Modified function of a stem cell regulator in monocots and dicots

单子叶植物和双子叶植物干细胞调节剂的功能修饰

• 批准号: BB/X000559/1
• 负责人: Peter Etchells
• 金额: $ 72.78万
• 依托单位: Durham University
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FX000559%2F1
• 关键词: Modified; function; stem; cell; regulator

The majority of our food crops come from flowering plants, which are referred to as angiosperms. Angiosperms are further divided into the dicots, which include many woody species, and the monocots, which include important cereal crops such as wheat and barley. One of the major differences between the monocot and dicot clades is the organisation of the stem. In dicots, the stem is characterised by a group of undifferentiated cells that are referred to as the cambium, which is responsible for most radial growth. The cambium is present in a ring around the stem and cells that divide within the cambium ultimately form the woody plant tissue at the centre of the stem (xylem), or a second transport tissue, the phloem, towards the outer part of the stem. Cell division in the cambium is controlled by a signalling mechanism referred to as TDIF-PXY. TDIF is a small ligand protein, and PXY a receptor protein that is localised to the plasma membrane of cambium cells. When TDIF binds to PXY, a signal is passed to the nucleus which promotes cell division. Monocots do not have a cambium. However, TDIF and PXY are still both encoded for within monocot genomes. This raises the question of what the function of TDIF and PXY is in monocots. To address this question, we deleted the PXY gene from barley. We found no changes to xylem or phloem formation, nor to radial growth, but we did observe fewer larger cells in barley mutant stems. Thus we hypothesised that barley PXY acts to promote cell divisions in a dividing cell population referred to as the intercalary meristem. Intercalary meristems predominate on monocots. They are areas of cell division within the stem that contribute to growth by adding length. The purpose of this proposal is to test the hypothesis that TDIF-PXY regulates the intercalary meristem in monocots. We propose to test this hypothesis by determining cell type specific expression domains of TDIF and PXY genes in barley stems. We already know that TDIF and PXY are expressed in stems, but we don't know in exactly which cell type. To perform this analysis we will use two methods. In the first, we will probe for mRNA derived from PXY and TDIF genes to visualise which cells it is present in. In the second, we will generate so-called reporter genes, where the DNA sequences that define where a gene is expressed (in this case, those for TDIF, PXY, and PXY-like genes) are placed upstream of a gene that can produce a coloured dye. We will further test our hypothesis by removing any TDIF-PXY redundancy from barley using genome editing. Redundancy is where several related genes can perform a similar function, such that mutating a single gene has only a minor effect on phenotype. We have already generated a pxy mutant but other closely related genes may contribute to PXY function. Removing those related genes may lead to clear changes to plant morphology which will help determine TDIF-PXY function in monocots. However, imaging changes to morphology in plant stems is challenging in barley. Thus to fully characterise cell- and tissue-specific changes we will use X-ray computer tomography. This method is a non-destructive technique for visualizing interior features within solid objects, and we propose to use a machine specifically developed for biological samples. This will allow us to fully characterise changes to morphology in the plant lines used in this proposal.Finally, we will determine the consequences of loss of TDIF-PXY on robust plant growth in challenging environmental conditions. Our preliminary data has suggested that TDIF-PXY is required to maintain robust growth under water limited conditions, so we will test the effectiveness of carbon assimilation in the plant lines generated on this proposal when limited amounts of water are available. Global heating is predicted to reduce water availability for crops. But understanding the relationships between plant development and physiology may provide indicators of how we can

我们的大多数粮食作物来自开花植物，被称为被子植物。被子植物进一步分为双子叶植物和单子叶植物，双子叶植物包括许多木本植物，单子叶植物包括重要的谷类作物，如小麦和大麦。单子叶植物和双子叶植物分支之间的主要区别之一是茎的组织。在双子叶植物中，茎的特征在于一组未分化的细胞，称为形成层，其负责大多数径向生长。形成层存在于茎周围的环中，并且在形成层内分裂的细胞最终形成茎中心处的木本植物组织（木质部）或朝向茎的外部的第二运输组织（韧皮部）。形成层中的细胞分裂由称为TDIF-PXY的信号传导机制控制。TDIF是一种小的配体蛋白，而PXY是一种定位于形成层细胞质膜的受体蛋白。当TDIF与PXY结合时，信号被传递到细胞核，促进细胞分裂。单子叶植物没有形成层。然而，TDIF和PXY仍然都在单子叶植物基因组内编码。这就提出了TDIF和PXY在单子叶植物中的功能是什么的问题。为了解决这个问题，我们从大麦中删除了PXY基因。我们没有发现木质部或韧皮部的形成，也没有径向生长的变化，但我们确实观察到更少的大麦突变体茎较大的细胞。因此，我们假设，大麦PXY的行为，以促进细胞分裂中的分裂细胞群体称为中间分生组织。在单子叶植物中，间生分生组织占优势。它们是茎内细胞分裂的区域，通过增加长度来促进生长。该提议的目的是检验TDIF-PXY调节单子叶植物中间分生组织的假设。我们建议通过确定大麦茎中TDIF和PXY基因的细胞类型特异性表达域来验证这一假设。我们已经知道TDIF和PXY在茎中表达，但我们不知道确切的细胞类型。为了进行这种分析，我们将使用两种方法。首先，我们将探测来自PXY和TDIF基因的mRNA，以观察它存在于哪些细胞中。在第二种方法中，我们将产生所谓的报告基因，其中定义基因表达位置的DNA序列（在这种情况下，TDIF，PXY和PXY样基因的DNA序列）被放置在可以产生有色染料的基因的上游。我们将通过使用基因组编辑从大麦中去除任何TDIF-PXY冗余来进一步测试我们的假设。Redundancy是几个相关基因可以执行类似功能的地方，例如突变单个基因对表型只有很小的影响。我们已经产生了一个pxy突变体，但其他密切相关的基因可能有助于PXY功能。去除这些相关基因可能导致植物形态的明显变化，这将有助于确定TDIF-PXY在单子叶植物中的功能。然而，在大麦中，对植物茎的形态变化进行成像是具有挑战性的。因此，为了完全确定细胞和组织特异性变化，我们将使用X射线计算机断层扫描。这种方法是一种非破坏性的技术，用于可视化固体物体内的内部特征，我们建议使用专门为生物样品开发的机器。这将使我们能够充分研究本提案中使用的植物品系的形态变化。最后，我们将确定TDIF-PXY缺失对植物在具有挑战性的环境条件下稳健生长的影响。我们的初步数据表明，TDIF-PXY需要在水分有限的条件下保持稳健的生长，因此，我们将在有限水量的情况下测试该提议产生的植物系中碳同化的有效性。据预测，全球变暖将减少农作物的可用水。但是，了解植物发育和生理学之间的关系可能会提供我们如何