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）：维持蛋白质稳态或蛋白质稳态对健康至关重要。蛋白质稳态在几种与衰老有关的疾病中受到干扰，包括神经退行性疾病，如阿尔茨海默病、帕金森病和亨廷顿病。照顾这些疾病的患者正在消耗我们医疗保健预算的越来越大的一部分。引人注目的是，仅老年痴呆症患者的护理每年就要花费2260亿美元，而且没有治疗方法。因此，需要新的办法。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应答。我们的长期目标是剖析宿主细胞上调蛋白质稳态途径以科普细胞内感染和复制的增加负担的机制。关闭此 我们理解的差距将提供关于蛋白质稳态的新见解，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，并增加蛋白质稳定能力，以科普这种增加的负担。本文的目的是确定线虫C.线虫响应于被属于微孢子虫门的天然细胞内微生物感染而上调泛素连接酶组分。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的影响，这可能是由于宿主的补偿机制。 我们最近的研究结果表明，C。秀丽线虫响应于包括微孢子虫和病毒在内的多种细胞内感染而上调泛素连接酶组分（Bakowski et al 2014）。在未发表的数据中，我们分离出了F盒相关基因（fbxr-1）缺陷的突变体，这些基因组成型表达泛素连接酶组分。FBXR-1突变体具有增加的病原体抗性以及大大增强的耐热性和降低的聚集蛋白水平，表明改善的蛋白质稳定能力。 我们假设C.线虫增加Skp-Cullin-F-box（SCF）泛素连接酶组分的转录，以靶向错误折叠的蛋白质进行泛素化和破坏，增加对蛋白毒性损伤的耐受性。在具体目标1中，我们将确定Cullin及其上游负调节因子FBXR-1在调节耐热性方面的作用。我们还将对Cullin进行结构/功能分析，以检验它是多亚基SCF泛素连接酶组分的一部分的假设。在具体目标2中，我们将使用遗传和生物化学方法鉴定这些其他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鉴定FBXR-1/Cullin途径的新组分。在具体目标4中，我们将确定FBXR-1在何处以及何时调节蛋白质聚集体的水平。这种方法是创新的，因为它利用宿主对专性细胞内感染的反应来了解如何改善蛋白质稳态。这项拟议中的研究意义重大，因为它可能导致蛋白质稳态受损疾病的新疗法，如阿尔茨海默病。