The roles of the Polo and MAP kinase signalling in driving polarised growth in fission yeast

在生物体内特化细胞类型的发育过程中，多样性部分是由极化生长和细胞内不对称产生的。通过细胞骨架元素之间的相互作用，这依赖于细胞命运决定因素的极化分布。细胞骨架是一种“内部支架”，对控制细胞功能的许多方面都很重要，包括分裂细胞以继承不同成分的能力。这种“不对称”分裂对于建立体内不同细胞的多样性非常重要。在这个项目中研究的两个细胞骨架元素，肌动蛋白和微管，是高度动态的聚合物，它们快速生长和收缩以响应外部和内部的信号，以建立和维持极化的细胞骨架。裂变酵母是研究极化生长的理想模型系统，因为它是一种简单的细胞类型。它的微管和肌动蛋白相互作用的方式和在更复杂的系统中一样。肌动蛋白和微管细胞骨架的联合作用严格限制了这种圆柱形细胞的细胞末端的生长。此外，负责细胞骨架极化变化的潜在控制与更高系统的控制相同或高度相似。Polo激酶是一种调节分子，以前被认为与细胞分裂的控制有关，但在细胞周期的非分裂阶段（称为间期）中没有控制。因此，我们最近发现polo激酶通过调节细胞骨架来控制间期细胞生长，这已经确定了一种对细胞骨架极化很重要的新的信号通路。我们的目标是进一步表征分裂酵母polo激酶Plo1的间期作用。我们知道Plo1控制细胞骨架的能力是由应激反应途径（SRP）控制的。SRPs使细胞能够快速响应环境中的各种变化，包括过热和饥饿。像人类这样的生物体有几个srp，每一个都对不同的压力做出反应。相反，在裂变酵母中，单个SRP响应各种外部刺激，使这种酵母成为一个特别有吸引力的模型，为更复杂的人类SRP的研究提供指导。本项目旨在研究细胞如何从应激诱导的生长扰动中恢复。在此过程中，它将扩展我们对肌动蛋白和微管细胞骨架如何相互交谈以启动新生长的理解。我们将利用特定突变可以用来研究酵母中蛋白质的功能的便利。我们将首先询问polo激酶物理修饰目标分子的能力是否对我们所看到的效果很重要，或者它是否只是必须存在（即具有纯粹的结构作用）。第二个目标是编目Plo1/SRP定位在细胞内随细胞周期和应激诱导的细胞骨架变化的变化。通过产生SRP成分与Plo1和荧光蛋白之间的融合，我们已经能够看到这些分子在活细胞中的去向。这使我们能够观察到细胞暴露于不同外部条件时细胞内蛋白质分布的变化。第三个目标是通过鉴定和表征SRP和Polo信号通路的靶标/相互作用物，确定和表征Polo和SRP信号通路协调调节细胞骨架极化变化的机制。总之，这个项目将增加我们对细胞如何响应来自环境的信号来调节和维持细胞生长的理解。

During the development of specialized cell types within an organism, diversity is generated in part by polarised growth and intracellular asymmetry. This relies on the polarised distribution of cell-fate determinants, through interplay between elements of the cytoskeleton. The cytoskeleton is an 'internal scaffold' that is important for controlling many aspects of cell function, including the ability of dividing cells to inherit different components. Such 'asymmetric' divisions are important in setting up the diversity of different cells within the body. The two cytoskeletal elements studied in this project, actin and microtubules, are highly dynamic polymers that grow and shrink rapidly to respond to external and internal cues, in order to establish and maintain a polarised cytoskeleton. The fission yeast is an ideal model system for studying polarised growth because it is a simple cell type. Its microtubules and actin interact in the same way as they do in more complex systems. The combined effect of the actin and microtubule cytoskeletons strictly limits growth to the cell ends of this cylindrical shaped cell. Furthermore the underlying controls responsible for changes in cytoskeletal polarisation are identical or highly reminiscent of those of higher systems. Polo kinases are regulatory molecules that have previously been linked to the control of cell division but not to control during the non-dividing phases of the cell cycle known as interphase. Therefore, our recent finding that polo kinases controls interphase cell growth by regulating the cytoskeleton has identified a novel signaling pathway important for cytoskeletal polarisation. We aim to further characterize this interphase role of the fission yeast polo kinase Plo1. We know that the ability of Plo1 to control the cytoskeleton is controlled by the stress response pathway (SRP). SRPs enable cells to rapidly respond to a variety of changes in their environment including excessive heat and starvation. Organisms such as humans have several SRPs, each of which responds to different stresses. In contrast in fission yeast a single SRP responds to a variety of external stimuli, making this yeast a particularly attractive model to provide guidance for the study of the more complex human SRPs. This project aim to study how cells recover from stress induced perturbations of growth. In doing this it will extend our understanding of how actin and microtubule cytoskeletons talk to each other to initiate new growth. We will take advantage of the ease with which specific mutations can be used to study the function of a protein in yeast. We will first ask whether the ability of the polo kinase to physically modify target molecules is important for the effects we see or whether it simply has to be present (i.e. has a purely structural role). The second aim is to catalogue the changes in Plo1/SRP localization within the cells that accompany cell cycle and stress induced changes in the cytoskeleton. By generating fusions between both SRP components and Plo1 and fluorescent proteins we have been able to see where in living cells the molecules go. This allows us to observe the changes in the distribution of the proteins within the cells as the cells are exposed to different external conditions. The third aim is to identify and characterize the mechanism by which Polo and SRP signaling are coordinated to regulate changes in cytoskeletal polarisation through the identification and characterization of targets/interactors of the SRP and Polo signaling pathways. In summary this project will increase our understanding of how cells respond to signals from the environment to regulate and maintain cell growth.