GIGANTEA基因响应干旱胁迫调控生殖生长的生理与分子机制

Moderate drought stress is one of the inducing factors of flower bud differentiation in fruit trees. It can not only promote normal flowering, but also induce off-season flowering. At the same time, it can also increase the quantity and quality of flowers, and achieve the purpose of changing the maturity of the fruit and improving the quality. The GIGANTEA (GI) gene encodes a specific nuclear protein, which is an important flowering regulator in the long-day flowering pathway, and participates in circadian rhythm regulation of flowering and stress-induced flowering. Our previous research found that GI not only responds to drought stress but also participates in citrus flowering. This project intends to analyze the function and regulation mechanism of citrus GI gene, and isolate the upstream and downstream regulatory genes by yeast one-hybrid and yeast two-hybrid, and transgenic knockout and overexpression to study the function of the gene, and then reveal the role of this gene in the process of citrus flowering and resistance to drought stress, follow the interaction of upstream and downstream genes, find more reproductive-related genes and reveal the mechanism of gene and environment, gene and gene network regulation. The results have important theoretical significance for people to understand the regulation mechanism of stress on plant flowering process from the molecular level. Meanwhile, it can also provide theoretical basis for citrus drought resistance mechanism, water saving irrigation, flower regulation, unseasonal cultivation and high yield cultivation techniques.

适度的干旱胁迫是果树花芽分化的诱导因子之一，其不仅可以促进正常开花，而且可以诱导反季节开花，同时还可以提高花的数量和质量，进而改变成熟期和提高果实品质。GIGANTEA（GI）基因编码一种特异性核蛋白，在长日照开花途径中作为重要的开花调控因子，参与生理节律开花与逆境调控。我们前期研究发现GI不仅响应干旱胁迫而且还参与柑橘成花。本项目拟对柑橘GI基因的功能及其调控机理进行解析，以酵母单双杂交分离上下游的调控基因，转基因敲除及超量表达研究该基因功能，进而揭示该基因在柑橘成花及抵抗干旱胁迫过程的作用，顺着上下游基因的相互作用，找到更多生殖发育相关的基因并揭示基因与环境、基因与基因网络调控的机理，对人们从分子水平上认识和理解逆境对植物生殖发育过程的调控机制具有重要的理论意义。同时也可为柑橘的抗旱机理、节水灌溉、调控开花、错季栽培及丰产栽培技术提供理论依据。

适度的干旱胁迫是果树花芽分化的诱导因子之一。我们前期研究发现GIGANTEA（GI）不仅响应干旱胁迫而且还参与柑橘成花。本项目主要基于上述线索完成了如下内容：CiGI在柑橘中存在CiGIa和CiGIβ两个选择性剪切本，遗传转化烟草发现，CiGIa具有早花作用并且增强了转基因植物对干旱的敏感性，而CiGIβ没有表现出干旱和开花的功能；进一步在柠檬中干涉与超表达该基因，其结果与烟草一致，这些结果表明CiGIa在干旱诱导柑橘开花的过程中充当关键角色；通过GUS组织化学染色和GUS活性定量发现CiGI的启动子也受干旱诱导并且发现CiFD可以结合到CiGI的启动子上，有意思的是CiFD也发生了选择性剪切并产生CiFDa和CiFDβ两个剪切本，转化烟草和柑橘发现CiFD的两个剪切本都可以促进转基因烟草和柑橘开花，说明CiFD在柑橘开花的过程中也充当了关键角色；利用酵母和Pull-down实验证实了CiLHY与CiGI蛋白互作，并且发现CiLHY可以结合到CiNF-YA1的启动子上促进其表达，进而增强柑橘对干旱的敏感性；同时CiLHY也与CiFT结合促进柑橘开花。基于上述结果，提出了在干旱条件下，CiGI调控柑橘生殖生长的模型：干旱胁迫下，CiFD被干旱诱导并结合到CiGI的启动子上，促进CiGIa的表达，随后招募大量的CiLHY蛋白与之形成复合物，CiGIa-CiLHY复合物可以直接结合到CiFT的启动子并且激活其表达，促使植物提早开花；CiGIa-CiLHY复合物也可以直接作用于CiNF-YA1的启动子激活其表达，在干旱逃逸中发挥作用。实验结果为柑橘的抗旱机理、调控开花及错季栽培提供了理论依据。