Brachyury-downstream gene networks in the notochord

描述(申请人提供)：转录因子brachyury(Bra)在脊索形成中的作用已有很好的文献记载，并在整个脊索门中进化保守。然而，关于Bra直接或间接控制的转录因子，以及与Bra协同作用的因子在脊索发育和进化中的作用，人们仍然知之甚少。特别是，关于脊索表达的顺式调节模块(增强子)的结构、共同特征和调控方式的知识是零散的和贫乏的。我们利用海鞘提供的实验优势作为模型系统，从已被描述为BRA靶标的基因以及与BRA关系未知的基因中分离和鉴定了一系列脊索增强子。我们从对脊索增强子的比较分析中收集的初步数据，使我们假设Ciona的脊索形成依赖于额外的、大多未描述的脊索转录因子，这些转录因子分为两类：BRA下游因子，由Bra直接或间接控制，作为其转录中介和与Bra协同作用的因子。我们将通过以下具体目标来验证这些假说：(1)分离脊索表达所需的最小调控序列；(2)识别控制最小增强子序列的激活子，并研究它们在脊索发育中的作用；(3)识别控制脊索激活子的增强子。 拟议的研究范围广泛，旨在快速解开脊索发育和分化不同步骤背后的基因网络，并获得有价值的见解，了解在脊索形成和进化过程中控制基因表达的分子机制。

DESCRIPTION (provided by applicant): The role of the transcription factor Brachyury (Bra) in notochord formation is well documented and evolutionarily conserved throughout the chordate phylum. However, still little is known about the transcription factors controlled directly or indirectly by Bra and about the function of factors acting in concert with Bra in notochord development and evolution. In particular, the knowledge of structure, common features and modalities of regulation of cis-regulatory modules (enhancers) directing expression in the notochord is fragmentary and poor. We have exploited the experimental advantages offered by the ascidian Ciona intestinalis as a model system to isolate and characterize a collection of notochord enhancers from genes that have already been described as Bra targets and from genes whose relationship with Bra is unknown. Our preliminary data, gathered from the comparative analysis of the notochord enhancers, lead us to hypothesize that notochord formation in Ciona relies upon additional, mostly uncharacterized notochord transcription factors that fall into two categories: Bra-downstream factors, controlled directly or indirectly by Bra and acting as its transcriptional intermediaries and factors acting in concert with Bra. We will test these hypotheses through the following specific aims: (1) Isolate minimal regulatory sequences necessary for notochord expression; (2) Identify the activators controlling the minimal enhancer sequences and study their role in notochord development; (3) Identify the enhancers controlling, in turn, selected notochord activators. The broad scope of the proposed research is to rapidly untangle the gene networks underlying the different steps of notochord development and differentiation and to gain valuable insights into the molecular mechanisms controlling gene expression in the notochord during its formation and evolution.