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Understanding mechanisms of coat assembly and membrane deformation.

了解涂层组装和膜变形的机制。

基本信息

批准号：
BB/T002670/1
负责人：
Giulia Zanetti
金额：
$ 66.92万
依托单位：
Birkbeck, University of London
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2020
资助国家：
英国
起止时间：
2020 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FT002670%2F1
关键词：
Understanding mechanisms coat assembly membrane

项目摘要

Cells can be likened to cities to describe their structure and organisation. The cell's citizens are the proteins, which carry out all the activities necessary to maintain cell homeostasis and ensure survival and proliferation.Eukaryotic cells are organised in regions delimited by membranes which are called compartments. These can be thought of as precisely defined neighbourhoods where particular activities take place. For example an organelle called the endoplasmic reticulum (shortened to ER) is where certain classes of proteins are made, and where the initial checks on their health are performed. Another important organelle is the Golgi: this is where proteins mature into their fully functional form and are sorted to where they'll perform their job.As in cities, proteins need to be transported from one compartment to another - a fundamental aspect that is necessary to maintain cell functionality. Because cell compartments are delimited by membranes, communication between compartments and exchange of material happens through vesicles: small sacs of membrane bud and pinch off from the originating compartment loaded with cargo, to then release it to the target compartment. The transport link that connects the ER to the Golgi, to get young proteins from where they were born to where they will fully mature is called COPII. COPII is a set of proteins which aid formation of vesicles from the ER by assembling into curved scaffolds around the ER membrane leading it to bud a vesicle (COPII thereby forms a so-called "coat"). While deforming the ER membrane, COPII also links with proteins inside the ER that need to be transported (cargo), so that these are incorporated in the forming vesicle. Deformation of the ER membrane follows the coat shape.We have a good functional understanding of COPII mechanisms, but our view of the coat in action on membranes is somewhat fuzzy. In particular we do not understand to the fullest detail what are the interactions between individual components that determine the shape of the coat, and therefore how the ER membrane will be deformed. This is important because the shape of the transport vesicle must be adapted to the type of cargo: COPII relies in fact on other proteins (regulators) to help form different shapes which are required for different cargoes. When individual coat components have some defects the transport system fails to do its job efficiently. In the worst cases this leads to dire consequences, with cells unable to survive. Sometimes, COPII defects only affect its ability to transport certain categories of cargo proteins: cells survive but they don't function in their every aspect, leading to genetic disease. For example, some mutations in COPII hamper ER exit of collagen precursors, leading to defects in development of skeletal and connective tissues.In order to understand the mechanisms that link assembly of coat components to determination of coat and membrane shape, and ability to transport all cargoes, we need to obtain a much clearer (higher resolution) picture of the COPII coat "in action".Cryo-electron tomography is a technique which allows 3D visualisation of biological specimens in near-native conditions. The resolution of this technique, especially when associated to in depth image processing, has leaped forward in the last few years, and today allows for views of biological events such as COPII budding which are a lot less "fuzzy" than before. In my lab, we have recently obtained one of the highest resolution reconstructions of part the coat. In this project, we aim to complete the picture by tackling the molecular interactions between coat components, and we also aim to study assemblies formed in increasingly physiological contexts. This will allow us to understand with the clearest molecular view how COPII assembled to form membrane-deforming scaffolds of various types, and how regulators can interact with COPII to modulate this process.

细胞可以被比作城市来描述它们的结构和组织。细胞的公民是蛋白质，它们执行维持细胞稳态并确保生存和增殖所需的所有活动。真核细胞组织在由称为隔室的膜界定的区域中。这些可以被认为是精确定义的社区，在那里进行特定的活动。例如，一种被称为内质网（简称ER）的细胞器是制造某些类型蛋白质的地方，也是对它们的健康进行初步检查的地方。另一个重要的细胞器是高尔基体：蛋白质在高尔基体中发育成熟，形成完整的功能形式，并被分类到它们将要执行任务的地方。就像在城市中一样，蛋白质需要从一个隔间运输到另一个隔间--这是维持细胞功能所必需的基本方面。由于细胞隔室由膜界定，隔室之间的通信和物质交换通过囊泡发生：膜的小囊芽和夹断从装载货物的起源隔室，然后将其释放到靶隔室。连接内质网和高尔基体的运输环节，将年轻的蛋白质从它们出生的地方运送到它们将完全成熟的地方，称为COPII。COPII是一组蛋白质，其通过组装成围绕ER膜的弯曲支架来帮助从ER形成囊泡，从而导致其出芽成囊泡（COPII由此形成所谓的“包衣”）。在使ER膜变形的同时，COPII还与ER内需要运输的蛋白质（货物）连接，使得这些蛋白质被并入形成的囊泡中。我们对COPII机制有了很好的功能理解，但是我们对膜上的涂层作用的看法有些模糊。特别是，我们不完全了解决定涂层形状的各个成分之间的相互作用，因此ER膜将如何变形。这很重要，因为运输囊泡的形状必须适应货物的类型：COPII实际上依赖于其他蛋白质（调节剂）来帮助形成不同货物所需的不同形状。当个别涂层组件存在一些缺陷时，运输系统就无法有效地完成其工作。在最坏的情况下，这会导致可怕的后果，细胞无法存活。有时，COPII缺陷只影响其运输某些类别货物蛋白的能力：细胞存活，但它们不能在各个方面发挥作用，导致遗传疾病。例如，COPII中的一些突变阻碍了胶原前体的ER退出，导致骨骼和结缔组织发育的缺陷。为了理解将被毛组分的组装与决定被毛和膜形状以及运输所有货物的能力联系起来的机制，我们需要得到一个更清晰的（更高分辨率）的COPII涂层“在行动中”的图片。冷冻电子断层扫描是一种技术，它允许在近距离内对生物标本进行3D可视化，原生条件。这种技术的分辨率，特别是当与深度图像处理相关联时，在过去几年中已经向前飞跃，并且今天允许生物事件的视图，例如COPII萌芽，这比以前少了很多“模糊”。在我的实验室里，我们最近获得了一个最高分辨率的部分外套重建。在这个项目中，我们的目标是通过解决外壳组件之间的分子相互作用来完成这一过程，我们还旨在研究在越来越多的生理环境中形成的组件。这将使我们能够以最清晰的分子视角了解COPII如何组装形成各种类型的膜变形支架，以及调控因子如何与COPII相互作用以调节这一过程。

项目成果

期刊论文数量（8）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Structure of the complete, membrane-assembled COPII coat reveals a complex interaction network.

DOI：
10.1038/s41467-021-22110-6
发表时间：
2021-04-01
期刊：
Nature communications
影响因子：
16.6
作者：
Hutchings J;Stancheva VG;Brown NR;Cheung ACM;Miller EA;Zanetti G
通讯作者：
Zanetti G

A Bayesian approach to single-particle electron cryo-tomography in RELION-4.0.

DOI：
10.7554/elife.83724
发表时间：
2022-12-05
期刊：
eLife
影响因子：
7.7
作者：
Zivanov J;Otón J;Ke Z;von Kügelgen A;Pyle E;Qu K;Morado D;Castaño-Díez D;Zanetti G;Bharat TAM;Briggs JAG;Scheres SHW
通讯作者：
Scheres SHW

A multivalent fuzzy interface drives reversible COPII coat assembly

多价模糊界面驱动可逆 COPII 涂层组装