A genome-wide RNAi screen for Neuronal Cell Fate Mutants

描述（由申请人提供）：“A genome-wide RNAi screen for neuronal cell fate mutants”在神经系统中产生单个神经元细胞类型的调节机制尚未完全理解。模型系统中的遗传突变体筛选以及脊椎动物中的反向遗传方法已经对神经元多样化的机制产生了有价值的见解，因为这些方法已经发现了参与这些过程的基因。我们在这里建议使用全基因组RNA干扰（RNAi）为基础的反向遗传学方法在顺从的模式生物C。elegans的基因，以揭示参与控制一对感觉神经元，称为ASE神经元的规范。我们要解决的问题是：（1）ASE神经元产生和正确指定所需的基因是什么？（2）由于两个ASE神经元以左/右不对称表达的化学感受器基因的形式显示出左/右不对称特征，我们希望确定参与这种左/右不对称发育程序的基因。到目前为止，我们已经完成了对染色体I（2446个克隆）的RNAi分析，并鉴定了14个导致ASE神经元发育中可重复缺陷的RNAi克隆。我们在这里建议将这种方法扩展到所有六条染色体，从而估计可能参与ASE神经元发育的6 x 14 = 84个基因的恢复。探索性的两年R 03资助框架为我们提供了建立这样一个基因集合的机会，这些基因在ASE发展中发挥作用。这个集合将提供一个宝贵的资源和起点，在未来进行深入的和假设驱动的基因分析，最终导致详细了解的发展计划，导致一个单一的神经元类的规范。公共卫生相关性：“神经元细胞命运突变体的全基因组RNAi筛选”这项拨款提案旨在利用线虫C. elegans作为一个模型系统。我们将使用基于RNA干扰的筛选来识别个体感觉神经元类发育所需的基因，并以适当的方式采用其左/右不对称特征。

DESCRIPTION (provided by applicant): "A genome-wide RNAi screen for neuronal cell fate mutants" The regulatory mechanisms that generate individual neuronal cell types in the nervous system are incompletely understood. Genetic mutant screens in model systems as well as reverse genetic approaches in vertebrates have yielded valuable insights into the mechanisms of neuronal diversification as these approaches have uncovered genes involved in these processes. We propose here to use a genome-wide RNA interference (RNAi)-based reverse genetics approach in the amenable model organism C. elegans to uncover genes involved in controlling the specification of a pair of sensory neurons, called the ASE neurons. The questions that we will address are: (1) what are the genes required for the ASE neurons to be generated and correctly specified; (2) as the two ASE neurons display the intriguing feature of showing left/right asymmetric features in the form of left/right asymmetrically expressed chemoreceptor genes, we wish to identify genes involved in this left/right asymmetric developmental program. So far, we have finished RNAi analysis of the chromosome I (2446 clones) and have identified 14 RNAi clones that lead to a reproducible defect in the development of the ASE neurons. We propose here to scale up this approach to all six chromosomes, leading to an estimated recovery of 6 x 14 = 84 genes potentially involved in ASE neuron development. The framework of an exploratory two-year R03 grant provides us with the opportunity to build such a collection of genes with a role in ASE development. This collection will provide an invaluable resource and starting point for a in-depth and hypothesis-driven gene-by-gene analysis in the future, eventually leading to a detailed understanding of the developmental program that leads to the specification of a single neuron class. PUBLIC HEALTH RELEVANCE: "A genome-wide RNAi screen for neuronal cell fate mutants" This grant proposal sets out to identify genes involved in neuronal development using the nematode C. elegans as a model system. We will use an RNA interference-based screen to identify genes that are required for an individual sensory neuron class to develop and to adopt its left/right asymmetric features in an appropriate manner.