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Elucidating the molecular mechanism of Arp2/3-independent actin nucleation by WASP family proteins

阐明 WASP 家族蛋白独立于 Arp2/3 的肌动蛋白成核的分子机制

基本信息

批准号：
BB/N007581/1
负责人：
Kathryn Ayscough
金额：
$ 65.31万
依托单位：
University of Sheffield
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=BB%2FN007581%2F1
关键词：
Elucidating molecular mechanism Arp2 independent

项目摘要

Cells are the basic unit of life and all organisms are composed of one or more cells. Central to the functioning of many cells, including human cells, is the internal skeleton, or cytoskeleton. This cytoskeleton is required for cells to have certain shapes that are often a necessary part of their functioning. However, unlike our own body skeleton that is static, the cytoskeleton is able to remodel itself to change cell shape, or allow a cell to move. One of the most important proteins in the cytoskeleton is called actin. It is an amazing protein because it is almost the same now as hundreds of millions of years ago, long before humans, or even vertebrates existed. Staying so similar over time is called evolutionary conservation. Proteins that are very important to cell functioning are the most highly conserved. Actin is a protein that can join together with other actin proteins to form long lines or filaments. These filaments can be organised by other proteins to form large structures that are part of the cytoskeleton. We are interested in how actin is controlled in cells and in particular, we are trying to determine how the filaments can be started from single actin proteins. This is a process called nucleation. Our study will help us understand the mechanism of this nucleation in cells. We think that it is important because actin is known to be necessary for cell movement, and this behaviour often changes in cancer cells that have become metastatic. Actin is also involved when pathogens invade our cells. One of the proteins known to be important in helping actin form new filaments is called WASP, which becomes defective in an immune-deficiency disorder called Wiskott Aldrich Syndrome. Because actin is a highly conserved protein (85% identical between yeast and humans), we have undertaken many of our studies in yeast to gain insight into fundamental aspects of actin function. Manipulating mammalian systems is not always straightforward and some experiments can take months to perform. Yeast provides a more simple system to investigate, and we can study things within the context of the whole organism as well as analysing different components individually. This complementarity of approaches is important to gain a deep understanding of a process. We also aim to exploit finding by undertaking informed experiments on mammalian proteins and cells. There are many examples of studies in yeast that have shed new light on processes in more complex organisms.Until recently it was considered that WASP and other proteins like it, work to activate a group of proteins called the Arp2/3 nucleation complex or Arp2/3. We have shown that the yeast WASP, called Las17, is able to nucleate actin without Arp2/3 present. We were also able to show that this nucleation activity is important for the function of Las17 inside cells. Importantly, we have also shown that two related mammalian proteins can generate filaments in similar conditions, suggesting that the property is conserved. We now want to learn more about the mechanism of actin nucleation by these proteins as it may underpin a new understanding of nucleation at specific sites in cells. Overall, this project is highly relevant to our understanding of key cell processes of cell organization, membrane trafficking and motility. While focused on the yeast WASP homologue our preliminary data indicates that the major findings will be of wide significance for many proteins both of the WASP family and also other proteins such as those expressed on pathogens that also drive actin filament formation. Our approach is highly complementary to, but distinct from, those used in other labs. We have already generated many tools and reagents for this work which means that we can make rapid progress towards our goals, and the outputs have the potential to be published in top international journals thus enhancing UK competitiveness in science.

细胞是生命的基本单位，所有生物体都是由一个或多个细胞组成的。许多细胞（包括人类细胞）的核心功能是内部骨架或细胞骨架。这种细胞骨架是细胞具有一定形状所必需的，这通常是细胞功能的必要组成部分。然而，不像我们自己的身体骨架是静态的，细胞骨架能够自我改造，改变细胞形状，或允许细胞移动。细胞骨架中最重要的蛋白质之一叫做肌动蛋白。它是一种令人惊奇的蛋白质，因为它现在与数亿年前几乎相同，远在人类甚至脊椎动物存在之前。随着时间的推移保持如此相似被称为进化守恒。对细胞功能非常重要的蛋白质是最高度保守的。肌动蛋白是一种可以与其他肌动蛋白结合形成长线或长丝的蛋白质。这些细丝可以由其他蛋白质组织形成大的结构，是细胞骨架的一部分。我们对肌动蛋白在细胞中是如何被控制的很感兴趣，特别是，我们试图确定纤维是如何从单个肌动蛋白开始的。这个过程叫做成核。我们的研究将帮助我们了解这种细胞成核的机制。我们认为这很重要，因为肌动蛋白是细胞运动所必需的，而这种行为在癌细胞转移后经常发生变化。当病原体侵入我们的细胞时，肌动蛋白也参与其中。已知在帮助肌动蛋白形成新纤维方面起重要作用的一种蛋白质叫做WASP，它在一种叫做Wiskott Aldrich综合征的免疫缺陷疾病中会出现缺陷。由于肌动蛋白是一种高度保守的蛋白质（酵母和人类之间85%相同），我们在酵母中进行了许多研究，以深入了解肌动蛋白功能的基本方面。操纵哺乳动物的系统并不总是那么简单，有些实验可能需要几个月的时间来完成。酵母提供了一个更简单的研究系统，我们可以在整个有机体的背景下研究事物，也可以单独分析不同的成分。这种方法的互补性对于获得对流程的深入理解非常重要。我们还旨在通过对哺乳动物蛋白质和细胞进行知情实验来开发发现。有许多研究酵母的例子，为更复杂的生物体的过程提供了新的线索。直到最近，人们还认为WASP和其他类似的蛋白质能够激活一组叫做Arp2/3成核复合体（Arp2/3）的蛋白质。我们已经证明，称为Las17的酵母WASP能够在没有Arp2/3存在的情况下使肌动蛋白成核。我们还能够证明这种成核活性对Las17在细胞内的功能很重要。重要的是，我们还证明了两种相关的哺乳动物蛋白质可以在类似的条件下产生细丝，这表明这种特性是保守的。我们现在想要更多地了解肌动蛋白由这些蛋白质成核的机制，因为它可能支持对细胞特定部位成核的新理解。总的来说，这个项目与我们对细胞组织、膜运输和运动的关键细胞过程的理解高度相关。虽然主要集中在酵母WASP同源物上，但我们的初步数据表明，主要发现将对WASP家族的许多蛋白质和其他蛋白质（如病原体上表达的驱动肌动蛋白丝形成的蛋白质）具有广泛的意义。我们的方法与其他实验室使用的方法高度互补，但又截然不同。我们已经为这项工作生产了许多工具和试剂，这意味着我们可以朝着我们的目标取得快速进展，并且产出有可能在顶级国际期刊上发表，从而提高英国在科学方面的竞争力。

项目成果

期刊论文数量（5）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

WASP family proteins, more than Arp2/3 activators.

DOI：
10.1042/bst20160176
发表时间：
2016-10-15
期刊：
Biochemical Society transactions
影响因子：
3.9
作者：
Tyler JJ;Allwood EG;Ayscough KR
通讯作者：
Ayscough KR

Phosphorylation of the WH2 domain in yeast Las17/WASP regulates G-actin binding and protein function during endocytosis.

DOI：
10.1038/s41598-021-88826-z
发表时间：
2021-05-06
期刊：
Scientific reports
影响因子：
4.6
作者：
Tyler JJ;Smaczynska-de Rooij II;Abugharsa L;Palmer JS;Hancock LP;Allwood EG;Ayscough KR
通讯作者：
Ayscough KR

AP-2-Dependent Endocytic Recycling of the Chitin Synthase Chs3 Regulates Polarized Growth in Candida albicans