Determination of the mechanisms of desmosome loss during EMT

EMT 过程中桥粒丢失机制的确定

• 批准号: BB/R001707/1
• 负责人: Christoph Ballestrem
• 金额: $ 60.51万
• 依托单位: University of Manchester
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2018
• 资助国家: 英国
• 起止时间: 2018 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FR001707%2F1
• 关键词: Determination; mechanisms; desmosome; loss; during

Cells in stress-exposed tissues, e.g. heart muscle and the coverings of body surfaces, are bound together by tiny, rivet-like structures called desmosomes. Aberrant function of these structures causes diseases such as sudden heart failure, defective wound healing, cancer spread and certain blistering diseases of the skin and oral cavity. Some of these conditions are among the most common causes of morbidity and death, while others are rare but extremely unpleasant, difficult to treat and can be fatal. Desmosomes are also important for normal development, where they stabilise developing tissues. It is therefore essential to understand how desmosome function is regulated.We have shown that an important factor contributing to the toughness of tissues is that desmosomes exhibit a highly adhesive state known as hyper-adhesion. Hyper-adhesion is important for tissue strength, but also locks cells together, thus restricting their movement. During wound healing, epidermal cells migrate to close the wound, their migration being triggered by wounding. The invasive spread of cancer cells also requires cell migration and in development cell movement generates the correct architecture of tissues. In order to move, cells need to reduce the degree of adhesion between them. We have shown that on wounding desmosomes rapidly lose hyper-adhesion, becoming more weakly adhesive. However, this weakening of adhesion may not be sufficient to permit the adequate movement. Instead, cells may need to lose some or all of their desmosomes. How they do this is not understood.Some evidence from electron microscopy studies of cancers and wounds suggests cells may be able to engulf whole desmosomes and therefore become stuck together more loosely. This is evident because desmosomes have a characteristic, easily recognisable structure. Normally desmosomes appear at the junction between cells but these studies have shown whole desmosome inside cells, as though one cell has "eaten" the desmosome! However unlikely this seems we have now induced cell separation in culture and shown that they do indeed engulf whole desmosomes. This is exciting because it enables us to investigate the mechanism behind a process that occurs in normal and diseased tissues.Desmosome engulfment resembles a process called phagocytosis whereby cells of the immune system engulf extracellular particles, e.g. bacteria. Phagocytosis requires active contractile activity by the engulfing cell so as to surround the particle and draw it inside. This, in turn, requires the action of the cell's contractile apparatus, which depends upon filamentous proteins called actin and myosin, similar to those involved in muscle contraction. Desmosomes are not normally associated with these proteins but instead are internally linked to other filaments called intermediate filaments (IF). The IF are linked from cell-to-cell by desmosomes, forming a scaffold that gives great strength to tissues. However, IF possess no contractile activity. So if the filaments they attach to cannot contract, how are desmosomes engulfed? Our pilot studies suggest the cell's contractile apparatus is somehow involved in desmosome engulfment and a regulatory enzyme called protein kinase C (PKC), known to be involved in phagocytosis and actin regulation, participates. To understand the mechanism in greater detail we will use state-of the-art microscopy to study how desmosomes become associated with the contractile machinery as they switch from hyper-adhesion to engulfment. Also we will use mass spectrometry to identify novel proteins involved in the process and to determine the role of PKC. Finally, we will use molecular cell biology to determine the functions of the key proteins in detail.Our results will both further our understanding of normal development and provide a basis for new therapies for major health problems such as chronic wounds, skin blistering diseases and, potentially, for limiting the spread of cancer.

暴露于压力的组织中的细胞，例如心肌和体表覆盖物，通过称为桥粒的微小铆钉状结构结合在一起。这些结构的异常功能会导致疾病，如突发性心力衰竭、伤口愈合不良、癌症扩散和某些皮肤和口腔起泡疾病。其中一些情况是发病和死亡的最常见原因，而另一些情况则很少见，但非常令人不快，难以治疗，可能是致命的。桥粒对正常发育也很重要，它们稳定发育中的组织。因此，了解桥粒功能是如何调节的至关重要。我们已经证明，有助于组织韧性的一个重要因素是桥粒表现出高度粘附状态，称为超粘附。超粘附对组织强度很重要，但也会将细胞锁定在一起，从而限制它们的运动。在伤口愈合期间，表皮细胞迁移以闭合伤口，它们的迁移由创伤触发。癌细胞的侵入性扩散也需要细胞迁移，并且在发育中细胞运动产生正确的组织结构。为了移动，细胞需要降低它们之间的粘附程度。我们已经表明，在创伤桥粒迅速失去超粘附，变得更弱的粘附。然而，这种粘附力的减弱可能不足以允许适当的移动。相反，细胞可能需要失去部分或全部桥粒。目前还不清楚它们是如何做到这一点的，一些来自癌症和伤口的电子显微镜研究的证据表明，细胞可能能够吞噬整个桥粒，因此变得更松散地粘在一起。这是显而易见的，因为桥粒具有特征性的、易于识别的结构。正常情况下桥粒出现在细胞之间的连接处，但这些研究表明整个桥粒都在细胞内，就好像一个细胞“吃掉”了桥粒一样！尽管这看起来不太可能，但我们现在已经在培养中诱导细胞分离，并表明它们确实吞噬整个桥粒。这是令人兴奋的，因为它使我们能够研究正常和病变组织中发生的过程背后的机制。桥粒吞噬类似于一种称为吞噬作用的过程，免疫系统的细胞通过吞噬细胞外颗粒，例如细菌。吞噬作用需要吞噬细胞的主动收缩活动，以便包围颗粒并将其吸入。这反过来又需要细胞收缩装置的作用，收缩装置依赖于称为肌动蛋白和肌球蛋白的丝状蛋白质，类似于参与肌肉收缩的那些蛋白质。桥粒通常不与这些蛋白质结合，而是与称为中间丝（IF）的其他纤维内部连接。IF通过桥粒在细胞与细胞之间连接，形成支架，为组织提供巨大的强度。然而，IF没有收缩活动。那么，如果桥粒附着的纤维不能收缩，那么桥粒又是如何被吞噬的呢？我们的初步研究表明，细胞的收缩装置在某种程度上参与了桥粒吞噬，一种称为蛋白激酶C（PKC）的调节酶参与了吞噬作用和肌动蛋白调节。为了更详细地了解机制，我们将使用最先进的显微镜来研究桥粒如何与收缩机制相关联，因为它们从超粘附转变为吞噬。我们还将使用质谱法来鉴定参与该过程的新蛋白质，并确定PKC的作用。最后，我们将使用分子细胞生物学来详细确定关键蛋白质的功能。我们的结果将进一步加深我们对正常发育的理解，并为主要健康问题（如慢性伤口，皮肤起泡疾病）的新疗法提供基础，并可能限制癌症的传播。

项目成果

Desmosomal dualism: the core is stable while plakophilin is dynamic

桥粒二元性：核心是稳定的，而亲斑蛋白是动态的

• DOI: 10.1101/2021.03.02.433631
• 发表时间: 2021
• 作者: Fülle J