FUNCTIONAL ANALYSIS OF GENES INVOLVED IN VERTEBRATE CRANIOFACIAL DEVELOPMENT

描述(申请人提供)：约75%的出生缺陷涉及头部、面部和口腔组织。虽然口面部裂和其他颅面畸形有明显的环境和遗传原因，但关于颅面发育机制的信息不足，使这些缺陷中的大多数能够在出生前被发现或预防。我们的目标是开发颅面畸形的动物模型，这将导致对相关人类出生缺陷的诊断和治疗的机械性洞察。我们最近发现了一大组新的基因，这些基因在老鼠面部形成的关键阶段表达。过去25年来积累的大量数据表明，不同物种的同源基因在相似的发育过程中往往具有保守的功能。考虑到这些观察，我们打算测试我们的假设，即在小鼠头面部隆起中差异表达的基因，以及在斑马鱼中在结构和表达上保守的基因，在脊椎动物面部的发育中发挥重要的保守功能，这将与人类头面部出生缺陷相关，包括口面部裂开和小颌症。斑马鱼提供了一个非常灵活的系统来快速有效地筛选多个候选基因的功能重要性。斑马鱼的发育是在外部进行的，可以在显微镜下连续检查。此外，斑马鱼胚胎中的基因表达可以被称为Morpholinos的特定反义试剂有效和容易地靶向。我们将执行两个相互关联的特定目标。目的1.分离斑马鱼在小鼠胚胎中发现的新的、特异的口面部转录本，并分析其在发育过程中的表达谱。目的2.基于钼的斑马鱼基因敲除筛选，分析其在颅面发育中的基因功能。最终，这项研究的结果将为分析相关人类出生缺陷的根本原因提供新的候选者。

DESCRIPTION (provided by applicant): About 75% of birth defects involve the head, face, and oral tissues. Although orofacial clefts and other craniofacial malformations have clear environmental and genetic causes, insufficient information exists concerning the mechanisms of craniofacial development to enable the majority of these defects to be detected or prevented pre-natally. Our goal is to develop animal models of craniofacial malformations that will lead to mechanistic insight into the diagnosis and treatment of related human birth defects. We have recently identified a large set of novel genes that are expressed during the critical stages of mouse face formation. Considerable data has accumulated over the past 25 years that orthologous genes in diverse species often share conserved functions in similar developmental processes. With these observations in mind, we intend to test our hypothesis that genes which are differentially expressed in the mouse craniofacial prominences, and which are conserved in structure and expression in the zebrafish, perform important conserved functions in the development of the vertebrate face that will be relevant to human craniofacial birth defects, including orofacial clefting and micrognathia. The zebrafish provides a very amenable system to rapidly and efficiently screen the functional importance of multiple gene candidates. Zebrafish development occurs externally and can be examined continuously under a microscope. Moreover, gene expression in the zebrafish embryo can be efficiently and easily targeted with specific anti-sense reagents, termed Morpholinos. We will perform two interrelated to Specific Aims. Aim 1. Isolation of zebrafish cDNAs corresponding to the novel and specific orofacial transcripts identified in the embryonic mouse and analysis of their expression profiles during development. Aim 2. MO based knockdown screen in zebrafish to analyze gene function in craniofacial development. Ultimately the results of this study will provide new candidates for an analysis of the underlying causes of related human birth defects.