The Intracellular Pathogen Response Triggers Defense Against Co-evolved Pathogens

项目摘要 传染病给人类健康带来重大负担，而上皮细胞处于第一线 这些致病微生物可以侵入这些细胞并在其中复制。除了 病原体引起的腹泻和肺炎等疾病，防御途径的不适当激活 会导致炎症性疾病。因此，了解更多关于上皮防御的知识是至关重要的。 对抗细胞内病原体特别是，对微孢子虫门的防御知之甚少。 真菌相关的细胞内寄生虫，其中14种可感染并导致人类疾病，最常见的是 感染肠上皮细胞我们已经开发了一种方便的整体动物模型来研究防御 通过对线虫C.优雅我们 长期目标是剖析上皮细胞防御共同进化的细胞内 病原体如微孢子虫。弥合我们理解上的这一差距将为治疗提供新的见解。 感染性疾病和炎性疾病。我们的核心假设是共同进化的病原体的攻击 导致宿主基因的扩展和多样化，使其成为“物种特异性”，尽管这些基因可能 控制保守的免疫途径。本文的目的是描述物种特异性的帕尔斯基因 该家族在C.而人类只有一个帕尔斯基因。 事实上，我们对帕尔斯蛋白质的结构或在任何系统中的生化功能都一无所知，尽管它们在某些系统中起着重要的作用。 通过序列分析和遗传学研究，C.优雅 我们发现帕尔斯-22和帕尔斯-25是对天然转录因子的共同转录应答的关键调节因子。 微孢子虫和病毒感染C.我们称之为细胞内病原体反应或IPR。的 IPR似乎定义了整个生理程序，我们的正向遗传筛选将帕尔斯-22鉴定为 作为IPR的负调节剂和作为正调节剂的帕尔斯-25。帕尔斯-22的丢失导致增强的 对细胞内病原体的免疫力，增加的RNA干扰，以及健身的后果， 所有这些都依赖于帕尔斯-25。在具体目标1中，我们将描述 基因表达与帕尔斯-22和帕尔斯-25调节的免疫应答之间的关系，鉴定 它们作用的组织，并确定它们所靶向的微孢子虫的阶段。具体目标2 重组表达帕尔斯-22和帕尔斯-25蛋白以表征它们的相互作用，以及它们的相互作用。 结构使用X射线晶体学和cryo EM。在具体目标3中，我们将分析其他帕尔斯的作用 蛋白质，RNAi机制，并确定新的监管机构的知识产权。该方法是创新的，因为它侧重于 调节一种新型上皮免疫的未表征蛋白质。拟议的研究意义重大， 因为它可能会导致传染病和炎症性疾病的新疗法。

Project Summary Infectious diseases impose a significant burden on human health, with epithelial cells on the front lines of attack by disease-causing microbes that can invade and replicate inside of these cells. In addition to diseases such as diarrhea and pneumonia caused by pathogens, inappropriate activation of defense pathways in epithelial cells can lead to inflammatory disease. Therefore, it is critical learn more about epithelial defense against intracellular pathogens. In particular, little is known about defense against the Microsporidia phylum of fungal-related intracellular parasites, 14 of which can infect and cause disease in humans, most commonly infecting intestinal epithelial cells. We have developed a convenient whole-animal model for studying defense against microsporidia through characterization of natural intestinal infection in the nematode C. elegans. Our long-term goal is to dissect the mechanisms by which epithelial cells defend against co-evolved intracellular pathogens like microsporidia. Closing this gap in our understanding will provide new insights for treating infectious disease and inflammatory disorders. Our central hypothesis is that attack from co-evolved pathogens causes expansion and diversification of host genes to become ‘species-specific’ although these genes may control conserved immune pathways. The objective here is to characterize the species-specific pals gene family, which expanded to 39 pals genes in C. elegans, whereas there is only one pals gene in humans. Virtually nothing is known about PALS protein structure or biochemical function in any system, although they have been connected to ubiquitin ligases through sequence analysis and genetic studies in C. elegans. We found PALS-22 and PALS-25 to be key regulators of a common transcriptional response to natural microsporidia and viral infections in C. elegans that we call the Intracellular Pathogen Response or IPR. The IPR appears to define an entire physiological program and our forward genetic screens identified PALS-22 as a negative regulator and PALS-25 as a positive regulator of the IPR. Loss of PALS-22 leads to enhanced immunity against intracellular pathogens, increased RNA interference, as well as fitness consequences such as delayed development, all of which depend on PALS-25. In Specific Aim 1 we will characterize the relationship between gene expression and immune responses regulated by PALS-22 and PALS-25, identify the tissues where they act, and define the stage of microsporidia they target. In Specific Aim 2 we will recombinantly express PALS-22 and PALS-25 proteins to characterize their interaction, as well as their structure using X-ray crystallography and cryo EM. In Specific Aim 3 we will analyze the role of other PALS proteins, RNAi machinery and identify new regulators of the IPR. The approach is innovative as it focuses on uncharacterized proteins that regulate a novel form of epithelial immunity. The proposed research is significant, because it could lead to new treatments for infectious diseases and inflammatory disorders.