Development of Validated Drosophila in vivo RNAi Models of Human Diseases

描述(申请人提供)：果蝇和人类在基因和基因功能水平上存在显著的保守性。例如，大约三分之二的人类疾病基因都有可识别的苍蝇同源基因，所有主要的信号转导途径在苍蝇和人类之间都是保守的。RNAi用于基因特异性敲除mRNA水平的可用性加快了我们进行使果蝇成为如此强大的系统的分子遗传学分析的速度。然而，尽管目前有一套令人印象深刻的工具可用，定位和/或产生果蝇疾病模型可能是耗时的，而且在大多数情况下，无法获得特征良好的功能丧失等位基因或验证的RNAi株。拟议的项目将建立在我们现有的专业知识和基础设施的基础上，以产生大量体内人类疾病的果蝇RNAi模型，即Hudis-Trip疾病模型资源。这一资源在规模上将是独一无二的，目标是大约900个人类疾病基因的同源果蝇，并在应用于每个RNAi苍蝇种群的高水平质量控制方面是独一无二的。对于基因功能丧失与疾病状态相关的人类疾病基因，Hudis-Trip苍蝇种群很可能通过在细胞、组织和/或生物水平上模拟人类疾病状态来作为疾病模型。具体地说，我们建议：目标1，利用在线孟德尔遗传in Man数据库和社区提名过程，编制人类疾病基因的优先列表；目标2，确定这些基因的900个高置信度果蝇同源基因；每个基因制作两个转基因果蝇RNAi种群；进行表型表征、定量PCR和救援，以验证资源；以及目标3，进一步表征最有希望的模型。值得注意的是，我们最初的保守疾病相关基因清单包括与几乎所有NIH研究所相关的基因。为了履行我们以社区为基础的努力的承诺，所有RNAi苍蝇种群将迅速转移到公共种群中心，所有数据集将通过我们自己的数据库、网站(www.flyrnai.org)和其他数据库提供。我们预计这一资源将被社区广泛使用，因为它将使科学家能够立即开始他们的研究，在一个强大的基因系统中建立高质量的、经过验证的疾病模型。 公共卫生相关性(申请人提出)：该项目将利用果蝇产生大量人类疾病的动物模型，在基因和基因功能水平上与人类有显著的相似之处。这些苍蝇模型将提供一个具有成本效益的工具，促进与各种疾病和几乎所有NIH研究所相关的创新分子和基因研究。

DESCRIPTION (provided by applicant): Remarkable conservation exists between Drosophila and humans at the level of genes and gene functions. For example, there are identifiable fly orthologs for about two-thirds of human disease genes and all of the major signal transduction pathways have been conserved between flies and humans. The availability of RNAi for gene-specific knockdown of mRNA levels has accelerated the pace at which we can undertake the type of molecular genetic analyses that make Drosophila such a powerful system. However, despite the impressive set of tools currently available, locating and/or generating Drosophila disease models can be time consuming, and in most cases, well-characterized loss-of-function alleles or validated RNAi strains are not available. The proposed project would build upon our existing expertise and infrastructure to produce a large collection of in vivo Drosophila RNAi models of human disease, the HuDis-TRiP Resource of Disease Models. This resource will be unique in its large scale, targeting approximately 900 Drosophila orthologs of human disease genes, and in the high level of quality control applied to each RNAi fly stock. For human disease genes for which loss-of-function of the gene is associated with disease state, HuDis-TRiP fly stocks are likely to serve as disease models by mimicking the human disease state at the cell, tissue and/or organism level. Specifically, we propose to: Aim 1, compile a prioritized list of human disease genes using the Online Mendelian Inheritance in Man database and a community nomination process; Aim 2, identify 900 high-confidence fly orthologs of these genes; make two transgenic Drosophila RNAi fly stocks per gene; perform phenotypic characterization, quantitative PCR and rescue to validate the resource; and Aim 3, further characterize the most promising models. Notably, our initial list of conserved disease-associated genes includes genes relevant to nearly all NIH Institutes. In keeping with our commitment to community-based efforts, all RNAi fly stocks will be rapidly transferred to a public stock center and all datasets will be made available through our own database, website (www.flyrnai.org) and other databases. We anticipate that this resource will be widely used by the community, as it will allow scientists to immediately begin their studies with high quality, validated disease models in a powerful genetic system. PUBLIC HEALTH RELEVANCE (proposed by applicant): The proposed project will generate a large number of animal models of human diseases using the fruit fly Drosophila, which shows remarkable similarity to humans at the level of genes and gene functions. These fly models will provide a cost-effective tool facilitating innovative molecular and genetic studies relevant to a wide variety of diseases and nearly all NIH Institutes.