Size Matters: A systems approach to understanding cell size control in a developing multicellular tissue

细胞是组织的基石，但如何从细胞中构建三维结构是生物学中一个尚未解决的基本问题。当我们考虑一个正在生长的组织时，情况就更加复杂了，在这个组织中，组成部分（细胞）的物理性质随着它们的生长和分裂而不断变化。研究了模式植物拟南芥生长的茎尖或茎分生组织。这种结构是植物生长过程中产生新叶和新花所必需的。器官发育的早期阶段需要有效的组织生长，此时通常观察到细胞大小的增加。几年来，我们已经开发了技术，使我们能够在共聚焦显微镜下长时间成像分生组织。我们可以随时间跟踪单个细胞并确定它们的生长和分裂。为了分裂，细胞要经历一系列被称为细胞周期的过程。利用我们开发的荧光报告器，我们首次可以在植物组织生长的成像时间过程中同时确定所有细胞在细胞周期中的位置。我们最近发表的研究表明，对于给定的组织和环境条件，细胞分裂时的大小平均是一致的，但是当这些变化时，细胞的可塑性很强。例如，我们发现，当植物生长在限制光合作用的环境条件下时，细胞更小，组织生长更慢。细胞大小的变化是由不均匀分裂引起的，必须加以消除。这是通过在每个细胞的基础上建立生长和分裂之间的平衡来实现的。系统的可塑性使我们认为，细胞分裂时的大小不是由直接的“细胞标尺”决定的，而是由生长和分裂过程的结果（或“涌现特性”）决定的。这种机制似乎在单细胞生物中是保守的，在充足的条件下，单细胞生物也可以实现更大的细胞大小和更高的绝对生长速率。在这些简单细胞中，细胞大小的调节是通过细胞的蛋白质合成能力来平衡细胞生长和分裂来实现的。我们开发了一种模型，可以根据细胞生长过程中细胞分裂所需的两种调节蛋白（CDKs）的生长速度和活性积累，准确预测植物细胞的大小。我们使用不同的突变体和生长条件进行了广泛的测试，并确定了导致细胞大小控制的关键过程。这些过程似乎是CDK活性的产生和阈值。在本提案中，我们将确定参与这些过程的“大小”分子，这些分子建立了细胞生长和分裂之间的联系，并使用我们最先进的成像和分析技术分析它们的功能。我们有许多候选尺寸，在CDK调节中具有已知的作用，但我们也将开展实验，利用基于确定蛋白质在不同条件下合成速率的全基因组方法，以公正的方式确定新的候选尺寸。我们将使用实验和数学相结合来开发一个模型，使我们能够了解这些大小分子是如何被调节的，以及这对细胞大小控制的影响。

Cells are the building blocks of tissues, but how 3D structures are built from cells is a fundamental unsolved problem in biology. It is even more complex when we consider a growing tissue, in which the physical properties of the building blocks (cells) change constantly as they grow and divide. We study the growing shoot tip or shoot meristem of the model plant Arabidopsis. This structure is required to produce new leaves and flowers as the plant grows. The early stages of organ development require efficient tissue growth and an increase in cell size is normally observed at this time. Over several years, we have developed techniques allowing us to image the meristem over extended periods in the confocal microscope. We can follow individual cells over time and determine their growth and division. In order to divide, a cell goes through a defined series of processes known as the cell cycle. Using fluorescent reporters we have developed, we can for the first time simultaneously determine the position of all cells in the cell cycle during imaging time courses of growing plant tissues. Our recently published study showed that the size cells reach when they divide is on average consistent for a given tissue and set of environmental conditions, but is highly plastic when these change. For example we found that cells were smaller, and tissue growth slower, when plants were grown under environmental conditions that restrict photosynthesis. Variation in cell size arises through unequal division and has to be removed. This is done by establishing a balance between growth and division on a cell by cell basis. The plasticity of the system leads us to consider that cell size at division is not determined by a direct "cellular ruler" but is instead determined as a consequence (or "emergent property") of the contributing processes of growth and division. This mechanism appears to be conserved from unicellular organisms which can also achieve a larger cell size and higher absolute growth rate under plentiful conditions. Regulation of cell size in these simple cells is achieved by balancing cell growth and division via the protein synthetic capacity of the cell. We have developed a model that can predict accurately the size of plant cells based on the rate of growth and the accumulation of activity of two regulatory proteins required for cell division (called CDKs) as the cell grows. We tested this extensively using different mutants and growth conditions and identified the key processes that lead to cell size control. These processes appear to be the production and thresholding of CDK activity. In this proposal we will identify the "sizer" molecules involved in these processes that establish the link between growth and division of cells and analyse their function using our state-of-the-art imaging and analysis techniques. We have a number of candidate sizers, with known roles in CDK regulation, but we will also carry out experiments to identify new candidates in an unbiased manner using a genome-wide approach based on identifying the rate at which proteins are being synthesized under different conditions. We will use a combination of experiments and mathematics to develop a model that will allow us to understand how these sizer molecules are regulated and what effect this has on cell size control.