Defining and Modulating Mechanisms of Collagen Proteostasis

作为骨骼、皮肤、软骨、基底膜等的主要蛋白质成分， 胶原蛋白是动物生命的分子支架。由于联合国系统的高度等级性质， 胶原超分子支架的性质基本上由细胞外基质的性质定义。 复杂的细胞内胶原蛋白折叠过程和质量控制。因此，毫不奇怪， 细胞内胶原蛋白的蛋白质沉积引起称为胶原病的多种疾病。这些缺陷 最常见的是由胶原基因中的常染色体显性突变引起的，并且可以是各种各样的。 归因于三个主要问题：（1）错误折叠或功能障碍的胶原蛋白链逃逸到 细胞外基质;（2）适当折叠的胶原蛋白分泌不足;和/或（3）细胞内积累 胶原蛋白分子的错误折叠导致慢性细胞功能障碍。所有这三个缺陷都与 随着内质网（ER）蛋白质稳定网络（一个高度整合的系统）的失败， 分子伴侣、质量控制机制和分泌机制）来适当地解决胶原蛋白的产生 问题，特别是在导致疾病的突变的背景下。阐明分子机制 因此，ER中胶原蛋白的蛋白质稳态对于疾病的发展至关重要， 改良疗法为此，目前的提案旨在回答三个关键问题：（1）胶原蛋白能否 通过合理的化学生物学调节ER蛋白抑制网络来挽救蛋白抑制缺陷？（二） ER质量控制机构如何识别错误折叠的胶原蛋白链？(3)胶原蛋白怎么样 组装是胶原蛋白折叠的关键第一步，对野生型胶原蛋白和 导致疾病的胶原蛋白变异在具体目标1中，最先进的化学生物学策略 针对ER蛋白质稳态网络和未折叠的蛋白质反应进行部署，以检验假设 与疾病相关的胶原蛋白蛋白质稳态缺陷可以通过蛋白质稳态网络调节来解决。在 具体目标2，胶原蛋白质量控制的机制（已知存在，但仍不明确）将 对于野生型和一系列错误折叠的胶原蛋白变体，可以进行详细研究。这一目标涉及质量 基于光谱的定量比较相互作用组学来检测这些机制，然后 生物化学验证和表征。在特定目标3中，胶原蛋白组装的分子代码将是 定义，并通过蛋白质稳态网络解决装配缺陷的策略将被追求。见解 使用这种生物化学和细胞和化学生物学实验策略的组合获得， 预计会产生积极的和最终可翻译的影响，因为它们很有可能产生新的 其他实验方法无法达到的各种胶原病治疗干预的靶点 接近。

As the primary proteinaceous component of bone, skin, cartilage, basement membranes, and more, collagen serves as the molecular scaffold for animal life. Owing to the highly hierarchical nature of the extracellular matrix, the properties of collagenous supramolecular scaffolds are fundamentally defined by the complex intracellular process of collagen folding and quality control. Unsurprisingly, therefore, defects in intracellular collagen proteostasis engender diverse diseases known as the collagenopathies. These defects are most commonly caused by autosomal dominant mutations in collagen genes, and can be variously ascribed to three primary issues: (1) Escape of misfolded or dysfunctional collagen strands into the extracellular matrix; (2) Insufficient secretion of properly folded collagen; and/or (3) Intracellular accumulation of misfolding collagen molecules that leads to chronic cell dysfunction. All three of these defects are associated with a failure of the endoplasmic reticulum's (ER's) proteostasis network (a highly integrated system of chaperones, quality control mechanisms, and secretory machineries) to properly solve the collagen production problem, particularly in the context of mutations that lead to disease. Elucidating molecular mechanisms of collagen proteostasis in the ER is therefore of paramount importance to enable the development of disease- modifying therapies. To this end, the current proposal aims to answer three key questions: (1) Can collagen proteostasis defects by rescued by rational chemical biologic modulation of the ER proteostasis network? (2) How is a misfolding collagen strand identified by the ER quality control machinery? (3) How is collagen assembly, which is the critical first step in collagen folding, regulated both for wild-type collagen and for misfolding, disease-causing collagen variants? In Specific Aim 1, state-of-the-art chemical biology strategies targeted at the ER proteostasis network and the unfolded protein response are deployed to test the hypothesis that disease-associated collagen proteostasis defects can be resolved by proteostasis network modulation. In Specific Aim 2, the mechanisms of collagen quality control (which are known to exist but remain ill-defined) will be studied in detail, both for wild-type and a range of misfolding collagen variants. This Aim involves mass spectrometry-based quantitative comparative interactomics to detect such mechanisms, followed by biochemical validation and characterization. In Specific Aim 3, the molecular code for collagen assembly will be defined, and strategies to address assembly defects via the proteostasis network will be pursued. Insights obtained using this combination of biochemical and cell and chemical biological experimental strategies are expected to have a positive and ultimately translatable impact, because they are highly likely to yield new targets for therapeutic intervention in diverse collagenopathies that are not accessible by other experimental approaches.