Protein/Nucleic Acid Interaction & Vertebrate Embryology

在脊椎动物中，在原肠胚形成过程中，外胚层被细分为四个主要部分：表皮和中枢神经系统（CNS）分别位于腹面和背面，神经嵴和感觉基板位于这两个区域之间。该项目的目标是了解决定外胚层细胞如何在这些途径之间选择的机制，以及随后的组织模式和分化是如何被调节的。本实验室采用的实验方法是使用依赖于细胞-细胞信号作为鉴定可能调节外胚层发育程序的基因的标准。人们的注意力集中在Msx1、Dlx3、Dlx5和Dlx6四个同型盒基因以及转录激活因子AP-2alpha上。我们发现这四个同源盒基因受骨形态发生蛋白（BMP）信号的分级反应的差异调控，从而导致体内不同的表达边界。我们推测，这至少可以解释原肠胚形成过程中外胚层空间格局的一些主要特征。Msx1似乎参与骨水泥腺（一种划分神经板最前方的结构）前边界的建立，并参与神经嵴的诱导。Dlx3似乎起着建立神经嵴外侧边界的作用。Dlx5和Dlx6的作用可能与神经板边界的建立有关，也可能与感觉基板的形成有关。

In vertebrates, the ectoderm is subdivided during gastrulation into four primary fates: epidermis and central nervous system (CNS) on the ventral and dorsal surfaces, respectively, and neural crest and sensory placodes located between these two domains. The goal of this project is to understand the mechanisms that determine how ectodermal cells choose between these pathways, and how the ensuing tissue patterning and differentiation are regulated. The experimental approach taken by this laboratory has been to use dependence upon cell-cell signaling as a criterion for identifying genes that may regulate the ectodermal developmental programs. Attention has been focused on four homeobox genes, Msx1, Dlx3, Dlx5 and Dlx6, and on the transcriptional activator AP-2alpha. We have found that the four homeobox genes are differentially regulated by a graded response to bone morphogenetic protein (BMP) signaling, resulting in different expression boundaries in vivo. We theorize that this can account for at least some of the major features of the spatial patterning of ectoderm during gastrulation. Msx1 appears to be involved in establishing the anterior boundary of the cement gland, a structure demarcating the most forward aspect of the neural plate, and in the induction of neural crest. Dlx3 seems to function in setting up the lateral boundary of neural crest. The roles of Dlx5 and Dlx6 may be related to establishing the boundary of the neural plate, and also in the formation of the sensory placodes. Neural crest induction in Xenopus requires two signals, a partially attenuated BMP signal and a Wnt/beta catenin signal. We have found that AP2alpha is responsible for conveying the BMP signal to the genome, and activation of a broad spectrum of neural crest-specific genes. AP2alpha is also imporant in the development of epidermis. We are using a hormone-inducible version of AP2alpha as a tool in conjunction with microarray analysis to identifiy target genes in both epidermis and neural crest. Several hundred such genes have now been identified, many of which have known functions such as transcription factors, signaling molecules, extracellular matrix-related enzymes and other structural or enzymatic functions. Many also have no known function, but are conserved in the mouse and human genomes and are thus likely to be important. We are now using antisense oligonucleotides and other strategies to study these functions, with the goal of learning how genes interact to control the formation and differentiation of epidermal and neural crest cells in the frog embryo. Since these tissues and genes are conserved throughout vertebrate phylogeny, our findings should have broad relevance to biomedical research and human health and development.