Cell-type specific wiring of the chaperone network

伴侣网络的细胞类型特定布线

• 批准号: RGPIN-2019-06230
• 负责人: Mok, SueAnn
• 金额: $ 2.19万
• 依托单位: University of Alberta
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=714291
• 关键词: Cell; type; specific; wiring; chaperone

My research program is aimed at dissecting the molecular mechanisms that allow the chaperone network to adapt to distinct and fluctuating protein folding environments in cells. Our current focus is to determine if cellular specialization (differences between cell types) modulates chaperone network function in two key aspects 1) the proteins vulnerable to misfolding and 2) the chaperones responsible for regulating specific protein misfolding targets. The human chaperone network is a cellular system composed of over 300 members that function in concert to maintain protein quality control. Chaperones recognize protein misfolding throughout the proteome and facilitate a multitude of responsive processes e.g. protein folding/refolding, sequestration of aggregates, and delivery to protein degradation systems. The chaperone network is incredibly complex; the majority of chaperones participate in numerous dynamic interactions with misfolded substrates, other chaperones, and the rest of the proteostasis machinery. Chaperone network functions regulate the folding of proteins required in all cellular processes and thus, it is critical that we decipher the underlying principles of this complex system. Human are composed of a number of distinct cell types that have specialized functions, and express the specific requisite proteins to carry out these functions. A fundamental question regarding chaperone networks is whether the system is specifically tailored to the protein quality control demands presented in a given cell type. There is evidence that expression of individual chaperones within the network varies across cell types. However, it is unknown if cell-type differences in chaperone and proteome profiles have profound effects on the pool of proteins susceptible to misfolding and the chaperones recruited to regulate specific protein misfolding targets. We will utilize iPSCs to generate three distinct cell types (undifferentiated, cardiomyocytes, neurons) in order to dissect and compare key aspects of chaperone network functions. Our first objective is to quantify proteome and chaperone network composition differences across cell types using mass spectrometry. Our second objective is to knockdown core chaperones (e.g. Hsc70, Hsp72) in all three cell types and determine 1) the protein targets susceptible to aggregation and if they are common across cell types and 2) if protein misfolding propensity within a given cell type is correlated with characteristics such as chaperone interactors, expression levels, and shared biochemical properties. Our third objective is to challenge cells with new chaperone substrates, not typically expressed in our cell types, and compare outcomes on substrate folding regulation, including the chaperones involved in these processes. Our studies will provide fundamental insights about the principles governing the relationships between the chaperone network, its substrates, and its resilience against protein folding stresses.

我的研究项目旨在剖析分子伴侣网络适应细胞中独特且波动的蛋白质折叠环境的分子机制。我们目前的重点是确定细胞特化（细胞类型之间的差异）是否在两个关键方面调节伴侣网络功能：1）容易发生错误折叠的蛋白质；2）负责调节特定蛋白质错误折叠目标的伴侣。 人类伴侣网络是一个由 300 多个成员组成的细胞系统，它们协同作用以维持蛋白质质量控​​制。分子伴侣识别整个蛋白质组中的蛋白质错误折叠，并促进多种响应过程，例如蛋白质折叠/重折叠、聚集体的隔离以及输送至蛋白质降解系统。伴侣网络极其复杂；大多数伴侣参与与错误折叠底物、其他伴侣和蛋白质稳态机制的其余部分的许多动态相互作用。伴侣网络功能调节所有细胞过程所需的蛋白质折叠，因此，我们破译这个复杂系统的基本原理至关重要。 人类由许多具有特殊功能的不同细胞类型组成，并表达执行这些功能所需的特定蛋白质。关于伴侣网络的一个基本问题是该系统是否专门针对给定细胞类型中提出的蛋白质质量控​​制需求而定制。有证据表明，网络内各个伴侣的表达因细胞类型而异。然而，尚不清楚分子伴侣和蛋白质组谱中的细胞类型差异是否对易发生错误折叠的蛋白质库以及为调节特定蛋白质错误折叠目标而招募的分子伴侣产生深远影响。 我们将利用 iPSC 生成三种不同的细胞类型（未分化细胞、心肌细胞、神经元），以便剖析和比较伴侣网络功能的关键方面。我们的第一个目标是使用质谱法量化不同细胞类型之间的蛋白质组和伴侣网络组成差异。我们的第二个目标是敲除所有三种细胞类型中的核心伴侣（例如 Hsc70、Hsp72），并确定 1) 易于聚集的蛋白质靶标以及它们是否在细胞类型中常见；2) 给定细胞类型内的蛋白质错误折叠倾向是否与伴侣相互作用、表达水平和共享生化特性等特征相关。我们的第三个目标是用新的伴侣底物（通常在我们的细胞类型中不表达）来挑战细胞，并比较底物折叠调节的结果，包括参与这些过程的伴侣。 我们的研究将提供关于伴侣网络、其底物及其对抗蛋白质折叠应力的弹性之间关系的原理的基本见解。