Dissecting the regulation and function of actin flows during cell motility

剖析细胞运动过程中肌动蛋白流的调节和功能

• 批准号: BB/V006169/1
• 负责人: Brian Stramer
• 金额: $ 84.17万
• 依托单位: King's College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FV006169%2F1
• 关键词: Dissecting; regulation; function; actin; flows

Cell migration is a fundamental cellular behaviour that occurs during normal physiology, such as embryogenesis, as well as pathologies, such as cancer metastasis. Understanding how cells control their movement is therefore important for numerous fundamental biological processes and is of translational relevance. For the past 50 years our view of cell migration has involved a step-wise cycle of subcellular behaviours beginning with extension of the membrane at the front of the cell. However, recent evidence in a number of cell types has revealed that the leading edge is instantaneously coordinated with other 'steps' of the migratory cycle: cell motility is therefore not a step-wise process. Our goal now is to understand how these migratory behaviours are coordinated in space and time to control cell motion. Our laboratory has pioneered the use of embryonic Drosophila macrophages (hemocytes) as an in vivo migration model to address fundamental processes controlling cell motility. These cells can be live imaged at very high resolution within the animal, and coupled with the genetic tractability of the fruitfly, this system is therefore a powerful model to understand the control of cell motility during normal physiology. Using this model we have recently revealed that the actin cytoskeletal network, which is known to be the driver of cell motility, undergoes constant flow within hemocytes. Through the development of computational tools we have revealed that this actin flow is highly coordinated across the entire cell resulting in stable sinks within the flowfield that appear to control actin flows across the cell and direct cell motion. We hypothesise that this orchestrated actin flow is a critical factor that controls the subcellular behaviours involved in cell migration, which will be dissected in further detail in this proposal. As actin flows are a central feature of all migrating cells, the results of this work will be relevant to many biological processes. In the first Objective we will further develop our computational tools to analyse actin flows and subsequently use these novel approaches to determine the timecourse of actin flow changes during normal hemocyte behaviours, such as responses to wound signals. We will also investigate how actin motion is correlated with the movement of myosin, a critical motor responsible for contraction and subsequent flow of the actin network. The results of this objective will form a baseline understanding that will allow us to precisely determine the function of regulators in subsequent objectives. In the next Objective we will examine how actin flows are regulated by testing flies containing mutations in genes hypothesized to control the actin network. Additionally, we will examine how the motion of the actin network is controlling and/or coordinated by the microtubule network, a second cytoskeletal network that is important in cell migration. Again, fly lines containing mutations in known regulators of the microtubule network will be tested to determine how this network is regulating actin motion. In the final Objective we will examine how actin flow may be directly controlling the activity of a regulator of cell migration responsible for defining cell polarity. Moesin is an actin regulator known to localize to the rear of many migratory cell types where it controls polarised motility. Preliminary data reveals that moesin localises to the rear of migrating hemocytes and that moesin mutants have defects in cell movement and actin flow. Previous mathematical modelling has suggested that any protein that binds to the actin network with sufficient strength will be carried rearward by the actin flow resulting in a gradient that may subsequently control cellular behaviour. We hypothesise that moesin is an ideal candidate for such direct regulation by actin flow and we will examine how moesin is controlled by the flow and subsequently feeds back to control the actin network.

细胞迁移是一种基本的细胞行为，发生在正常生理过程中，如胚胎发生，以及病理过程中，如癌症转移。因此，了解细胞如何控制它们的运动对于许多基本的生物学过程是重要的，并且具有翻译相关性。在过去的50年里，我们对细胞迁移的看法涉及到亚细胞行为的逐步循环，从细胞前部膜的延伸开始。然而，最近在许多细胞类型中的证据表明，前沿与迁移周期的其他“步骤”是即时协调的：因此，细胞运动不是一个循序渐进的过程。我们现在的目标是了解这些迁移行为是如何在空间和时间上协调以控制细胞运动的。我们的实验室率先使用胚胎果蝇巨噬细胞（血细胞）作为体内迁移模型来解决控制细胞运动的基本过程。这些细胞可以在动物体内以非常高的分辨率进行实时成像，再加上果蝇的遗传易感性，因此该系统是一个强大的模型，可以理解正常生理过程中细胞运动的控制。利用这个模型，我们最近揭示了肌动蛋白细胞骨架网络，它被认为是细胞运动的驱动因素，在血细胞内经历恒定的流动。通过计算工具的发展，我们发现这种肌动蛋白流动在整个细胞中是高度协调的，导致流场内稳定的汇，这似乎控制了肌动蛋白在细胞中的流动并指导细胞运动。我们假设这种精心安排的肌动蛋白流动是控制参与细胞迁移的亚细胞行为的关键因素，这将在本提案中进一步详细剖析。由于肌动蛋白流动是所有迁移细胞的核心特征，因此这项工作的结果将与许多生物过程相关。在第一个目标中，我们将进一步开发我们的计算工具来分析肌动蛋白流动，并随后使用这些新方法来确定正常血细胞行为（如对伤口信号的反应）期间肌动蛋白流动变化的时间进程。我们还将研究肌动蛋白运动如何与肌凝蛋白运动相关，肌凝蛋白是负责肌动蛋白网络收缩和随后流动的关键运动。这一目标的结果将形成一个基线理解，使我们能够精确地确定监管机构在后续目标中的功能。在下一个目标中，我们将通过测试含有控制肌动蛋白网络的基因突变的果蝇来研究肌动蛋白流动是如何调节的。此外，我们将研究肌动蛋白网络的运动是如何由微管网络控制和/或协调的，微管网络是细胞迁移中重要的第二个细胞骨架网络。同样，将测试含有已知微管网络调节因子突变的果蝇系，以确定该网络是如何调节肌动蛋白运动的。在最后的目标中，我们将研究肌动蛋白流动如何直接控制负责定义细胞极性的细胞迁移调节剂的活性。Moesin是一种肌动蛋白调节剂，已知位于许多迁移细胞类型的后部，在那里它控制极化运动。初步数据显示，moesin定位于迁移血细胞的后方，并且moesin突变体在细胞运动和肌动蛋白流动方面存在缺陷。先前的数学模型表明，任何与肌动蛋白网络结合的蛋白质，只要强度足够，就会被肌动蛋白流带往后面，从而产生一个梯度，从而可能控制细胞行为。我们假设moesin是由肌动蛋白流动直接调节的理想候选者，我们将研究moesin是如何被肌动蛋白流动控制的，并随后反馈控制肌动蛋白网络。

项目成果

Imaging actin organisation and dynamics in 3D

• DOI: 10.1242/jcs.261389
• 发表时间: 2024-01-01
• 期刊: JOURNAL OF CELL SCIENCE
• 影响因子: 4
• 作者: Phillips，Thomas A.;Marcotti，Stefania;Parsons，Maddy
• 通讯作者: Parsons，Maddy

Bridging Imaging Users to Imaging Analysis - A community survey.

将影像用户与影像分析联系起来 - 一项社区调查。

• DOI: 10.1101/2023.06.05.543701
• 发表时间: 2023
• 期刊: bioRxiv : the preprint server for biology
• 作者: Sivagurunathan,Suganya;Marcotti,Stefania;Nelson,CarlJ;Jones,MartinL;Barry,DavidJ;Slater,ThomasJA;Eliceiri,KevinW;Cimini,BethA
• 通讯作者: Cimini,BethA