Perturbation of core processes triggers host defense against pathogens

 描述(申请人提供)：肠道上皮细胞必须检测并对导致食源性和水源性疾病的微生物病原体做出反应，但也要将它们与肠道中数万亿种无害甚至有益的微生物区分开来。宿主细胞做出这种区分的机制还知之甚少。我们的长期目标是更好地了解病原体是如何通过肠道细胞以及其他细胞类型与其他微生物区分开来的。缩小我们在认识上的这一差距可能会改善传染病的治疗，以及更好地控制炎症性疾病。我们的中心假设是，区分病原体和其他微生物的一个主要策略是通过感知核心宿主过程的扰动，这些过程通常是病原体衍生毒素的靶标。这里的目的是确定线虫检测这种病原体诱导的扰动的机制。线虫提供了一个很好的系统来解决这个问题，因为它依赖于上皮防御，并且非常容易驯服。我们还将把我们的发现扩展到哺乳动物身上，以研究类似的保护性宿主反应。我们最近的发现表明，像mRNA翻译这样的核心过程的扰动会触发线虫bZIP转录因子ZIP-2的激活，以提供宿主防御(Dunbar等人，2012)。Zip-2上调了一组参与细胞内防御的基因，并促进了对感染的抵抗力(Estes等人，2010年)。我们未发表的结果表明，ZIP-2与CEBP-2协同工作，CEBP-2是线虫与哺乳动物C/eBP-的同源基因，后者是一种转录因子，介导哺乳动物对感染的急性反应。我们假设1)ZIP-2和CEBP-2形成一个促进宿主防御的异源二聚体转录因子，2)ZIP-2的翻译被上游开放阅读框架(UORF)上调，uORF作为翻译衰减的传感器，3)哺乳动物使用类似的宿主防御系统。我们将在特定目标1中测试我们的第一个假设，在那里我们将检查CEBP-2和ZIP-2的表型、表达和相互作用。我们将在特定目标2中测试我们的第二个假设，在那里我们调查翻译衰减增加ZIP-2蛋白水平这一令人惊讶的结果的潜在机制。我们将分析Zip-2uORF(Dunbar等人，2012年)，并从ZIP-2调控因子的遗传筛选中鉴定HITS。在目标3中，我们将在哺乳动物中建立我们的初步发现，测试C/eBP-在对毒素的保护性反应中的作用，并研究这种反应在体内的功能意义。这种方法是创新的，因为它分析了细胞如何检测病原体靶向的核心过程中的扰动，这是一种新的动物宿主防御模式。它还研究了uORF的作用和功能，uORF存在于30%-50%的小鼠和人类基因中，但人们对此知之甚少。拟议的研究具有重要意义，因为它将深入了解细胞如何对病原体攻击做出具体反应，并可能导致肠道和其他组织感染的新治疗方法，以更好地对抗传染病。

 DESCRIPTION (provided by applicant): Intestinal epithelial cells must detect and respond to microbial pathogens that cause foodborne and waterborne diseases, yet also discriminate them from the trillions of other microbes in the intestine that are innocuous or even beneficial. The mechanisms by which host cells make this distinction are poorly understood. Our long-term goal is to better understand how pathogens are discriminated from other microbes by intestinal cells, as well as by other cell types. Closing this gap in our understanding may allow for improved treatment of infectious diseases, as well as better control of inflammatory disorders. Our central hypothesis is that a major strategy for discriminating pathogens from other microbes is through sensing perturbations of core host processes that are commonly targeted by pathogen-derived toxins. The objective here is to identify the mechanism by which the nematode C. elegans detects such pathogen-induced perturbations. C. elegans provides an excellent system to address this question because it relies on epithelial defense and is extremely tractable. We will also extend our findings to mammals, to investigate a similar protective host response. Our recent findings have shown that perturbation of core processes like mRNA translation triggers activation of the C. elegans bZIP transcription factor ZIP-2 to provide host defense (Dunbar et al, 2012). ZIP-2 upregulates a suite of genes involved in intracellular defense, and promotes resistance to infection (Estes et al, 2010). Our unpublished results suggest that ZIP-2 works together with CEBP-2, which is the C. elegans ortholog of mammalian C/EBP-, a transcription factor that mediates acute response to infection in mammals. We hypothesize that 1) ZIP-2 and CEBP-2 form a heterodimeric transcription factor that promotes host defense, 2) ZIP-2 translation is upregulated upon infection by an upstream open reading frame (uORF) that acts as a sensor for translational attenuation, and 3) mammals deploy a similar host defense system. We will test our first hypothesis in Specific Aim 1, where we will examine CEBP-2 and ZIP-2 phenotypes, expression and interaction. We will test our second hypothesis in Specific Aim 2, where we investigate the underlying mechanisms of the surprising result that translational attenuation increases ZIP-2 protein levels. We will analyze the zip-2 uORF (Dunbar et al, 2012), and characterize hits from a genetic screen for regulators of ZIP- 2. In Aim 3, we will build off our preliminary findings into mammals, testing a role for C/EBP- in the protective response to toxins, and investigating the functional significance of this response in vivo. This approach is innovative, because it analyzes how cells detect perturbations in core processes targeted by pathogens, which is a newly appreciated mode of animal host defense. It also investigates the role and function of uORFs, which are found in 30-50% of all mouse and human genes, but poorly understood. The proposed research is significant because it will provide insight into how cells respond specifically to pathogenic attack, and may lead to new treatments for infections in the intestine as well as other tissues, to better combat infectious disease.