High-resolution imaging and time-resolved proteomic profiling of COPII-dependent procollagen packaging.

COPII 依赖性前胶原包装的高分辨率成像和时间分辨蛋白质组学分析。

• 批准号: MR/P000177/1
• 负责人: David Stephens
• 金额: $ 58.94万
• 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: 2016
• 资助国家: 英国
• 起止时间: 2016 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FP000177%2F1
• 关键词: resolution; imaging; time; resolved; proteomic

Collagen is the most abundant protein in our bodies. It is also one of the largest. This size presents a significant problem to cellular machinery involved in shipping procollagen from its site of synthesis (inside the endoplasmic reticulum of cells) to its site of action in the extracellular matrix. The machinery that controls the first step in this pathway normally generates transport carriers that are simply too small to accommodate procollagen. Nearly all cell types secrete one or more types of collagen and this process is absolutely essential during development to provide the extracellular matrix within which cells migrate to their final position and differentiate. It is also essential for those very differentiation processes. Later in life ongoing secretion of collagen is vital for tissues to maintain themselves by renewal and regeneration. Major pathologies are also associated with defects in collagen secretion including fibrosis. The field of stem cell biology increasingly relies on accurate reproduction of the "stem cell niche", of which the cohort of secreted collagens is a major component. The development and exploitation of engineered cells and tissue for therapeutic benefit will in many cases rely on a complete understanding of these processes. Consequently it is perhaps surprising how little we know about this process. The machinery that directs the first step in the secretion of small proteins (for which a 2013 Nobel Prize was awarded) has been very well characterised. This machinery generates small (60-80 nanometre) transport vesicles that are not sufficient to encapsulate the ~300 nanometre size of collagen molecules. While there have been some advances in this area, they either relate to specific collagen types (such as collagen VII) or are underdeveloped in terms of our understanding (such as modification of the core machinery using a small protein called ubiquitin). Our work aims to use a combination of cutting-edge imaging and proteomics methods to precisely define this machinery, and the nature of these transport carriers exactly at the point of procollagen export. Importantly we will focus on two major procollagen isotypes - I and II to provide insight into the pathway for these forms that are of major clinical interest. We have generated new tools to examine this process with unprecedented spatial and temporal precision. As such we anticipate making major advances in this important field.

胶原蛋白是我们体内含量最多的蛋白质。它也是最大的城市之一。这种大小对于将前胶原从合成点（细胞内质网内）运输到细胞外基质中的作用点的细胞机制来说是一个重大问题。控制这一途径第一步的机制通常产生的运输载体太小，无法容纳前胶原蛋白。几乎所有类型的细胞都会分泌一种或多种胶原蛋白，这一过程在细胞发育过程中提供细胞外基质是绝对必要的，细胞在细胞外基质中迁移到最终位置并分化。它对那些分化过程也是必不可少的。在生命的后期，胶原蛋白的持续分泌对组织的更新和再生至关重要。主要病理也与胶原分泌缺陷有关，包括纤维化。干细胞生物学领域越来越依赖于“干细胞生态位”的精确复制，其中分泌胶原的队列是一个主要组成部分。在许多情况下，工程细胞和组织的开发和利用将依赖于对这些过程的完全理解。因此，我们对这一过程所知之少也许令人惊讶。指导小蛋白分泌第一步的机制（为此获得了2013年诺贝尔奖）已经得到了很好的描述。这种机制产生的小运输囊泡（60-80纳米）不足以封装约300纳米大小的胶原蛋白分子。虽然在这一领域已经取得了一些进展，但它们要么与特定的胶原蛋白类型（如胶原VII）有关，要么就我们的理解而言尚不发达（如使用一种名为泛素的小蛋白质修饰核心机制）。我们的工作旨在结合尖端成像和蛋白质组学方法来精确定义这种机制，以及这些运输载体在前胶原蛋白输出点的性质。重要的是，我们将重点关注两种主要的前胶原同种型- I和II，以深入了解这些具有重要临床意义的形式的途径。我们已经产生了新的工具，以前所未有的空间和时间精度来检查这一过程。因此，我们期望在这一重要领域取得重大进展。

项目成果

ER-to-Golgi trafficking of procollagen in the absence of large carriers

在没有大载体的情况下，前胶原从内质网到高尔基体的运输

• DOI: 10.1101/339804
• 发表时间: 2018
• 作者: McCaughey J

P4HB recurrent missense mutation causing Cole-Carpenter syndrome

• DOI: 10.1136/jmedgenet-2017-104899
• 发表时间: 2018-03-01
• 期刊: JOURNAL OF MEDICAL GENETICS
• 影响因子: 4
• 作者: Balasubramanian, Meena;Padidela, Raja;Stephens, David J.
• 通讯作者: Stephens, David J.

The Golgi matrix protein giantin is required for normal cilia function in zebrafish.

斑马鱼中正常的纤毛功能需要Golgi基质蛋白巨蛋白。

• DOI: 10.1242/bio.025502
• 发表时间: 2017-08-15
• 期刊: Biology open
• 影响因子: 2.4
• 作者: Bergen DJM;Stevenson NL;Skinner REH;Stephens DJ;Hammond CL
• 通讯作者: Hammond CL

Giantin-knockout models reveal a feedback loop between Golgi function and glycosyltransferase expression.

• DOI: 10.1242/jcs.212308
• 发表时间: 2017-12-15
• 期刊: Journal of cell science
• 影响因子: 4
• 作者: Stevenson NL;Bergen DJM;Skinner REH;Kague E;Martin-Silverstone E;Robson Brown KA;Hammond CL;Stephens DJ
• 通讯作者: Stephens DJ

COPII-dependent ER export in animal cells: adaptation and control for diverse cargo.

• DOI: 10.1007/s00418-018-1689-2
• 发表时间: 2018-08
• 期刊: Histochemistry and cell biology
• 影响因子: 2.3
• 作者: McCaughey J;Stephens DJ
• 通讯作者: Stephens DJ