Probing organismal proteostasis through the response to intracellular infection

通过对细胞内感染的反应探索机体蛋白质稳态

• 批准号: 9353488
• 负责人: Emily R Troemel
• 金额: $ 38.75万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-30 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9353488
• 关键词: Aging; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Animals; Binding; Biochemical; Boxing; Budgets; CUL1 gene; Caenorhabditis elegans; Caring; Cells; Complex; Cullin Proteins; Data; Defect; Degenerative Disorder; Degradation Pathway; Disease; F Box Domain; Gene Expression; Genes; Genetic; Genetic Epistasis; Genetic Screening; Genetic Transcription; Glutamine; Goals; Growth; Health; Healthcare; Healthcare Systems; Homeostasis; Human; Huntington Disease; Immune response; Infection; Intestines; Lead; Ligase; Maintenance; Mediating; Methods; Microbe; Microsporidia; Molecular; Monitor; Nematoda; Neurodegenerative Disorders; Outcome; Parkinson Disease; Pathway interactions; Patient Care; Patients; Proteasome Inhibition; Proteins; Proteomics; RNA Interference; RNA Viruses; Research; Resistance; Role; Signal Pathway; Source; Stress; Structure; Testing; Tissues; Ubiquitination; Up-Regulation; Virus; age related; base; coping; improved; innovation; insight; mutant; novel; novel strategies; novel therapeutics; pathogen; polyglutamine; pressure; protein aggregate; protein misfolding; response; stressor; transcriptome sequencing; ubiquitin ligase; ubiquitin-protein ligase

 DESCRIPTION (provided by applicant): Maintenance of protein homeostasis, or proteostasis, is critical for health. Proteostasis is perturbed in several aging-related diseases including neurodegenerative diseases like Alzheimer's, Parkinson's, and Huntington's diseases. Caring for patients with these diseases is consuming an increasing fraction of our health care budget. Strikingly, care for Alzheimer's patients alone is responsible for $226 billion in spending per year, and no treatments are available. Thus, new approaches are needed. One such approach involves leveraging the host/pathogen response to infection with intracellular microbes. Our long-term goal is to dissect the mechanisms by which host cells upregulate proteostasis pathways to cope with the increased burden of intracellular infection and replication. Closing this gap in our understanding will provide new insights about proteostasis, and could provide novel treatments for neurodegenerative diseases. Our central hypothesis is that hosts can sense the effects of intracellular infection and increase proteostasis capacity to cope with this increased burden. The objective here is to determine the mechanisms by which the nematode C. elegans upregulates ubiquitin ligase components in response to infection by a natural intracellular microbe that belongs to the microsporidia phylum. Microsporidia commonly infect all animals including humans, and can replicate to very high levels without causing overt effects on the host, likely due to host compensatory mechanisms. Our recent findings indicate that C. elegans upregulates ubiquitin ligase components in response to diverse intracellular infections including microsporidia and virus (Bakowski et al 2014). In unpublished data we have isolated mutants defective in an F-box-related gene (fbxr-1) that constitutively express these ubiquitin ligase components. fbxr-1 mutants have increased pathogen resistance, as well as greatly enhanced thermotolerance and reduced levels of aggregated proteins, indicating improved proteostasis capacity. We hypothesize that C. elegans increases transcription of Skp-Cullin-F-box (SCF) ubiquitin ligase components in order to target misfolded proteins for ubiquitination and destruction, increasing tolerance of proteotoxic insults. In Specific Aim 1 we will determine where the Cullin, as well as its upstream negative regulator FBXR-1 act to regulate thermotolerance. We also will perform structure/function analysis on the Cullin to test the hypothesis that it is part of a multi-subunit SCF ubiquitin ligase component. In Specific Aim 2 we will identify these other SCF ligase components using genetic and biochemical approaches. In Specific Aim 3 we will identify new components of the FBXR-1/Cullin pathway using RNAseq analysis, genetic epistasis, and a forward genetic screen. In Specific Aim 4 we will determine where and when FBXR-1 regulates levels of protein aggregates. This approach is innovative because it leverages the host response to obligate intracellular infection to understand how proteostasis can be improved. The proposed research is significant because it could lead to new treatments for diseases of compromised proteostasis, such as Alzheimer's disease.

 描述(由申请人提供)：维持蛋白质平衡，或蛋白质平衡，对健康至关重要。蛋白质平衡在几种与衰老相关的疾病中受到干扰，包括阿尔茨海默氏症、帕金森氏症和亨廷顿病等神经退行性疾病。护理这些疾病的患者在我们的医疗保健预算中所占的比例越来越大。引人注目的是，仅照顾阿尔茨海默氏症患者一项就花费了226亿美元，而目前还没有可用的治疗方法。因此，需要新的方法。一种这样的方法涉及利用宿主/病原体对细胞内微生物感染的反应。我们的长期目标是剖析宿主细胞上调蛋白平衡途径以应对细胞内感染和复制增加的负担的机制。结束这件事 我们在理解上的差距将为蛋白质代谢提供新的见解，并可能为神经退行性疾病提供新的治疗方法。我们的中心假设是，宿主可以感觉到细胞内感染的影响，并增加蛋白稳定能力，以应对这种增加的负担。这里的目的是确定线虫上调泛素连接酶组分的机制，以响应属于微孢子虫门的一种自然细胞内微生物的感染。微孢子虫通常感染包括人类在内的所有动物，并且可以复制到非常高的水平，而不会对宿主造成明显影响，这可能是由于宿主的补偿机制。我们最近的发现表明，线虫在应对包括微孢子虫和病毒在内的各种细胞内感染时，上调泛素连接酶组分(Bakowski等人，2014年)。在未发表的数据中，我们分离出了F-box相关基因(fbxr-1)中有缺陷的突变株，该突变株结构性地表达这些泛素连接酶成分。Fbxr-1突变体的抗病性增强，耐热性显著增强，聚集态蛋白水平降低，蛋白稳定能力增强。我们假设线虫增加了Skp-cullin-F-box(SCF)泛素连接酶组分的转录，以便靶向错误折叠的蛋白质进行泛素化和破坏，从而增加对蛋白毒性侮辱的耐受性。在具体目标1中，我们将确定cullin及其上游负调控因子FBXR-1在哪里调节耐热性。我们还将对cullin进行结构/功能分析，以验证它是多亚基SCF泛素连接酶组件的假设。在特定目标2中，我们将使用遗传和生化方法鉴定其他SCF连接酶组分。在具体目标3中，我们将使用RNAseq分析、遗传上位性和正向遗传筛选来识别FBXR-1/Cullin途径的新组件。在具体目标4中，我们将确定FBXR-1在哪里以及何时调节蛋白质聚集体的水平。这种方法是创新的，因为它利用宿主的反应来专职细胞内感染，以了解如何改善蛋白稳定。这项拟议的研究意义重大，因为它可能导致蛋白质平衡受损的疾病的新疗法，如阿尔茨海默病。