Molecular Mechanisms of Prion Protein Amyloid Formation

朊病毒蛋白淀粉样蛋白形成的分子机制

基本信息

批准号：
10927847
负责人：
SUZETTE Alise PRIOLA
金额：
$ 36.7万
依托单位：
NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10927847
关键词：
Affect Alzheimer&apos s Disease Amyloid Amyloid Proteins Amyloid beta-Protein Annual Reports Biochemical Biochemistry Biological Assay Biological Models Bovine Spongiform Encephalopathy Brain Cells Cellular Structures Characteristics Chronic Wasting Disease Collaborations Complex Creutzfeldt-Jakob Syndrome Data Deer Deposition Diffuse Disease Environment Equine mule Exposure to Familial Creutzfeldt-Jakob Disease Goals Human Iatrogenesis In Vitro Incidence Infection Infectious Agent Laboratories Manuscripts Medical Modeling Molecular Molecular Chaperones Monitor Mutation Nerve Degeneration Neurodegenerative Disorders Parkinson Disease Pathogenesis Pathogenicity Pathway interactions Peptide Hydrolases Persons Physiological Pilot Projects Postdoctoral Fellow PrP PrP amyloid PrP gene PrPSc Proteins Prion Diseases Prions Process Proteins Publishing Regulation Resistance Science Scrapie Sheep Structure Transgenic Mice Work Writing alpha synuclein amyloid formation cell type in vivo Model insight instrument interest misfolded protein mouse model prion-like protein aggregation protein misfolding tau Proteins uptake

项目摘要

Transmissible spongiform encephalopathies (TSEs or prion diseases) are a group of rare neurodegenerative diseases which include scrapie in sheep, bovine spongiform encephalopathy (BSE), and chronic wasting disease (CWD) in mule deer and elk. In humans, the most common type of prion disease is Creutzfeldt-Jakob disease (CJD) which can occur in several forms. Sporadic CJD (sCJD) makes up the majority of CJD cases and occurs randomly at an incidence of 1-2 per million people worldwide. Iatrogenic CJD (iCJD) is associated with exposure to prion contaminated medical instruments or products while familial CJD (fCJD) is associated with mutations in the prion protein gene. The infectious agent of prion diseases is called a prion and is largely composed of an abnormally refolded, protease resistant form (PrPSc) of the normal, protease-sensitive prion protein, PrPC. PrPSc can be deposited in the brain as either diffuse amyloid negative deposits or as dense amyloid positive deposits. For reasons that are not yet clear, amyloid forms of prion disease appear to be less transmissible than non-amyloid forms. Furthermore, it is unknown if prion diseases where PrPSc is deposited primarily as amyloid follow the same pathogenic processes as prion diseases where PrPSc is primarily deposited as non-amyloid. Multiple studies in transgenic mouse models have shown that amyloid formed from amyloid beta (A) protein, alpha synuclein and tau also propagate via prion-like mechanisms and spread from cell-to-cell (e.g. Science 313: 1781-1784 (2006), Nat Cell Biol 11: 909-913 (2009), J Exp Med 209: 975-986 (2012)). Based on these data, it has been suggested that amyloid formation in neurodegenerative proteinopathies such as Alzheimers Disease (AD) and Parkinsons disease (PD) occurs via prion-like mechanisms and that proteins such as AD-associated A may also be transmissible, infectious prions. Co-deposition of misfolded proteins during neurodegeneration, such as the co-localization of PrPSc and A to plaques in some cases of sCJD (ACTA Neuropathol 96:116-122 (1998)), also suggest that interactions between these proteins could contribute to disease pathogenesis. Laboratory models of prion infection therefore represent a way of studying prion and prion-like mechanisms of disease that can be potentially applied to other neurodegenerative diseases triggered by misfolded proteins. We are interested in understanding the molecular mechanisms underlying PrP amyloid formation and have begun to approach this issue