VvYABBY-5 基因调控葡萄多心皮子房形成分子机制研究

Grape fruit generally has two carpels which encloses two ovules respectively. However, a few cultivers produce fruits with three or four carpels in natural condition. Comparing to the two carpels, Fruit size, bunch weight and yield of multiple carpels are be improved. However, the mechanism of the increase in carpel number is still unknown and the study for this mechanism will offer the insights into physiologic regulation and breeding big fruit grape. In this work, we are ready to use "xiangfei" grape as the plant material because of the high three carpel percentage in this cultivar. Firstly, the spatiotemporal expression of the candidate gene VvYABBY-5 which controlling the carpel number during development of the ovaries with different number carpel (two, three and four carper) will be analyzed by Realtime-PCR and In situ hybridization. Secondly, the role of VvYABBY-5 in multiple carpels forming will be analyzed by overexpressing and silencing VvYABBY-5 in "xiangfei" grape and calculating multiple carpel percentage in transgenic grape. Finally, the functional genes those are the downstream targets of VvYABBY-5 will be found through transcriptome profiling and chromatin immunoprecipitation. Then the role of these genes will be analyzed by transgenic technology. The aim of this proposal is to reveal the he genetic networks in controlling the formation of multiple carpel.

葡萄果实一般为2心皮4胚珠植物，但极少数品种在自然条件下果实可以发育成多心皮（3-4心皮）。多心皮比2心皮果粒大，果穗重，产量高，具有重要的生产价值，但目前有关多心皮报道极少，其形成机理尚不清晰，而研究多心皮发生机制能为大果粒育种及生理调控提供依据。本研究以多心皮发生率较高的葡萄香妃为试材，在前期研究基础上，首先利用realtime-PCR和原位杂交对心皮发生有重要作用的VvYABBY-5基因在不同心皮数和发育阶段的子房进行时空表达分析，明确VvYABBY-5基因表达与多心皮子房的相关关系；其次转基因增强和抑制VvYABBY-5基因表达，观察转基因植株心皮变化，确定VvYABBY-5基因调控心皮数目的作用；最后利用转录组测序结合染色质免疫共沉淀筛选鉴定被转录因子VvYABBY-5基因调控的下游功能基因，转基因研究其功能。最终揭示以VvYABBY-5基因为核心的调控葡萄多心皮发生的分子机制。

大多数葡萄品种果实为2心皮4胚珠植物，但极少数品种在自然条件下果实可以发育成多心皮（3-4心皮）。多心皮比2心皮果粒大，果穗重，产量高，具有重要的生产价值。研究多心皮果实的发生机制能为大果粒育种提供依据。本研究以多心皮发生率较高的葡萄品种香妃为试材，研究了3心皮果实的形成途径，对不同心皮子房进行了转录组测序，根据测序结果分析了 VvYABBY家族基因、VvSUP、VvAG1和VvSEP3基因在不同心皮形成和发育阶段的作用。研究结果显示，‘香妃’葡萄3心皮子房发生比例多年稳定在40%左右，心皮形成的主要方式是先天融合，后天融合的比例较低。3心皮果粒单粒重大于2心皮果粒20%以上，种子数也多。VvSUP基因在心皮形成过程中发挥作用，其作用部位在心皮原基形成的边界，过表达VvSUP转基因番茄植株表现为心皮数目减少。VvSUP基因可能通过与下游的VvWUS和VvAG1基因共同调控心皮形成。VvYABBY5基因主要在心皮发育过程中起作用，转VvYABBY5基因番茄使其开花提前，但对心皮数目没有影响。VvYABBY5基因在开花前的3心子房中表达量少于2心皮子房，但其启动子的甲基化程度并不影响其表达。VvYABBY5蛋白与VvAG1，VvSEP3和VvSEP3蛋白存在着物理互作。说明VvYABBY5是通过与VvAG1，VvSEP3和VvSEP3形成蛋白复合体来影响葡萄心皮子房的发育。此外转VvSEP3和VvAG基因番茄都可引起开花性状的改变。