MUTAGENESIS SCREEN FOR MUTATIONS AFFECTING ZEBRAFISH

神经嵴是脊椎动物胚胎前体细胞的一个短暂群体，它产生各种各样的细胞类型，包括周围神经元和胶质细胞、内分泌细胞、色素细胞和颅面骨骼的元素。由于神经嵴发育几乎涉及胚胎发育的所有基本过程，神经嵴已被广泛用作研究胚胎细胞分化的模型系统。此外，大量人类临床相关疾病是由于神经嵴发育过程中的错误导致的，其严重程度从功能不便到灾难。最后，因为神经嵴是严格意义上的脊椎动物胚胎组织，理解神经嵴的发育具有重要的革命性意义。综上所述，阐明神经嵴发育是如何被调控的，对于理解各种功能重要细胞类型的产生、提供对胚胎发育的一般见解、理解并最终减轻涉及神经嵴和神经嵴衍生细胞的许多不利的人类疾病是非常重要的。我们建议系统筛选破坏神经嵴发育的斑马鱼突变，作为最终确定控制神经嵴发育的分子机制的一种手段。我们将通过监测作为神经嵴细胞标记的冠蛋白基因表达来识别突变，重点关注胚胎发生的四个不同阶段，这些阶段对应于神经嵴发育的主要事件，首先通过原位杂交，最终使用诱变的冠蛋白gfp转基因系。所有确定的突变系将通过分析大量神经嵴亚谱系和衍生物特异性分子探针的表达来表征。突变体将在这些结果的基础上进行分类，并进一步通过互补分析和连锁图谱进行表征。大量破坏神经嵴发育的突变的分离和初步表征将为研究界提供重要的资源，并对阐明调节神经嵴发育的机制具有宝贵的价值。

The neural crest is a transient population of vertebrate embryonic precursor cells that generates a wide variety of cell types of peripheral neurons and glia, endocrine cells, pigment cells and elements of the craniofacial skeleton. Because neural crest development involves nearly all of the basic processes common to embryonic development generally, the neural crest has been extensively utilized as a model system to study embryonic cell diversification. In addition, a large number of clinically relevant conditions in humans result from miscues during neural crest development that can range in severity from functional inconvenience to catastrophe. Lastly, because neural crest is strictly a vertebrate embryonic tissue, understanding neural crest development has important revolutionary implications. Taken together, elucidating how neural crest development is regulated is important in order to understand the generation of a variety of functionally significant cell types, to provide insights into embryonic development generally, and to understand and ultimately relieve a number of adverse human afflictions that involve the neural crest and neural crest- derived cells. We propose to systematically screen for zebrafish mutations that disrupt neural crest development as a means to ultimately identify the molecular mechanisms that control neural crest development. We will identify mutations by monitoring crestin gene expression as a marker for neural crest cells, focusing on four different stages of embryogenesis that correspond to major events in neural crest development, first by in situ hybridization and ultimately using a mutagenized crestin-GFP transgenic line. All identified mutant lines will then be characterized by analyzing the expression of a large battery of neural crest sublineage- and derivative- specific molecular probes. Mutants will be classified on the basis of these results and further characterized by complementation analysis and linkage mapping. The isolation and initial characterization of large numbers of mutations that disrupt neural crest development will provide an important resource for the research community and be invaluable for elucidating mechanisms that regulate neural crest development.