A new role for intermediate filaments in the secretory pathway

中间丝在分泌途径中的新作用

• 批准号: BB/T000945/1
• 负责人: Martin Lowe
• 金额: $ 61.31万
• 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: 2019
• 资助国家: 英国
• 起止时间: 2019 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FT000945%2F1
• 关键词: new; role; intermediate; filaments; secretory

Proteins are major building blocks within our cells, and are also secreted from our cells to mediate intercellular communication, as well as generating the extracellular matrix to which cells attach within our bodies. This matrix is vital for tissue formation and maintenance, and provides mechanical strength, which is especially apparent in bone and tendon. Cells therefore constantly make and secrete proteins via a process that can be regulated in response to various external influences, including the mechanical environment of the cell. Approximately one third of all cellular proteins, including those proteins secreted to the cell exterior, are made within the secretory pathway. This pathway is comprised of various compartments through which cargo proteins are sequentially transported, receiving functionally important modifications as they do so. The compartment at the centre of the secretory pathway is the Golgi apparatus, which serves as a hub for transport, as well as being a major site of cargo modification. The organization of the secretory pathway and movement of proteins between secretory compartments, including the Golgi apparatus, is dependent upon a fibrous network, generally referred to as the cytoskeleton. Defects in secretory pathway function and/or cytoskeleton organization can cause disease, highlighting the importance of these processes for human health. The focus of this study is a Golgi resident protein called GORAB, whose mutation causes a skin and bone disorder in humans. We have shown that GORAB forms discrete structures called domains at the Golgi membrane, that this process is important for cargo modification at the Golgi, and that its loss affects assembly of the extracellular matrix. More recently, we have found a novel interaction between GORAB and a type of cytoskeletal structure called cytoplasmic intermediate filaments. Although intermediate filaments are very well studied and known to be important for cell integrity, we know very little about how they might affect the organization or function of the secretory pathway. Our identification of a novel association of the Golgi apparatus with intermediate filaments suggests an important role for this interaction in secretory pathway organization and function. In this proposal we aim to identify how GORAB forms domains at the Golgi membrane, how the assembly of domains is controlled, and how the domains bind to cytoplasmic intermediate filaments. A major goal is then to determine the functional significance of the GORAB-intermediate filament interaction, testing our hypothesis that it is important for Golgi organization and function, with mechanistic experiments to determine how this occurs. Another key aspect will be to determine how the secretory pathway, and the Golgi apparatus in particular, responds to changes in the mechanical environment of the cell, which is vital for the correct production of proteins within the secretory pathway. Because cytoplasmic intermediate filaments play an important mechanical role within cells, we will test the extent to which they directly couple changes in the mechanical environment of cells to the functional organization of the Golgi apparatus, focusing on the interaction with GORAB, but also exploring other possible mechanisms at play. The work will be important for determining how the secretory pathway is organized, how intermediate filaments interact with this pathway, and how the secretory pathway is coupled to its mechanical environment, all of which are fundamentally important for human health.

蛋白质是我们细胞内的主要组成部分，也是从我们的细胞中分泌出来的，以调节细胞间的沟通，以及产生细胞在我们体内附着的细胞外基质。这种基质对组织的形成和维持至关重要，并提供机械强度，这在骨骼和肌腱中尤为明显。因此，细胞不断地通过一个过程来制造和分泌蛋白质，这个过程可以受到各种外部影响的调节，包括细胞的机械环境。大约三分之一的细胞蛋白质，包括那些分泌到细胞外部的蛋白质，是在分泌途径内制造的。这一途径由不同的隔间组成，货物蛋白通过这些隔间依次运输，在运输过程中接受重要的功能修饰。位于分泌途径中心的隔室是高尔基体，它是运输的枢纽，也是货物修饰的主要部位。分泌途径的组织和蛋白质在包括高尔基体在内的分泌室之间的运动依赖于纤维网络，通常被称为细胞骨架。分泌途径功能和/或细胞骨架组织的缺陷可导致疾病，突出了这些过程对人类健康的重要性。这项研究的重点是一种名为Gorab的高尔基体驻留蛋白，它的突变会导致人类的皮肤和骨骼疾病。我们已经证明，Gorab在高尔基体膜上形成称为结构域的离散结构，这一过程对于高尔基体的货物修饰是重要的，并且它的损失影响细胞外基质的组装。最近，我们发现了Gorab和一种叫做细胞质中间丝的细胞骨架结构之间的一种新的相互作用。尽管人们对中间丝进行了很好的研究，并且知道中间丝对细胞的完整性很重要，但我们对它们如何影响分泌途径的组织或功能知之甚少。我们发现了高尔基体与中间丝之间的一种新的联系，这表明这种相互作用在分泌途径、组织和功能中扮演着重要的角色。在这个建议中，我们的目标是确定Gorab如何在高尔基膜上形成结构域，如何控制结构域的组装，以及结构域如何与细胞质中间丝结合。然后，一个主要的目标是确定Gorab-中间丝相互作用的功能意义，验证我们的假设，即它对高尔基体的组织和功能是重要的，并通过机械实验来确定这是如何发生的。另一个关键方面将是确定分泌途径，特别是高尔基体如何对细胞机械环境的变化做出反应，这对分泌途径中蛋白质的正确产生至关重要。由于细胞质中间丝在细胞内扮演着重要的机械角色，我们将测试它们在多大程度上直接将细胞机械环境的变化与高尔基体的功能组织联系起来，重点放在与Gorab的相互作用上，同时也探索其他可能发挥作用的机制。这项工作对于确定分泌途径是如何组织的，中间丝如何与这一途径相互作用，以及分泌途径如何与其机械环境相耦合将是重要的，所有这些对人类健康都是至关重要的。

项目成果

Vimentin intermediate filaments provide structural stability to the mammalian Golgi complex.

• DOI: 10.1242/jcs.260577
• 发表时间: 2023-10-15
• 期刊: JOURNAL OF CELL SCIENCE
• 影响因子: 4
• 作者: Vitali, Teresa;Sanchez-Alvarez, Rosa;Witkos, Tomasz M.;Bantounas, Ioannis;Cutiongco, Marie F. A.;Dudek, Michal;Yan, Guanhua;Mironov, Alexander A.;Swift, Joe;Lowe, Martin
• 通讯作者: Lowe, Martin

Immunofluorescence Microscopy of the Mammalian Golgi Apparatus.

哺乳动物高尔基体的免疫荧光显微镜。

• DOI: 10.1007/978-1-0716-2639-9_8
• 发表时间: 2023
• 期刊: Methods in molecular biology (Clifton, N.J.)
• 作者: Arab M

GOLPH3 keeps the Golgi residents at home.

• DOI: 10.1083/jcb.202108147
• 发表时间: 2021-10-04
• 期刊: The Journal of cell biology
• 作者: Lowe M