菊花CmYAB1调控花瓣发育的分子机制

Flower is one of the most important ornamental organ of ornamental plants, and the shape, size and quantity of petals directly affect ornamental. Petal development is not only the focus of horticulture researches, but also one of the hotspots of developmental biology. The inflorescences of Chrysanthemum (Chrysanthemum morifolium), which is one of top ten traditional flowers in China and one of the four major cut flowers all over the world, are generally composed of two different type of florets (ray and disk) that tightly packed into a varied capitulum as the rich petal morphology. Thus, Chrysanthemum is the ideal material for the researches of development petals, but the molecular mechanism of the petal development the is largely unknown. We previous results showed that the expression level of CmYAB1 was different in petals between ray and disk florets. Here we aimed to explore the temporal and spatial expression pattern of CmYAB1 by RNA in situ hybridization and gene function in petal development via genetic transformation in chrysanthemum. In addition, the transcriptome profiles in transgenic plants will be investigated via RNA-seq, and genes regulated downstream by CmYAB1 were screened using bioinformatics analysis. Moreover, protein interaction analysis will be performed via yeast two hybridization. Taking all these experimental data, we will identify the function and shed light on the mechanism of CmYAB1 involved in the petal development, which in turn will be aid to genetic manipulation of flower type in chrysanthemum. The findings of present research would also provide clues for dissecting molecular mechanisms involved in flower development in other ornamental plants.

花是观赏植物最重要的观赏部位之一，其中花瓣的形状、大小和数量等直接影响其观赏性；花瓣发育研究不仅是园艺学关注的焦点，也是发育生物学研究的热点之一。菊花作为我国十大传统名花和世界四大切花之一，其瓣型极为丰富，且同一个花序中通常既有舌状花又有管状花，是研究花瓣发育的理想材料，但目前菊花花瓣发育的分子机制尚不明晰。我们前期的研究表明，CmYAB1的表达量与花瓣形状有关，在舌状花和管状花花瓣中差异表达。本项目拟以菊花为试材，利用RNA原位杂交技术明确CmYAB1的时空表达情况；构建超表达载体和干扰载体对菊花进行遗传转化，解析其在菊花花瓣发育中的功能；利用转录组测序和生物信息学技术分析其参与的调控网络；通过酵母双杂交技术筛选其互作蛋白，并利用BiFC和Pull-down技术进行验证。本项目的实施可揭示菊花花瓣发育的分子机制，为菊花瓣型的分子改良奠定基础，也可为显花植物的花瓣形态建成机理研究提供借鉴。

菊花作为我国十大传统名花和世界四大切花之一，深受人们的喜爱。作为菊科植物，它具有典型的头状花序，外轮为艳丽多姿的舌状花，内轮通常为黄色的管状花。舌状花根据花瓣融合程度的不同可分为平瓣、匙瓣和管瓣等类型。其丰富多样的瓣型是研究花瓣发育的理想材料，但目前菊花花瓣发育特别是调控花瓣融合的分子机制尚不清楚。我们前期的研究表明，YABBY家族基因CmYAB1在舌状花和管状花花瓣中差异表达，可能参与花瓣形态发育。本项目以切花菊‘神马’为研究对象，对CmYAB1的功能和调控机制进行了研究。RNA原位杂交试验表明CmYAB1在分生组织中表达且在所有侧生器官的远轴端特异表达。构建超表达载体和干扰表达载体对菊花进行遗传转化，结果发现转基因植株花瓣的融合程度发生改变，表明CmYAB1参与调控菊花花瓣的融合。通过对干扰转基因株系进行转录组测序和RT-PCR验证，初步发现边界基因CmCUC2的同源基因在两个干扰株系中显著降低。对其启动子进行克隆并采用生物信息学分析预测出5个CmYAB1的结合元件，进一步通过酵母单杂、EMSA、ChIP-qPCR、LUC等实验验证出CmYAB1能够直接结合CmCUC2的启动子，并促进CmCUC2的表达。另外，通过酵母双杂交技术筛选到其互作蛋白CmYAB3.1，并利用BiFC和Pull-down等技术验证了CmYAB1与CmYAB3.1蛋白互作，且双荧光素酶实验证明了CmYAB1与CmYAB3.1蛋白互作后增强了对下游基因CmCUC2的促进作用。本项目阐释了CmYAB1调控菊花花瓣融合的分子机制，CmYABs形成蛋白复合体调控边界基因CmCUC2表达，进而调控菊花花瓣的形态建成，为菊花瓣型的分子改良奠定基础，也可为显花植物的花瓣形态建成机理研究提供借鉴。