Whole organism transcriptional profiling of innate immune response

描述（由申请人提供）：脊椎动物的免疫反应由多个分子和细胞成分组成，它们必须相互连接，为宿主物种提供足够的防御病原体的能力。尽管关于个体分子或细胞如何对感染作出反应的信息很多，但由于免疫系统的动态复杂性及其相互依存的先天和适应性功能，对整个生物体对病原体暴露的反应的完整理解仍未得到解决。斑马鱼幼虫为克服这一障碍提供了一个独特的模型，因为幼虫在生命的最初4-6周内成功地保护自己免受病原体的侵害，同时缺乏功能性适应性免疫系统，这使得在整个生物体环境中专门检查先天免疫反应成为可能。在初步的转录谱研究中发现，在斑马鱼幼虫中对病原体相关分子模式作出反应的新基因，也对成年斑马鱼和小鼠的感染刺激作出反应，强调了这种新的先天免疫模型对基因发现的实用性。据推测，斑马鱼幼虫对感染刺激的转录反应将揭示介导哺乳动物先天免疫的新基因。为了验证这一假设，将实现两个目标：1)采用一种新的斑马鱼幼虫实验来确定在缺乏适应性免疫的情况下对病原体刺激的全生物体转录反应；2)验证功能上未表征的病原体应答基因在斑马鱼和哺乳动物先天免疫反应中的作用。该研究的长期目标是在从个体基因到转录组的多个尺度上，在整个生物体的背景下建立先天免疫反应的综合模型。这些实验的成功完成将促进新的未来研究，包括透明斑马鱼幼虫体内感染反应的可视化，基于整个生物体的化学遗传和诱变筛选，以确定免疫反应的药理学和遗传修饰因子，以及基因敲除/敲除研究，以评估这些免疫反应基因在斑马鱼和哺乳动物免疫防御和恢复中的作用。这些数据、试剂和工具将构成一种新的基于系统的、多生物的研究、评估和调节脊椎动物先天免疫反应的范式。

DESCRIPTION (provided by applicant): The vertebrate immune response is comprised of multiple molecular and cellular components that must interface to provide the host species with an adequate defense against pathogens. Although much information is available on how individual molecules or cells respond to infection, a complete understanding of the whole-organism response to pathogen exposure remains unresolved, due to the dynamic complexity of the immune system and its interdependent innate and adaptive functionality. The zebrafish larva provides a unique model for overcoming this obstacle as the larva successfully defends itself from pathogens while lacking a functional adaptive immune system for the first 4-6 weeks of life, making it possible to examine exclusively the innate immune response in a whole-organism context. In preliminary transcriptional profiling studies it was found that novel genes that respond to pathogen associated molecular patterns in the zebrafish larva, also respond to infection stimuli in adult zebrafish and mice, underscoring the utility of this novel innate immune model for gene discovery. It is hypothesized that the transcriptional response of zebrafish larvae to infection stimuli will reveal novel genes that mediate innate immunity in mammals. In order to test this hypothesis, two Aims will be achieved, 1) a novel zebrafish larvae assay will be employed to determine the whole-organism transcriptional response to pathogen stimuli in the absence of adaptive immunity, and 2) the role of functionally uncharacterized pathogen-responsive genes will be validated in zebrafish and mammalian innate immune response. The long range goal of the proposed research is to develop an integrative model of innate immune response in the context of the whole organism, on multiple scales, from the individual gene to the transcriptome. Successful completion of the proposed experiments will facilitate novel future investigations, including in vivo visualization of infection response in the transparent zebrafish larva, whole-organism based chemical genetic and mutagenesis screens to identify pharmacological and genetic modifiers of immune response, and gene knock-out/knock- down studies for assessing the role of these immune response genes in immune defense and recovery in both zebrafish and mammals. These data, reagents and tools will constitute a novel systems-based, multi- organism paradigm for investigating, evaluating and modulating the vertebrate innate immune response. PUBLIC HEALTH RELEVANCE The complete, dynamic process by which a vertebrate responds to infection on the organismal level is not well understood. The proposed research will utilize a novel strategy to characterize the genomic response of zebrafish to immune stimuli and use this information to identify uncharacterized mammalian genes that respond to pathogen exposure. The data generated by these methods will provide unprecedented insight into the organism based genomic response to infection, facilitating novel diagnosis, treatment and drug discovery strategies for modulating immune response.