ORGANOGENESIS OF THE PHARYNX IN C ELEGANS

驱动前肠发育的基因和机制主要是 这是未知的，代表了发育生物学的一个新领域。 我们 研究前肠的发展，通过调查咽的形成， 线虫线虫先前，我们确定了一个位点， PHA-4，其是建立咽和直肠原基所需的： 携带PHA-4突变的动物在缺乏咽部的幼虫期停止生长， 直肠 我们最近克隆了PHA-4，并表明它编码一种 果蝇基因分叉头的同源物。 这一结果，加上我们的 表型分析表明，前肠（咽）和后肠（直肠） 蠕虫和苍蝇之间的发展已经保存了500多年 百万年。 引人注目的是，imperianus加上食道 闭锁是一种常见的人类出生缺陷，这表明类似的 可能存在于人类中的发育途径。 这一建议将扩大我们的研究咽形成在三个 的方式 目标1。咽部的发育依赖于不同细胞谱系的细胞 组装成一个完整的器官 PHA-4基因标志着 这些不同的细胞谱系通过表型（即， 所有细胞谱系的咽细胞在PHA-4突变体中丢失）， 表达（即，在咽细胞中表达alpha-4：：GFP报告基因 所有的血统）。 要了解多个细胞的会聚 血统到一个器官原基的实现，我们将分析如何 在咽细胞中建立PHA-4表达。 我们将使用一个 缺失分析的组合，报告基因构建体，不同突变体 背景和迁移率改变测定来解决这个问题。 目标2.我们将调查PHA-4是否直接激活两个 用于区分PHA-4的两种模型的候选咽基因 功能：1）PHA-4可以提前启动下游的层次结构， 调节因子2）PHA-4可以是许多或所有 早期和晚期的咽基因。 目标3。咽部发育所需的基因很少被发现。 为了发现形成咽所必需的新位点，我们将 在表型上表现为纯合缺陷， 是咽部发育所必需的 它们分为三种表型 分类：i）分叉咽ii）小咽iii）停滞咽 原基 根据我们的初步调查，我们将选择最 有希望的候选人进行进一步分析。

The genes and mechanisms that drive foregut development are largely unknown and represent a new area of developmental biology. We are studying foregut development by investigating pharynx formation in the nematode Caenorhabditis elegans. Previously, we identified a locus, pha-4, that is required to establish the pharynx and rectum primordia: animals bearing pha-4 mutations arrest as larvae that lack a pharynx and a rectum. We have recently cloned pha-4 and shown that it encodes a homolog of the Drosophila gene fork head. This result, coupled with our phenotypic analyses, suggest that foregut (pharynx) and hindgut (rectum) development have been conserved between worms and flies for over 500 million years. Strikingly, imperforate anus coupled with esophageal atresia is a common human birth defect, suggesting that similar developmental pathways may exist in humans. This proposal will extend our studies of pharynx formation in three ways. Aim 1. Pharynx development depends on cells from different cell lineages assembling into an integrated organ. The pha-4 gene marks the convergence of these different cell lineages both by phenotype (i.e. pharynx cells from all cell lineages are lost in pha-4 mutants) and by expression (i.e. a pha-4::GFP reporter is expressed in pharynx cells from all lineages). To understand how the convergence of multiple cell lineages into one organ primordium is achieved, we will analyze how pha-4 expression is established in pharynx cells. We will use a combination of deletion analysis, reporter constructs, different mutant backgrounds, and mobility shift assays to address this issue. Aim 2. We will investigate whether PHA-4 directly activates two candidate pharynx genes to distinguish between two models of pha-4 function: 1) pha-4 could act early to initiate a hierarchy of downstream regulators 2) pha-4 could be an obligate co-factor for many or all pharynx genes, early and late. Aim 3. Few genes required for pharynx development have been identified. To discover new loci that are necessary to form the pharynx, we will phenotypically characterize homozygous deficiencies that remove genes needed for pharynx development. These fall into three phenotypic classes: i) bifurcated pharynx ii) small pharynx iii) arrested pharynx primordium. On the basis of our initial survey, we will choose the most promising candidates for further analysis.