The role of host endogenous small RNAs in innate immunity

描述(申请人提供)：包括哺乳动物、无脊椎动物和植物在内的生物体已经进化出复杂和保守的先天免疫反应，以保护自己免受病原体的攻击。在这些系统中，宿主免疫反应和抗病是通过调节大量基因来实现的，但这些过程是如何调控的在很大程度上仍不清楚。小RNA是一种非编码的调节RNA分子，通过介导mRNA降解、翻译抑制或染色质修饰来控制基因的表达。我们和其他人的工作证明了宿主内源性小RNAs，包括microRNAs(MiRNAs)和小干扰RNAs(SiRNAs)在抗菌天然免疫中的作用。我们发现，几种病原菌诱导的内源siRNAs，包括天然反义转录本衍生的siRNAs(NAT-siRNAs)和长siRNAs(LsiRNAs)，通过转录后沉默植物免疫系统的负调控因子来促进宿主的免疫反应。在感染细菌的植物中也观察到了表观遗传变化，然而，小RNA引导的DNA甲基化和染色质修饰在植物免疫中的作用尚未被探索。最近，我们以拟南芥为模型系统，以Illumina深度测序为平台，对细菌攻击后的小RNA种群进行了分析，并鉴定了受细菌病原体感染调控的不同类别的内源小RNA。我们假设，这些病原体响应的小RNA通过转录或转录后沉默宿主免疫调节因子来调节宿主免疫反应中涉及的基因的表达。我们建议研究这些病原体响应的小RNA的调节、生物发生和功能。具体地说，我们将对几个新发现的miRNAs、NAT-siRNAs、lsiRNAs和簇状HC-siRNAs进行深入的鉴定，这些miRNAs受到不同菌株丁香假单胞菌的特殊调控。预测的这些小RNA的靶基因将得到验证并进行功能分析。这些靶标很可能是参与植物免疫反应途径的成分。我们还将描述在细菌感染时引导DNA甲基化的HC-siRNAs。我们将生成细菌感染前后的全基因组DNA甲基化图谱，并将siRNAs、DNA甲基化和病原体反应基因表达联系起来。这项拟议的研究将极大地促进我们对小RNA参与宿主天然免疫的机制和功能的理解。 公共卫生相关性：多细胞真核生物，包括哺乳动物、无脊椎动物和植物，已经进化出复杂和保守的先天免疫反应，以保护自己免受病原体的攻击。这项研究将极大地促进我们对小RNA介导的基因调控机制和功能的理解，以模式生物拟南芥为例。

DESCRIPTION (provided by applicant): Organisms, including mammals, invertebrates, and plants have evolved sophisticated and conserved innate immune responses to protect themselves from pathogen attacks. In these systems, host immune responses and disease resistance are achieved by modulation of a large array of genes, but how these processes are regulated is still largely unclear. Small RNAs are non-coding regulatory RNA molecules that control gene expression by mediating mRNA degradation, translational inhibition, or chromatin modification. Our work and those of others have demonstrated a role of host endogenous small RNAs, including microRNAs (miRNAs) and small interfering RNAs (siRNAs), in antibacterial innate immunity. We discovered that several pathogen-induced host endogenous siRNAs, including natural antisense transcripts-derived siRNAs (nat-siRNAs) and long siRNAs (lsiRNAs), facilitate host immune responses by posttranscriptionally silencing the negative regulators of plant immune systems. Epigenetic changes were also observed in bacteria-infected plants, however, the role of small RNA-guided DNA methylation and chromatin modification in plant immunity has not been explored. Using Arabidopsis as a model system and Illumina deep sequencing as a platform, we recently profiled the small RNA population after bacterium-challenge and identified diverse classes of endogenous small RNAs that are regulated by the infection of bacterial pathogens. We hypothesize that these pathogen-responsive small RNAs regulate the expression of genes involved in host immune responses by silencing regulators of host immunity transcriptionally or post-transcriptionally. We propose to investigate the regulation, biogenesis, and function of these pathogen-responsive small RNAs. Specifically, we will perform in depth characterization of several newly identified miRNAs, nat-siRNAs, lsiRNAs, and clustered hc-siRNAs that are particularly regulated by various strains of the bacterium Pseudomonas syringae. Predicted target genes of these small RNAs will be validated and subjected to functional analysis. These targets are likely to be components involved in plant immune response pathways. We will also characterize the hc-siRNAs that directed DNA methylation in response to bacterial infection. We will generate a whole-genome DNA-methylation map before and after bacterial infection, and correlate the siRNAs, DNA methylation, and pathogen-responsive gene expression. The proposed research will significantly advance our understanding of the mechanisms and functions of small RNAs involved in host innate immunity. PUBLIC HEALTH RELEVANCE: Multicellular eukaryotes, including mammals, invertebrates, and plants have evolved sophisticated and conserved innate immune responses to protect themselves from pathogen attack. The proposed research will significantly advance our understanding of the mechanisms and function of small RNA-mediated gene regulation in innate immunity using the model organism Arabidopsis thaliana.