Role of Thrombospondins in interstitial Extracellular Matrix 3-Dimensional Organisation: Mechanisms and Functions

血小板反应蛋白在间质细胞外基质三维组织中的作用：机制和功能

• 批准号: MR/K018043/1
• 负责人: Josephine Adams
• 金额: $ 97.16万
• 依托单位: University of Bristol
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2013
• 资助国家: 英国
• 起止时间: 2013 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FK018043%2F1
• 关键词: Role; Thrombospondins; interstitial; Extracellular; Matrix

In our bodies, our skin is supple and our bones are rigid because of the properties of extracellular matrix (ECM) that surrounds the cells in each tissue. ECM is built up from the beginning of development as a suite of large proteins released by cells that bind with each other to form insoluble fibrils and networks. These structures are maintained throughout life, are repaired after injury, and can cause health problems if disrupted in disease. The physical and chemical properties of ECM inform cells whether to be stationary, move, or divide, and modulate the expression of genes. These signals enable cells to function properly throughout life but how the 3-dimensional organisation of ECM is set up and tuned is poorly understood. For all these reasons, research into principles of ECM organisation is important and necessary to identify new therapeutic targets and materials. The most abundant proteins in ECM are collagens that form elongated fibrils. Many accessory proteins act to maintain the number, sizes, and packing of collagen fibrils, and how the fibrils interact with other ECM components. Accessory proteins are attractive as possible therapeutic targets because they are less abundant (so could be more easy to modulate with a small drug) and yet can act as "levers" to change ECM organisation. Our recent research identified that thrombospondin proteins (TSPs) are conserved with collagens and a few other ECM components throughout animals, from sponges to human. Because of this conservation we consider that the mechanism(s) by which TSPs bind into ECM and modulate collagen fibril organisation are likely to be important and a promising therapeutic target. In studying how TSPs bind into ECM, we identified a protein sequence motif that is widely conserved between TSPs and is required for them to accumulate into ECM made by cultured cells. TSPs mutated at this site are secreted normally by cells but are not held in the ECM. We discovered recently that this site works by mediating interactions between separate TSP molecules. These interactions cause TSP clusters to form small dot-like structures or "puncta". From this research we developed a model, (a hypothetical diagram), of how TSPs can enter and cluster in ECM to impact on collagen fibril organisation. Our model is novel because neither the molecular identity of the motif or its role in interactions between TSP molecules were known before. Our central goals are: 1), To further understand mechanisms of TSPs within cell culture ECM, to extend the model and strengthen its accuracy. 2), To test how the site we have identified in cell culture works in living animals. To make this test is essential to understand how our discoveries in cell culture relate to protein function in real tissues. With the mouse we can examine the role of the TSP interaction site in the skin of an animal close to human anatomy and physiology that is used widely in biomedical research. 3). From the new genetic strains of mice we will also isolate cells from the skin so that we can closely study their properties in culture and relate the mechanisms that we identify in goal 1 to effects of TSPs on cell functions and ECM properties. These experiments will generate additional materials from the experimental mice and help us understand how our new knowledge might be adapted for clinical materials.

在我们的身体里，我们的皮肤是柔软的，我们的骨骼是坚硬的，这是因为包围着每个组织中细胞的细胞外基质(ECM)的特性。细胞外基质是从发育开始就建立起来的，是细胞释放的一套大蛋白，它们相互结合形成不溶的纤维和网络。这些结构在一生中都会得到维护，在受伤后会得到修复，如果在疾病中受到破坏，可能会导致健康问题。细胞外基质的物理和化学性质告诉细胞是静止的、移动的还是分裂的，并调节基因的表达。这些信号使细胞在整个生命过程中都能正常运作，但人们对细胞外基质的三维组织是如何建立和调整的知之甚少。因此，对ECM组织原理的研究对于确定新的治疗靶点和材料是重要和必要的。细胞外基质中最丰富的蛋白质是形成细长纤维的胶原蛋白。许多辅助蛋白的作用是维持胶原纤维的数量、大小和包装，以及纤维与其他细胞外基质成分的相互作用。辅助蛋白作为可能的治疗靶点很有吸引力，因为它们不那么丰富(因此可能更容易用一种小药物调节)，但可以充当改变细胞外基质组织的“杠杆”。我们最近的研究发现，从海绵到人类，凝血酶原蛋白(TSPs)与胶原蛋白和其他一些ECM成分在动物中都是保守的。由于这种保守性，我们认为TSP与细胞外基质结合并调节胶原纤维组织的机制(S)可能是重要的，也是一个有前景的治疗靶点。在研究TSP如何与细胞外基质结合的过程中，我们发现了一个在TSP之间广泛保守的蛋白质序列基序，它是TSP积累到培养细胞产生的ECM所必需的。在该位点突变的TSP由细胞正常分泌，但不存在于ECM中。我们最近发现，这个站点通过调节不同TSP分子之间的相互作用来发挥作用。这些相互作用导致TSP集群形成小的点状结构或“点状”。通过这项研究，我们开发了一个模型(假设图)，说明TSP如何进入并聚集在ECM中，从而影响胶原原纤维组织。我们的模型是新颖的，因为以前既不知道该基序的分子同一性，也不知道它在TSP分子之间相互作用中的作用。我们的中心目标是：1)进一步了解TSP在细胞培养ECM中的作用机制，扩展模型并增强其准确性。2)，测试我们在细胞培养中确定的位置在活动物中是如何工作的。进行这项测试对于了解我们在细胞培养中的发现如何与真实组织中的蛋白质功能有关是至关重要的。通过小鼠，我们可以研究TSP相互作用部位在动物皮肤中的作用，这种作用与人体解剖学和生理学密切相关，在生物医学研究中被广泛使用。3)。我们还将从新的小鼠遗传品系中分离皮肤细胞，以便我们能够密切研究它们在培养中的特性，并将我们在目标1中确定的机制与TSP对细胞功能和细胞外基质特性的影响联系起来。这些实验将从实验小鼠身上产生额外的材料，并帮助我们了解我们的新知识如何适用于临床材料。

项目成果

Modulation of the extracellular matrix patterning of thrombospondins by actin dynamics and thrombospondin oligomer state.

• DOI: 10.1042/bsr20140168
• 发表时间: 2015-05-20
• 期刊: Bioscience reports
• 影响因子: 4
• 作者: Hellewell AL;Gong X;Schärich K;Christofidou ED;Adams JC
• 通讯作者: Adams JC

A Rapid, Scalable Method for the Isolation, Functional Study, and Analysis of Cell-derived Extracellular Matrix.

• DOI: 10.3791/55051
• 发表时间: 2017-01-04
• 期刊: Journal of visualized experiments : JoVE
• 作者: Hellewell AL;Rosini S;Adams JC
• 通讯作者: Adams JC

Intermolecular interactions of thrombospondins drive their accumulation in extracellular matrix.

• DOI: 10.1091/mbc.e14-05-0996
• 发表时间: 2015-07-15
• 期刊: Molecular biology of the cell
• 影响因子: 3.3
• 作者: Kim DJ;Christofidou ED;Keene DR;Hassan Milde M;Adams JC
• 通讯作者: Adams JC

Emergence of a Thrombospondin Superfamily at the Origin of Metazoans

• DOI: 10.1093/molbev/msz060
• 发表时间: 2019-03
• 期刊: Molecular Biology and Evolution
• 影响因子: 10.7
• 作者: D. Shoemark;B. Ziegler;Hiroshi Watanabe;Jennifer Strompen;R. Tucker;S. Özbek;J. Adams