Mechanisms of proximo-distal polarity establishment and tissue patterning in Hibiscus petals

开花植物花瓣上的彩色图案是吸引传粉者和直接参与物种繁殖成功的关键。这些模式通常非常复杂，结合了色素沉着、细胞形状和角质层纹饰的差异，因此它们的发育必须受到严格调节，以产生具有不同特性的邻近组织。当花瓣表皮生长时，如此清晰的边界是如何在花瓣表皮内建立起来的仍有待了解。剑桥大学塞恩斯伯里实验室（Sainsbury Laboratory）的Moyroud小组开发了一种木槿，作为一种新的模型系统，以了解花瓣上的图案是如何形成的。利用转录组学方法，他们确定了在花瓣形成模式之前的早期发育阶段，其表达仅限于花瓣不同部分的基因。这些构成了可能作为模式形成上游调控者的候选基因。拟议的项目将旨在(i)测试一个关于组织花瓣基部到尖端极性的信号性质的新假设，（ii）探索花瓣中细胞极性和组织极性之间的相互作用，以及（iii）确定下游基因调控网络(s)负责模式形成。为此，一系列分子生物学技术将与新兴模型系统中最先进的遗传操作相结合（克隆，定量PCR，构建过表达和CRISPR/Cas9敲除的木槿系，生产表达感兴趣代谢物荧光传感器的转基因植物和转录组学）。该项目还将涉及外部分析：使用定制的体外花瓣生长系统，将候选分子应用于植物芽发育或分离花瓣上，以评估其干扰花瓣图案的能力。最后，将使用共聚焦、拉曼和扫描电子显微镜方法分析处理过的芽/花瓣、转基因系和最近在遗传筛选中发现的模式形成缺陷的突变体的表型。

PhD project strategic theme: Understanding the rules of life The colourful patterns on the petal of flowering plants are key to attracting pollinators and participate directly in the reproductive success of a species. These patterns are often highly elaborate and combine differences in pigmentation, cell shape and ornamentation of the cuticle so their development must be tightly regulated to generate neighbouring tissues with distinct properties. How such clear boundaries are established within the petal epidermis as it grows remains to be understood.The Moyroud group at the Sainsbury Laboratory (University of Cambridge) has developed a species of Hibiscus as a new model system to understand how those patterns form on petals. Using a transcriptomic approach, they identified genes whose expression is restricted to distinct parts of the petal in very early stages of development, before pattern formation. Those constitute candidate genes likely to act as upstream regulators of pattern formation.The proposed project will aim at (i) testing a new hypothesis regarding the nature of the signal that organises the petal base-to-tip polarity, (ii) exploring the interplay between cell polarity and tissue polarity in petals, and (iii) identifying the downstream gene regulatory network(s) responsible for pattern formation. To do so, a range of molecular biology techniques will be used in combination with state-of-the art genetic manipulations in an emerging model system (cloning, quantitative PCR, construction of overexpression and CRISPR/Cas9 knock-out Hibiscus lines, production of transgenic plants expressing fluorescent sensors for metabolites of interest and transcriptomics). This project will also involve external assays: candidate molecules will be applied to developing buds in planta or on isolated petals using a custom-made in vitro petal growth system to assess their ability to interfere with petal patterning. Finally, confocal, Raman and scanning electron microscopy methods will be used to analyse phenotypes of treated buds/petals, transgenic lines and mutants with defect in pattern formation recently identified in a genetic screen.