Circadian regulation of cell division in cyanobacteria: mechanism and function

许多生物体，从简单的细菌到复杂的哺乳动物，都有一个内部的“生物钟”，帮助调节24小时周期中的基本生命过程。生物钟是一个相互作用的基因和蛋白质的网络，这些基因和蛋白质反过来又与细胞中的其他过程相互作用，以打开或关闭基因。一个这样的过程是细胞分裂周期--亲本细胞复制其DNA并分裂成两个新细胞的过程。时钟控制着这个周期的速度和定时，时钟缺陷会导致严重的问题。在人类中，这些疾病包括癌症(不受调控的细胞周期是癌症进展的标志)和代谢综合征。我们的目标是了解时钟和细胞周期如何以及为什么一起工作。为了做到这一点，我们将研究已知的最简单的时钟，它是在一种细菌--光合作用的蓝藻细长聚球藻中发现的。通过研究这个简单的系统，我们可以建立时钟和细胞周期如何相互作用的原理，然后我们可以将其应用于更复杂的生物体。多年来，人们一直认为蓝藻的生物钟在一天中的特定时间阻止细胞分裂，而在其他时间没有影响。在之前的工作中，我们展示了时钟全天持续协调细胞分裂。然而，分子的身份和它们如何协同工作(即这种控制的机制)仍然是未知的。我们假设，生物钟通过调节一种名为FtsZ的蛋白质的水平来控制细胞何时分裂，这种蛋白质在蓝藻细胞中积累在细胞分裂的地方。为了验证我们的假设，我们将结合数学建模--以定量验证我们的理解--与尖端的实验技术相结合。我们的方法将提供关于单个细胞(而不是整个群体平均在一起)行为的高分辨率数据。单个细胞之间的差异通常是区分假说的关键。同样重要的是，这些数据是动态的，否则无法确定因果关系。我们将在显微镜下跟踪数千个细胞，持续测量每个细胞中FtsZ产生的位置和速度。我们将使细胞中产生的FtsZ的数量与24小时周期内细胞分裂的模式相匹配。结合这些观察结果和我们的数学模型，我们将确定时钟FtsZ和细胞周期之间的精确关系，并确定是否有其他相关分子。至于为什么时钟调节细胞周期的问题，我们需要探索时钟如何与环境变化接合并做出反应。为了做到这一点，我们将调节环境光线水平，并使用微流控技术，在这种环境中，单个细胞的局部环境可以被精确控制和快速修改。刺激细胞产生FtsZ的模式与时钟和环境昼夜周期都不同步，我们将观察细胞是如何受到影响的，并由此推断时钟控制细胞周期的目的。通过全面、定量地了解蓝藻生物钟调节细胞周期的机制和结果，本项目将为进一步研究这两个关键细胞系统之间的相互作用奠定坚实的基础。我们的概念框架将适用于更复杂的生物，如人类，在那里，时钟和细胞周期也密切相互作用，以维持健康，机制的理解仍然极其有限。彻底了解细胞分裂控制是发展控制细胞分裂能力的第一步，这可能与未来涉及细胞分裂的不同领域的研究以及在生物医学和合成生物学中的应用有关。

Many organisms, from simple bacteria to complex mammals, have an internal 'circadian' clock that helps regulate essential life processes on a 24-hour cycle. The circadian clock is a network of interacting genes and proteins, which in turn interact with other processes in cells to switch genes on or off. One such process is the cell division cycle - the process by which a parent cell replicates its DNA and divides into two new cells. The clock controls the speed and timing of this cycle and clock defects cause serious problems. In humans, these include cancer (an unregulated cell cycle is a hallmark of cancer progression) and metabolic syndrome.Our goal is to understand both how and why the clock and the cell cycle work together. To do so, we will study the simplest known clock, which is found in a type of bacteria, the photosynthetic cyanobacterium Synechococcus elongatus. By studying this simple system, we can establish the principles of how clocks and the cell cycle interact, which we can then apply to more complicated organisms.For many years, it was believed that the circadian clock of cyanobacteria 'blocks' cell division at certain times of day, and has no influence at other times. In previous work, we showed that the clock continuously orchestrates cell division throughout the day. The identities of the molecules and how they work together (i.e. the mechanism of this control) remain, however, unknown.We hypothesise that the circadian clock controls when cells divide by regulating the levels of a protein called FtsZ, which accumulates in cyanobacterial cells at the place where the cell will divide. To test our hypothesis, we will combine mathematical modelling - to quantitatively validate our understanding - with cutting-edge experimental techniques. Our approach will provide high-resolution data on the behaviour of individual cells (rather than whole populations averaged together). Differences between individual cells are often crucial to distinguish between hypotheses. It is also important that this data is dynamic, as otherwise causality cannot be determined.We will track thousands of cells over time under a microscope, continuously measuring the location and rate of FtsZ production in each cell. We will match the amount of FtsZ produced in cells to the patterns of cell division over 24-hour cycles. Combining these observations with our mathematical models, we will determine the precise relation between the clock, FtsZ and the cell cycle, and identify any other molecules involved.Turning to the question of why the clock regulates the cell cycle, we need to explore how the clock engages with and responds to environmental changes. To do this, we will regulate the ambient light levels and use microfluidics, where the local environment of individual cells can be precisely controlled and rapidly modified. Stimulating cells to produce FtsZ in a pattern that is 'out of sync' with both the clock and the environmental day/night cycle, we will observe how the cells are affected and, from this, infer the purpose of the clock's control of the cell cycle. By providing a comprehensive, quantitative understanding of the mechanism and consequences of the regulation of the cell cycle by the circadian clock in cyanobacteria, this project will establish a firm foundation for further studies of the interplay between these two critical cell systems. Our conceptual framework will be applicable to more complex organisms, such as humans, where the clock and the cell cycle also interact tightly to maintain health and mechanistic understanding remains extremely limited. Having a thorough understanding of cell division control is the first step towards developing the capabilities to control it, which could be relevant for future research in the diverse areas involving cell division as well as for applications in biomedicine and synthetic biology.