using both in vitro and in vivo model systems. This project focuses primarily on 1) understanding how protein aggregation and disaggregation are controlled by the cell and, 2) understanding the pathways of PrP amyloid formation and spread. Since PrPSc formation and spread appear to be mechanistically similar to the formation and spread of amyloid in other neurodegenerative diseases, the results of our prion studies will likely be broadly applicable to other diseases of protein misfolding and deposition. The ordered aggregation of PrPSc, Abeta, and other amyloid proteins during neurodegeneration is thought to be critical to the pathogenesis of neurodegenerative protein misfolding diseases such as prion disease and AD. However, the processes by which these aggregates form and the mechanisms by which the cell can degrade them remains poorly understood. In earlier studies of how prions interact with cells, we showed that the uptake and disaggregation of prions varied by prion strain (J. Virol. 87: 11552-61 (2013), Annual Report 2013; Am. J. Pathol. 184: 3299-3307 (2014), Annual Report 2014) suggesting that the composition of PrPSc aggregates differed between strains. The post-doctoral fellow in the lab, Dr. Daniel Shoup, has demonstrated that the sizes and stabilities of PrPSc aggregates change during cellular uptake and degradation. He further showed that these changes vary with the prion strain, potentially impacting the ability of a given prion strain to infect cells (Shoup and Priola, Biochemistry 60: 398-411 (2021), Annual Report 2021). In 2023, Dr. Shoup published studies showing that prion aggregates contain both protease-sensitive and resistant forms of prion protein that appear to interact in a regular manner. He used this biochemical characteristic to monitor how the cell tries to unfold and degrade prions during the initial stages of prion infection. His data suggest that the ability of a prion strain to infect a cell may correlate with its ability to protect its core structure from cell-induced structural changes. In 2023, Dr. Shoup used an in vitro protein re-folding assay using purified mammalian chaperones, which he developed, to study how different chaperones interact to unfold PrPSc under physiological relevant conditions. His results show that only some chaperones are able to interact with and alter PrPSc structure. These interactions are prion strain specific and dependent upon pH. His work suggests that only certain combinations of cellular chaperones and environments are conducive to unfolding PrPSc and provides insights into why only some prion strains can infect only certain cell types. He is currently writing the manuscript for this study which should be submitted later this year. We have discovered that prion aggregates have different sizes and stabilities that may affect its ability to infect a cell and replicate (Shoup and Priola, Biochemistry 60: 398-411 (2021), Annual Report 2021). In 2023, this discovery formed the basis of a collaboration with Dr. Byron Caughey's laboratory to characterize protein aggregates found in different human neurodegenerative diseases caused by the same misfolded protein. The goal of this study is to determine if these aggregates differ depending upon the type of disease, and whether these differences impact the ability of the aggregates to induce their own formation. These studies will provide mechanistic insights into how the same misfolded protein can cause different forms of neurodegenerative disease. Finally, in 2023 Dr. Shoup initiated a pilot study to look at how regulation of chaperone expression may impact the interaction of PrP with A. If successful, his work will significantly expand our understanding of any functional relationship between PrP and A and how that may impact the progression of neurodegenerative diseases.

