ABA胁迫诱导茶树富集γ-氨基丁酸的代谢机理研究

Gamma amino butyric acid (GABA) a four carbon non-protein amino acid with an amino group attached to γ-carbon instead of α-carbon exists in different organisms including bacteria, plants and animals. A range of physiological and biochemical analyses from previous studies have unraveled the role of GABA and GABA metabolism. GABA metabolism has different functions in plant metabolism including osmotic and pH regulation, pollen tube growth and guidance, bypass of the tricarboxylic acid (TCA) cycle, C:N balance, nitrogen metabolism, and preventing the accumulation of reactive oxygen species under different environmental stress. Moreover, GABA, a well known neurotransmitter in mammalian brain tissues is also suggested to be a signaling molecule in plants. . In the previous work, Some genes about GABA shunt have been screened from the tea whole organ transcriptome database, and which have been induced by abscisic acid then to increase GABA levels in roots and leaves of tea. But the relationship between the two GABA synthesis pathways in tea are unclear, and the role of GABA metabolism in adversity stress responses in relation to the effect of nitrogen metabolism in tea are also unclear. Therefore, we considered that the inhibiting the GAD gene expression and supplying different exogenous nitrogen substrate would be good tools to study the role of GABA in nitrogen metabolism of tea plant in relation to the response of ABA stress. On the one hand, using the stable isotope labeling the substrate clarify the mutual relations between two GABA synthesis ways; On the other hand, together with metabolite and gene expression profiling analysis method, screening key nitrogen compounds and genes response to ABA stress in tea plant, illuminating the nitrogen metabolism model. This study will clarify the regulatory relationship between exogenous stress treatments and enrichment of GABA, then provide scientific basis for further enhance the development and utilization of tea plant resources.

γ-氨基丁酸不仅具有极高保健功能，还具有协助植物抵御环境胁迫，调节C-N代谢平衡等生理功能，但对于GABA在茶树体内合成途径的相互关系以及胁迫诱导茶树富集GABA过程对氮代谢的调节尚不明确。项目组对前期获得的转录组数据进行挖掘，获得GABA支路和多胺途径上多条序列，发现ABA能显著诱导茶树根和叶部GAD的表达并促进GABA含量的提高。本项目拟在前期研究基础上，通过抑制GAD基因表达和外源饲喂不同氮素底物这两种正反结合的研究手法，对ABA诱导下茶树应激机制及氮代谢进行研究，一方面利用稳定同位素标记底物对GABA两条合成途径的相互关系进行解析；一方面结合代谢组与表达谱的分析方法，筛选出ABA诱导下茶树以GABA为节点的氮代谢过程中的关键含氮化合物和基因，解析氮代谢模式。本研究将明确逆境诱导茶树体内GABA合成的关键途径与调节氮平衡的分子机理，为茶树栽培与资源开发提供科学依据。

γ-氨基丁酸（GABA）是一类四碳非蛋白氨基酸，广泛存在于哺乳动物体内，具有抑制神经递质的作用，因而对人体具有广泛的健康功效。同时，研究表明，其代谢过程与植物响应多种生物与非生物胁迫相关，对维持植物碳氮代谢平衡起到重要的作用。但对于GABA在茶树体内合成途径的相互关系以及胁迫诱导茶树富集GABA过程对氮代谢的调节尚不明确。本项目前期主要针对ABA诱导茶树快速积累GABA的调控过程进行研究，结果发现在采用厌氧处理方式快速增加茶鲜叶中GABA含量制备“Gabaron Tea”的过程中，也检测到ABA有应激性增加，因此转为研究不同厌氧处理（充N2和充CO2）过程中，茶叶片积累GABA响应胁迫应激的调节机制。本研究采用代谢组学（UPLC-QTOF-MS）与转录组（qRT-PCR）整合的检测方法，通过比较厌氧胁迫下茶树叶片以GABA为节点的碳氮代谢过程中的关键化合物的动态变化，筛选出主要代谢通路和差异基因，并解析基因表达调控模式，结果从代谢组学水平和核酸水平揭示了离体茶树叶片响应不同厌氧胁迫应激时采取的差异响应模式和动态变化过程。本研究将明确逆境诱导茶树体内GABA合成的关键途径与调节碳氮平衡的分子机理，为茶树基因工程育种，以及生产上指导合理合宜的“Gabaron Tea”制备提供重要的理论与实践基础。