plan includes three specific aims. First, a combination of bioinformatics methods and experimental assays will be used to identify new enhancers that mediate either notochord-specific or muscle-specific gene expression in electroporated Ciona embryos. Conserved sequence motifs will be identified within coordinately regulated enhancers, and efforts will be made to identify the specific transcription factors that interact with these newly identified motifs. Second, a variety of methods will be used to determine whether the elegant endomesoderm gene regulation network defined in sea urchins is conserved in Ciona. Most of our efforts will focus on the cis- and trans- regulation of two key endomesoderm patterning genes: Krox1 and GataE. Third, newly established GFP visualization methods will be used to characterize the mutant phenotypes produced by overexpressing endomesoderm patterning genes such as Krox1, FoxA, and Brachyury. The successful execution of the proposed research plan will provide the first systematic comparision of a well-defined gene regulation network in two related, but morphologically distinct, animals.