Mechanisms to generate inter-individual variability from single-cell heterogeneity

基因相同的个体，从细菌群体到人类双胞胎，往往表现不同，即使他们拥有相同的基因。我们有兴趣知道这些差异是如何产生的，以及这些差异可能会产生什么影响。我们选择的模式系统是一种小型模式植物拟南芥，它每一代产生数千个基因相同的种子。植物生长在多变和嘈杂的环境中，包括波动的光、水和温度。植物也有一个嘈杂的内部环境；我们实验室最近的工作表明，即使在基因相同且生长在相同环境中的植物中，大约9%的基因的激活水平在不同植物之间也会高度不同。我们发现，帮助植物在干旱、高盐度或极端温度等环境条件下生存的应激反应基因在不同植物之间的表达水平特别可能不同。基因激活的这种变异在自然界中可能对遗传相似的植物种群有用，以对冲对不可预测的环境压力的押注。这是因为可变的基因表达意味着种群中总是有一些植物由于其可变的基因激活而准备好承受不同的压力。胁迫反应基因的高表达可能是有代价的，这可能使可变基因表达比所有始终高表达同一基因的植物更有利。除了在野外有潜在的好处外，植物之间的差异也可能是一个问题，例如在农业中，环境受到更多的控制，农民要求统一的作物同时发芽和开花，并对化肥和水的应用做出同样的反应。在这项建议中，我们试图了解在相同条件下生长的基因相同的植物之间的基因活性差异是如何由细胞水平的过程产生的。为了做到这一点，我们将使用两种不同的实时成像技术来监控候选“嘈杂”途径随时间的基因活动。我们将使用发光记者(使用萤火虫的荧光素酶基因)来读出我们感兴趣的基因在整个幼苗水平上的表达。这项技术将使我们能够监测特定基因的表达如何随时间以及叶、茎和根等组织的变化而变化。我们还将使用荧光记者(使用来自水母的绿色荧光蛋白)来对感兴趣组织中的单个活细胞进行更高分辨率的成像。我们将测试组织和单个细胞中基因活性水平的变异性如何导致整个植物水平的差异。为了理解我们的实验结果，我们将建立基因激活动力学的数学模型，这将有助于我们测试我们的理解并设计新的实验。我们将重点关注一种由胁迫反应激素ABA控制的途径，因为我们已经发现，受ABA调控的许多基因在不同植物之间似乎是高度可变的。最后，我们将确定我们所描述的噪声基因激活对植物生长和生存的影响。我们将跟踪添加胁迫前后的基因活性，并检查胁迫前途径的激活水平是否与胁迫后植物的生存相关。这可能是在胁迫前随机激活的途径允许高表达的植物在条件的突然变化中存活下来(一种更好的对冲策略)。在未来，了解植物如何在基因活动中产生和调节噪音，对于开发更均匀的作物，以及了解野生植物种群如何在由于气候变化而导致的更频繁的极端天气中生存下来，将是重要的。

Genetically identical individuals, from populations of bacteria to human twins, often behave differently, even though they have identical genes. We are interested to know how these differences are generated, and what effects these differences might have. Our model system of choice is a small model plant, Arabidopsis, which produces 1000s of genetically identical seeds each generation. Plants grow in a variable and noisy environment, including fluctuating light, water and temperature. Plants also have a noisy internal environment; recent work from our laboratory has revealed that even in plants that are genetically identical and grown in the same environment, around 9% of genes will be highly variable in their level of activation from plant to plant. We found that stress response genes that help plants survive environmental conditions such as drought, high salinity or extreme temperatures were particularly likely to be variable in their expression levels between plants. This variation in gene activation could be useful in nature for populations of genetically similar plants to hedge bets against unpredictable environmental stresses. This is because variable gene expression would mean that there are always a few plants in the population that are prepared to survive different stresses due to their variable gene activation. There might be a cost to having high expression of stress response genes, which could make variable gene expression more advantageous than all plants having high expression levels of a gene all of the time. As well as having potential benefits in the wild, variability between plants can also be a problem, such as in agriculture where environments are more controlled and farmers require uniform crops that germinate and flower at the same time and respond equally to applications of fertilisers and water.In this proposal, we seek to understand how the differences in gene activity between genetically identical plants grown in the same conditions are generated by cellular-level processes. To do this, we will use two different live imaging techniques to monitor gene activity of candidate 'noisy' pathways over time. We will use luminescent reporters (using a luciferase gene from fireflies) to give us a read out of the expression of our genes of interest at the level of whole seedlings. This technique will allow us to monitor how expression of a particular gene varies over time and across tissues such as leaves, stems and roots. We will also use fluorescent reporters (using green fluorescent protein from jelly fish) to allow higher resolution imaging of individual living cells in the tissues of interest. We will test how the variability in the levels of activity of genes in tissues and individual cells leads to the differences at the level of whole plants. To understand the results of our experiments, we will build mathematical models of the gene activation dynamics that will help us test our understanding and design new experiments. We will focus on a pathway controlled by a stress response hormone, ABA, as we have found that many genes regulated by it appear to be highly variable from plant to plant.Finally, we will determine the effects on plant growth and survival of the noisy gene activation we have characterised. We will track gene activity before and after the addition of stress, and examine whether the level of activation of a pathway before the stress correlates with the survival of a plant after stress. It could be that the random activation of the pathway before stress allows the plants that have higher expression to survive the sudden change in conditions (a bet-hedging strategy). In the future, understanding how plants produce and regulate noise in gene activity will be important for the development of more uniform crops and to understand how populations of wild plants can survive more frequent weather extremes due to climate change.