可传播的海绵状脑病（TSE或PRION疾病）是一组罕见的神经退行性疾病，包括绵羊中的刮饼，牛海绵状脑病（BSE）和M子鹿和Elk中的牛皮脑病（BSE）和慢性浪费疾病（CWD）。在人类中，最常见的prion病类型是可以以几种形式发生的克鲁特兹菲尔特 - 贾科布疾病（CJD）。零星的CJD（SCJD）构成了大多数CJD案件，并且在全球范围内以每百万人民为1-2人的发病率随机发生。医源性CJD（ICJD）与暴露于Prion受污染的医疗工具或产品有关，而家族性CJD（FCJD）与Prion蛋白基因中的突变有关。 Prion疾病的感染因子称为prion，主要由正常的，蛋白酶敏感的prion蛋白PRPC的异常重塑，抗蛋白酶抗性形式（PRPSC）组成。 PRPSC可以作为弥漫性淀粉样蛋白负沉积物或致密淀粉样蛋白阳性沉积物沉积在大脑中。由于尚不清楚的原因，淀粉样蛋白的疾病形式似乎不如非淀粉样蛋白形式传播。此外，尚不清楚PRPSC主要作为淀粉样蛋白的pr疾病是否遵循与PRPSC主要沉积为非淀粉样蛋白的prion疾病相同的致病过程。转基因小鼠模型中的多项研究表明，由淀粉样蛋白（A）蛋白，α突触核蛋白和Tau形成的淀粉样蛋白也通过类似prion的机制传播并从细胞到细胞传播（例如Science 313：1781-1784（2006）（2006），Nat Cell Biol 11：909-913（2009），J.99999999.99.99.99.99.99（2009）。基于这些数据，已经提出，诸如阿尔茨海默氏病（AD）和帕金森氏病（PD）的神经退行性蛋白质病中的淀粉样蛋白形成是通过类似prion的机制发生的，并且诸如AD相关A之类的蛋白质也可能是可传播的，感染性的蛋白质也可能是可传播的。神经退行性过程中错误折叠蛋白的共沉积，例如在某些SCJD的情况下，PRPSC和A对斑的共定位（ACTA Neuropathol 96：116：116-122（1998））也表明，这些蛋白质之间的相互作用可能有助于疾病病原病原体。因此，王室感染的实验室模型代表了一种研究疾病的prion和prion样机制的方法，该方法可能可能应用于错误折叠蛋白触发的其他神经退行性疾病。我们有兴趣了解PRP淀粉样蛋白形成的分子机制，并开始使用体外和体内模型系统来解决此问题。该项目主要集中在1）了解蛋白质聚集和分解如何受细胞控制和2）了解PRP淀粉样蛋白形成和扩散的途径。由于PRPSC的形成和扩散似乎在机械上与淀粉样蛋白在其他神经退行性疾病中的形成和扩散相似，因此我们的prion研究结果可能广泛适用于其他蛋白质错误折叠和沉积的疾病。神经退行性过程中PRPSC，ABETA和其他淀粉样蛋白的有序聚集被认为对神经退行性蛋白误折叠疾病（如prion病和AD）的发病机理至关重要。但是，这些聚集体形成的过程以及细胞可以降解它们的机制仍然很少理解。在对王子如何与细胞相互作用的早期研究中，我们表明，prion毒的摄取和分裂因pr菌株而异（J.Virol。87：11552-61（2013），2013年年度报告；Am。J.Pathol。184：3299-3307（2014），2014年年度报告，2014年年度报告，表明PRPS PRPC PRECC的组成不同。实验室的博士后研究员丹尼尔·舒普（Daniel Shoup）博士证明，PRPSC聚集体的大小和稳定性在蜂窝摄取和降解过程中发生了变化。他进一步表明，这些变化随prion菌株的变化而有所不同，可能会影响给定的prion菌株感染细胞的能力（Shoup和Priola，生物化学60：398-411（2021），年度报告2021）。 SHOUP博士在2023年发表了研究，表明prion骨料既包含蛋白酶敏感和抗性形式的prion蛋白，它们似乎是定期相互作用的。他使用这种生化特征来监测细胞在prion感染的初始阶段如何尝试展开和降解prions。他的数据表明，prion菌菌株感染细胞的能力可能与保护其核心结构免受细胞诱导的结构变化的能力相关。在2023年，Shoup博士使用纯化的哺乳动物伴侣进行了体外蛋白质重折叠测定，他开发了这些伴侣，以研究不同的伴侣在生理相关条件下如何相互作用与PRPSC相互作用。他的结果表明，只有一些伴侣能够与PRPSC结构进行交互并改变。这些相互作用是特定于pr菌株的prion菌株，并取决于pH。他的工作表明，只有某些细胞伴侣和环境的组合有利于展开PRPSC，并提供了有关为什么只有某些prion菌株只能感染某些细胞类型的原因。他目前正在为这项研究撰写手稿，该手稿应在今年晚些时候提交。我们发现，prion骨骨料具有不同的大小和稳定性，可能会影响其感染细胞和复制的能力（Shoup and Priola，生物化学60：398-411（2021），年度报告2021）。在2023年，这一发现构成了与拜伦·科伊（Byron Caughey）博士实验室合作的基础，以表征由不同人类神经退行性疾病中发现的蛋白质骨料，由相同错误折叠的蛋白质引起的。这项研究的目的是确定这些骨料是否取决于疾病的类型，以及这些差异是否影响骨料诱导自己的形成的能力。这些研究将提供机械洞察，以了解相同错误折叠的蛋白如何引起不同形式的神经退行性疾病。最后，Shoup博士在2023年开始了一项试点研究，以研究伴侣表达的调节如何影响PRP与A的相互作用。如果成功，他的工作将大大扩展我们对PRP与A之间的任何功能关系的理解以及如何影响神经退行性疾病的进展。