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

基因相同的个体，从细菌种群到人类双胞胎，即使他们有相同的基因，也往往表现不同。我们很想知道这些差异是如何产生的，以及这些差异可能产生什么影响。我们选择的模型系统是一种小型的模式植物，拟南芥，它每一代产生1000个遗传相同的种子。植物生长在一个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，即使在基因相同并生长在相同环境中的植物中，大约9%的基因在不同植物之间的激活水平也会有很大差异。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植物之间的表达水平特别可能是可变的。这种基因激活的变化在自然界中可能对遗传相似的植物种群有用，以对冲不可预测的环境压力。这是因为可变的基因表达意味着种群中总是有少数植物由于其可变的基因激活而准备好在不同的胁迫下生存。胁迫应答基因的高表达可能是有代价的，这可以使可变基因表达比所有植物都具有高表达水平的基因更有利。除了在野外具有潜在的益处之外，植物之间的变异性也可能是一个问题，例如在环境受到更多控制的农业中，农民需要同时发芽和开花的统一作物，并且对肥料和水的应用做出同样的反应。我们试图理解在相同条件下生长的遗传上相同的植物之间的基因活性差异是如何通过细胞水平的过程产生的。为此，我们将使用两种不同的实时成像技术来监测候选“噪声”途径随时间的基因活性。我们将使用发光报告基因（使用萤火虫的荧光素酶基因），在整个幼苗的水平上为我们读出我们感兴趣的基因的表达。这项技术将使我们能够监测特定基因的表达如何随着时间的推移以及在叶，茎和根等组织中变化。我们还将使用荧光报告分子（使用来自水母的绿色荧光蛋白），以允许对感兴趣的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和设计新的实验。我们将集中在一个途径控制的压力反应激素，阿坝，因为我们已经发现，许多基因调控它似乎是高度可变的从植物到plant.Finally，我们将确定的影响植物生长和生存的嘈杂的基因激活，我们的特点。我们将跟踪基因活性之前和之后的压力，并检查是否激活的途径在压力前的水平与压力后植物的生存。这可能是因为在胁迫之前随机激活该途径，使得具有较高表达的植物能够在条件的突然变化中存活下来（一种赌注对冲策略）。在未来，了解植物如何产生和调节基因活动中的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气候变化导致的更频繁的极端天气中生存将是非常重要的。

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.