Integration of cell division and plant development: control of leaf growth and cell number by AINTEGUMENTA

植物的叶子和花（“器官”）在芽的尖端形成。在茎的顶端有一个微观结构，像一个小的浅圆顶状结构，被称为茎尖分生组织。在这个圆顶的两侧，小突起或“原基”形式，将生长成一片新叶。原基中的细胞在一段时间内迅速分裂，直到叶子长到几毫米长，有足够的细胞形成最后的叶子，然后它们停止分裂并扩张。因此，叶子从几毫米的大小生长到完全成熟的大小是由现有细胞的膨胀引起的，而不是由更多的细胞形成引起的。是什么控制着叶子中细胞的数量和最终的大小？我们的目标是在拟议的项目中找到这个问题的答案。我们所知道的是，有一些突变会影响器官的大小和器官中细胞的数量。其中一种鉴定了一种被称为ANT的基因。ANT的突变会减少器官中的细胞数量，并对花朵产生其他影响。增加ANT基因的表达量会增加叶片的大小和叶片中细胞的数量。像ANT这样的基因被称为“发育调节剂”，因为它们控制着植物的生长和发育。特别是，蚂蚁控制叶子中细胞的数量和叶子的总体大小。我们还知道，细胞分裂的决定取决于被称为d型细胞周期蛋白（CYCDs）的蛋白质水平。然而，cycd和发育器官大小调节因子（如ANT）之间没有已知的联系。因此，我们不知道细胞分裂在发育过程中是如何被控制的。更一般地说，我们不了解细胞——器官和生物体的基本组成部分——是如何被控制以产生更大的结构的。我们认为ANT调节了一种叫做CYCD3的CYCD基因；并有证据支持这一点。ANT水平升高导致CYCD3表达增加；1、改变CYCD3水平；1也会影响叶片中细胞的数量，与改变ANT的效果类似。这两个观察结果表明，ANT可以通过CYCD3起作用；由ANT基因编码的蛋白质是一种“转录因子”，可以结合DNA并控制其他基因，因此ANT蛋白可以直接打开CYCD3；1基因。已知蚂蚁蛋白结合位点的序列，在CYCD3的启动子（控制区）中发现；而不是在其他CYCD基因的启动子中。因此，情况很有可能就是这样。在这个项目中，我们将证明ANT蛋白调控CYCD3；1通过结合其启动子，并鉴定其他受ANT调控的基因。这将是植物中细胞分裂和发育调节因子之间的第一个联系，并将帮助我们理解细胞的行为在发育过程中是如何协调的。首先，我们要证明ANT的作用取决于CYCD3的存在；我们可以这样做是因为我们在CYCD3；1中有一个突变体。我们预测当CYCD3；1缺失，证明CYCD3；1受ANT调控。然而，尚不能证明这是否是ANT与CYCD3结合的直接影响；1基因。因此，我们将证明ANT与CYCD3结合；1启动子。最后，我们希望进一步了解蚂蚁控制的其他基因在叶片发育中的作用。

Plant leaves and flowers ('organs') form at the tips of shoots. At the very tip of the shoot is a microscopic structure like a small shallow dome-like structure, known as the shoot apical meristem. On the sides of this dome, small protrusions or 'primordia' form, that will grow into a new leaf. Cells in primordia divide rapidly for a time until the leaf is a couple of millimetres long and there is enough cells to make the final leaf, and then they stop dividing and expand. The growth of the leaf from the size of a few millimetres up to its fully mature size is therefore caused by expansion of existing cells, and not by more cells being formed. What controls the number of cells in the leaf and its eventual size? We aim to find answers to this question in the proposed project. What we know is that there are mutants that affect organ size and the number of cells in an organ. One identifies a gene known as AINTEGUMENTA (ANT). Mutations in ANT reduce the number of cells in organs and also have other effects in the flowers. Increasing the amount of expression of the ANT gene increases leaf size and the number of cells in the leaf. Genes like ANT are known as 'developmental regulators' because they control the growth and development of the plant. In particular, ANT acts to control the number of cells in the leaf and the its overall size. We also know that cell makes a decision to divide depending on the level of proteins known as D-type cyclins (CYCDs). However, there is no known link between CYCDs and developmental organ size regulators such as ANT. We therefore do not know how cell division is controlled in development. More generally, we do not understand how cells -the fundamental building blocks of organs and organisms- are controlled to produce larger structures. We believe that ANT regulates a particular CYCD gene called CYCD3;1 and have evidence to support this. Increasing ANT levels leads to increased expression of CYCD3;1, and altering levels of CYCD3;1 also affects the number of cells in leaves, in a similar way to the effects of altering ANT. These two observations suggests that ANT could work through CYCD3;1. The protein encoded by the ANT gene is a 'transcription factor' that binds DNA and controls other genes, so ANT protein could directly switch on the CYCD3;1 gene. The sequence of the binding site of the ANT protein is known and it is found in the promoter (controlling region) of the CYCD3;1 gene and not in the promoters of other CYCD genes. There is therefore a good likelihood that this may be the case. In this project, we are going to prove that the ANT protein regulates CYCD3;1 by binding its promoter, and also identify other genes regulated by ANT. This will be the first link between cell division and a developmental regulator in a plant, and will help us understand how the behaviour of cells is co-ordinated during development. First we are going to prove that the action of ANT depends on the presence of CYCD3;1. We can do this because we have a mutant in the CYCD3;1. We predict that ANT will have no effect when CYCD3;1 is missing, proving that CYCD3;1 is regulated by ANT. However it will not prove whether this a direct effect of ANT binding to the CYCD3;1 gene. Thus we will then prove that ANT binds to the CYCD3;1 promoter. Finally we wish to understand more about the other genes ANT controls in leaf development.