Elucidation of novel pathways components of C. elegans viral immunity

这项研究计划代表了一个新的方向，在我的实验室对发现新的途径参与病毒的先天免疫使用C。我的实验室里的本科生们获得了初步的数据。感兴趣的途径目前在蠕虫中是未知的，但存在于脊椎动物中。他们在C.秀丽隐杆线虫的发现将在这一模式生物的知识方面提供重要的进展，这将使我们对进化过程中这些过程的保护以及它们在脊椎动物中的重要性有新的认识。梭线虫的先天免疫系统在区分非病原性细菌和病原性细菌方面非常有效。但它的基因组中不含有参与哺乳动物感染病毒免疫应答的重要途径成分。在脊椎动物中，先天性免疫应答由识别病毒核酸的特异性受体（称为病原体识别受体（PRR））启动。这些受体通过称为寡核苷酸结合结构域（OB）的结合结构域识别病毒核酸。在感染过程中，这些PRR识别病毒核酸，因此发生一系列信号传导事件以上调抗病毒机制。In C.这些PRR以及激活病毒免疫应答的分子组分是未知的，并且仍有待发现是否C. elegans是PRR。我们在实验室里建立了C。线虫感染系统的dsRNA奥赛病毒。我们现在可以使用反向遗传学工具，已经广泛开发了这种模式生物，并从其中许多试剂可以提供给我们，以确定该病毒激活的途径组件。长期：我们的长期目标是确定未知的病毒免疫途径的分子组成部分激活的奥赛病毒感染的模式生物C。优雅我们可以揭示一个古老的dsRNA抗病毒免疫途径，目前是未知的，可能是保守的动物。短期：目前的方案可以用两个较短的目标来描述。(i)确定OB折叠在C.奥赛野田病毒感染的线虫。哪些OB折叠是奥赛病毒核酸的模式识别受体，触发了C。优雅它们存在吗？我们将使用OB折叠敲低菌株和/或过表达OB基因启动子或编码区的转基因系来确定病毒是否干扰这些基因中的一些基因的基因表达。(ii)确定哪些抗病毒宿主效应物在奥赛病毒感染后作为途径激活的结果而受到调节。我们将使用RNA测序技术来分析感染和未感染菌株之间差异表达的基因。影响：目标1中的拟议研究可能会识别出在C.优雅识别它们是对这种生物体基础知识的重要进步。目标2将深入了解C.尽管已知的脊椎动物途径成分很少，但秀丽线虫对病毒感染产生先天免疫应答。这种新型信号级联的组分的鉴定对于在脊椎动物中发现新型抗病毒效应物和调节物具有真实的潜力。病毒先天免疫反应的这些新特征可能是脊椎动物中进化上保守的古老机制。对这些效应子和调节子的进一步研究可能会为新型抗病毒和抗炎疗法带来候选靶点，以及可能在脊椎动物中错过的新的感染生物标志物。

This research program represents a new direction in my laboratory towards the discovery of novel pathways involved in viral innate-immunity using C. elegans as a model organism that build from preliminary data obtained by undergraduate students in my laboratory. The pathways of interest are currently unknown in the worm, but exist in vertebrates. Their discovery in C. elegans would provide an important advance in knowledge in this model organism that would give us novel insights into the conservation of these processes during evolution and therefore, their importance in vertebrates. The C. elegans innate immune system is remarkably efficient at distinguishing non-pathogenic from pathogenic bacteria. But its genome does not contain the important pathway components that are involved in viral immune response for infection in mammalian. In vertebrates the innate immune response is mounted by specific receptors that recognize viral nucleic-acids called Pathogens Recognition Receptors (PRR). These receptors recognize viral nucleic acids by a binding domain called Oligonucleotide Binding domain (OB). During infection these PRR recognize viral nucleic-acids and as a result a series of signaling events take place to up regulate anti-viral mechanisms. In C. elegans these PRR’s as well as the molecular components that activate the viral immune response are unknown and it remains to be discovered if some members of the OB fold family in C. elegans are PRR. We have established in my laboratory the C. elegans infection system by the dsRNA Orsay virus. We can now use reverse genetics tools that have been extensively developed for this model organism and from which many reagents can be made available to us to identify the pathways components activated by this virus.Long term: Our long term goal is to identify the molecular components of the unknown viral immunity pathways activated by orsay virus infection in the model organism C. elegans. We may reveal an ancient dsRNA antiviral immune pathway that is currently unknown and possibly conserved in animals. Short term: The current program can be described by two shorter objectives. (i) Determine the role of the OB fold in C. elegans infected by nodavirus orsay. Which OB fold are the pattern recognition receptors of the orsay-virus nucleic-acids that trigger the immune response in C. elegans . Do they exist? We will use OB fold knockdown strains and/or transgenic line over-expressing OB gene promotor or coding region to determine if the virus interfere with gene expression for some of these genes. (ii) Determine which antiviral host effectors are regulated as a result of pathway activation after orsay virus infection. We will use RNA sequencing techniques to profile differentially expressed genes between infected and non infected strains.Impact : The proposed research in objective 1 may identify the unknown Pattern Recognition Receptor(s) that are mounting the viral immune response in C. elegans. Identifying them is an important advance in basic knowledge of this organism. Objective 2 will give insight into how C. elegans mounts the innate immune response against viral infection, despite having few of the known vertebrate pathway components. The identification of components of such a novel signaling cascade has real potential for the discovery of novel antiviral effectors and regulators in vertebrates. These novel features of viral innate immune response may be an ancient mechanism evolutionarily conserved within vertebrates. Further studies on these effectors and regulators could lead to candidate targets for novel antiviral and anti-inflammatory therapies as well as new biomarkers for infection that may have been missed in vertebrates.