Analysis of Ret signaling in Drosophila enteric nervous system development.

描述(申请人提供)：大约每5000名婴儿中就有一名出生时肠道最下部缺乏神经元。这种情况被称为先天性巨结肠(HSCR)，需要进行矫正手术。受影响神经元的祖细胞出生在发育中的脊髓附近，随后迁移到整个消化道，形成肠道神经系统(ENS)。神经前体使用受体酪氨酸激酶，RET，检测并迁移到一种有吸引力的配体，胶质细胞系源性神经营养因子(GDNF)。人类和小鼠的遗传分析支持RET信号在HSCR中的核心作用，但在活体中的实际作用仍在争论中。ENS领域面临的一个主要挑战是确定在体内形成ENS所需的分子信号。我们相信，利用果蝇强大的遗传学，我们可以发现这些信号。初步数据表明，果蝇Ret是形成ENS所必需的，我们已经确定了另一条平行的信号通路。具体目标1将证实并推广这些发现。无脊椎动物和人类之间的分子功能有显著的保守性，所以我们相信我们的结果将与人类的HSCR相关。直接的遗传分析将确定体内ENS细胞的迁移、增殖、分化或轴突引导是否需要Ret和一条新的无关途径(目标1)。这些信息将与HSCR相关，并与成人ENS情况相关，如糖尿病。我们已经确定了候选的共同受体和配基，并正在组织培养中测试它们的物理联系和信号潜力(特定目标2)。这可能会创造一个机会来分析简单生物体中的GDNF信号，这将与帕金森氏症等非ENS疾病相关。我们还将开发基于Ret启动子分析的新试剂来研究果蝇的形成(特定目标#3)。初步证据表明，我们发现的平行信号通路将与脊椎动物ENS的形成有关。我们的研究可能为HSCR提供新的治疗方法，包括成功进行肠内神经元移植所必需的因素。 公共卫生相关性：Ret基因在肠道(肠道)神经系统的形成中起核心作用，在先天性巨结肠(HSCR)中经常发生突变。尽管动物模型对我们理解HSCR做出了贡献，但还没有建立起无脊椎动物模型。本研究旨在建立果蝇HSCR模型，旨在阐明Ret及其相互作用基因的基本作用机制。

DESCRIPTION (provided by applicant): About one in five thousand babies are born lacking neurons in the lowest part of the intestine. This condition is known as Hirschsprung Disease (HSCR) and requires corrective surgery. The progenitors of the affected neurons are born adjacent to the developing spinal cord and subsequently migrate to populate the entire length of the digestive tract, forming the enteric nervous system (ENS). The neural precursors use a receptor tyrosine kinase, RET, to detect and migrate towards a chemoattractive ligand, Glial Cell Line-Derived Neurotrophic Factor (GDNF). Genetic analysis in humans and mice supports a central role for RET signaling in HSCR, yet the actual role in vivo is under debate. A major challenge for the ENS field is to identify the molecular signals required for ENS formation in vivo. We believe that using Drosophila, with its powerful genetics, we can uncover these signals. Preliminary data indicates that Drosophila Ret is required for ENS formation, and we have identified an additional, parallel signaling pathway. Specific aim #1 will confirm and extend these findings. There is a remarkable conservation of molecular function between invertebrates and humans, so we believe our results will be relevant to HSCR in humans. Straightforward genetic analysis will determine whether Ret and a novel unrelated pathway are required for migration, proliferation, differentiation or axon guidance of ENS cells in vivo (aim #1). This information will be relevant to HSCR, and to adult ENS conditions, such as seen in diabetes. We have identified a candidate co-receptor and ligand and are testing these in tissue culture for physical association and signaling potential (specific aim #2). This could create an opportunity to analyze GDNF signaling in a simple organism, which would have relevance to non-ENS conditions such as Parkinson's disease. We will also develop new reagents for studying Drosophila ENS formation based on Ret promoter analysis (specific aim #3). Preliminary evidence suggests that the parallel signaling pathway we have uncovered will be relevant to vertebrate ENS formation. Our research may offer novel therapeutic approaches to HSCR, including factors necessary for successful enteric neuron transplantation. PUBLIC HEALTH RELEVANCE: The Ret gene plays a central role formation of the enteric (gut) nervous system, being frequently mutated in Hirschsprung's Disease (HSCR). Despite the contribution of animal models to our understanding of HSCR, no invertebrate model has yet been established. This proposal aims to develop a Drosophila model of HSCR with the aim of elucidating the basic mechanisms of action of Ret and interacting genes.