Proteome-Driven Holistic Reconstruction of Organ-Wide Multi-Scale Networks

摘要 人类的器官，如大脑，复杂得惊人。它们由成百上千个独立的 功能区，每个功能区包含相当数量的不同细胞类型和无数分子。 了解这些多尺度组件如何协同工作以生成系统级响应 对于生物学的许多领域来说都是必不可少的，但这一领域的进展受制于流行的 将生物系统分成已知的细胞类型，然后分别研究每个种群。虽然 这种强大的简约主义方法使得在多个层面上审问复杂的相互作用变得困难- 分子(如蛋白质)、亚细胞(如突触)、细胞和区域水平。此外，这种方法可能会 忽略许多潜在的重要但未确定的功能网络。我们无法彻底识别多个 扩展功能网络并询问其系统范围内的多因素交互作用限制了我们的能力 了解复杂生物系统的功能和功能障碍。在这里，我们的目标是从根本上 通过开发尖端的平台，将我们的方法从简约主义转变为整体主义 全器官多尺度网络的蛋白质组学重建。利用小鼠和人类的临床样本 作为我们的模型，我们将开发四个广泛适用的跨学科平台，将 化学和材料工程技术。这些平台将实现：(1)可扩展的组织转化 变成一个坚不可摧的、含有蛋白质组的三维(3D)框架；(2)无限轮的 单个完整组织的分子表型与多个数据集的精确体积共配准；(3) 通过在器官范围内同步靶-探针结合反应，快速、可扩展和统一的组织标记；(4) 完整器官的超分辨率蛋白质组成像。如果成功，我们提议的工作将使蛋白质组- 完整生物系统的驱动整体重建和高维定量表型 史无前例的分辨率。使用技术平台和人脑有机模型，既健康又 我们将调查以下基本问题：(Q1)存在多少种细胞类型/区域 大脑器官的不同发育阶段？(Q2)这些单元和区域如何形成网络？(第三季度) 亚细胞成分、单个细胞、回路和区域的蛋白质组状态在整个过程中如何变化 发展。(Q4)细胞的形态特征如何变化？(Q5)疾病患者的Q1-4是如何改变的 有机化合物？这项研究可能为理解人体器官在健康和健康中的发育提供新的见解 疾病。

Abstract Human organs such as the brain are stunningly complex. They consist of hundreds to thousands of separate functional areas, each containing a comparable number of distinct cell types and innumerable molecules. Understanding how these multi-scale components work together to generate systems-level responses is essential for many fields of biology, but advancement in this area is hampered by the prevalent methodology of dividing biological systems into known cell types and then separately studying each population. Although powerful, this reductionistic approach makes it difficult to interrogate complex interactions at multiple levels — molecular (e.g., proteins), subcellular (e.g., synapses), cellular, and area level. Moreover, this approach could ignore many potentially important but unidentified functional networks. Our inability to thoroughly identify multi- scale functional networks and interrogate their system-wide, multifactorial interactions has limited our ability to understand the function and dysfunction of complex biological systems. Here, we aim to fundamentally transform our approach from a reductionistic to a holistic one by developing cutting-edge platforms for proteomic reconstruction of organ-wide multi-scale networks. Using murine and human clinical samples and organoids as our models, we will develop four broadly applicable cross-disciplinary platforms that integrate chemical and material engineering technologies. These platforms will enable: (1) scalable tissue transformation into an indestructible, proteome-containing three-dimensional (3D) framework; (2) unlimited rounds of molecular phenotyping of a single intact tissue with precise volume co-registration of multiple datasets; (3) rapid, scalable, and uniform tissue labeling by synchronizing target-probe binding reactions organ-wide; (4) superresolution proteomic imaging of intact organs. If successful, our proposed work will enable proteome- driven holistic reconstruction and high-dimensional quantitative phenotyping of intact biological systems at unprecedented resolution. Using the technology platforms and human brain organoid models, both healthy and diseased, we will investigate the following fundamental questions: (Q1) How many cell types/regions exist at different developmental stages of the brain organoids? (Q2) How these cells and regions form networks? (Q3) How proteomic states of subcellular components, individual cells, circuits, and regions change throughout the development. (Q4) How morphological features of cells change? (Q5) how Q1-4 are altered in diseased organoids? This study may provide new insights into understanding human organ development in health and disease.