Corbo J C, Levine M, Zeller R W
Department of Biology, University of California, San Diego, La Jolla 92093-0347, USA.
Development. 1997 Feb;124(3):589-602. doi: 10.1242/dev.124.3.589.
We present evidence that the embryo of the ascidian, Ciona intestinalis, is an easily manipulated system for investigating the establishment of basic chordate tissues and organs. Ciona has a small genome, and simple, well-defined embyronic lineages. Here, we examine the regulatory mechanisms underlying the differentiation of the notochord. Particular efforts center on the regulation of a notochord-specific Ciona Brachyury gene (Ci-Bra). An electroporation method was devised for the efficient incorporation of transgenic DNA into Ciona embryos. This method permitted the identification of a minimal, 434 bp enhancer from the Ci-Bra promoter region that mediates the notochord-restricted expression of both GFP and lacZ reporter genes. This enhancer contains a negative control region that excludes Ci-Bra expression from inappropriate embryonic lineages, including the trunk mesenchyme and tail muscles. Evidence is presented that the enhancer is activated by a regulatory element which is closely related to the recognition sequence of the Suppressor of Hairless transcription factor, thereby raising the possibility that the Notch signaling pathway plays a role in notochord differentiation. We discuss the implications of this analysis with regard to the evolutionary conservation of integrative enhancers, and the subdivision of the axial and paraxial mesoderm in vertebrates.
我们提供的证据表明，海鞘（Ciona intestinalis）的胚胎是研究基本脊索动物组织和器官形成的一个易于操作的系统。海鞘基因组较小，胚胎谱系简单且明确。在此，我们研究脊索形成过程中的调控机制。特别关注的是脊索特异性海鞘短尾基因（Ci-Bra）的调控。我们设计了一种电穿孔方法，用于将转基因DNA高效导入海鞘胚胎。该方法使得能够从Ci-Bra启动子区域鉴定出一个最小的434 bp增强子，该增强子介导绿色荧光蛋白（GFP）和乳糖操纵子（lacZ）报告基因在脊索中的特异性表达。该增强子包含一个负调控区域，可排除Ci-Bra在不适当胚胎谱系中的表达，包括躯干间充质和尾部肌肉。有证据表明，该增强子由一个与无毛转录因子抑制子识别序列密切相关的调控元件激活，从而增加了Notch信号通路在脊索分化中发挥作用的可能性。我们讨论了这一分析对于整合增强子的进化保守性以及脊椎动物中轴和近轴中胚层细分的意义。
