细胞质甘油醛-3-磷酸脱氢酶在ABA和蔗糖协同调控草莓果实成熟中的机理研究

Strawberry is becoming a model for studying non-climacteric fruit. As fewer fruit are produced in winter, the “winter strawberry” is gaining attention from farmers because of its higher economic value. Abscisic acid (ABA) and sucrose play important roles in the regulation of strawberry fruit ripening. To further understanding the physiological and molecular mechanisms of strawberry fruit ripening, ABA, sucrose or ABA plus sucrose applications in the field experiment had been conducted first. The results showed that all the above three treatments can accelerate strawberry fruit ripening, and the mixed treatment was the best. Then, RNA-sequence analysis of the treated fruit transcriptome revealed only seven genes were specific response to the mixed treatment significantly among the differentially expressed genes. Furthermore, four of them were key enzymes in glycolysis pathway, and the differentially expression of cytoplasmic glyceraldehydes-3-phosphate dehydrogenase gene among them was the best. Based on the results, it could be indicated that glycolysis was involved in regulating strawberry fruit ripening through the synergistic effect of sucrose and ABA. However, information on the role of glycolysis in fruit development and ripening is limited. To analyse the role of glycolysis in strawberry fruit ripening, followed by the preliminary studies, we focused on cytoplasmic glyceraldehydes-3-phosphate dehydrogenase gene. In this study, gene cloning, subcellular localizationa, tobacco transient expression system, gene silencing and gene overexpression techniques in strawberry fruit were used to verify the biological function of glycolysis key enzyme cytoplasmic glyceraldehydes-3-phosphate dehydrogenase.

草莓是研究非呼吸跃变的典型材料。在生产上，“冬草莓”售价较高，因此其冬季产量与农户种植效益密切相关。ABA和蔗糖是调控草莓果实成熟的两个关键因素。我们前期研究发现，田间喷施ABA和蔗糖能有效缩短草莓果实成熟期至少4~12天，因此是一种提高草莓冬季产量的潜在方法。转录组进一步分析发现，仅有7个基因显著地特异响应ABA和蔗糖混合处理，其中4个都是糖酵解途径关键酶，尤以GAPC2对处理响应变化最为明显。由此说明，糖酵解参与了ABA和蔗糖协同调控草莓果实成熟的进程。为进一步揭示糖酵解在草莓果实成熟中的关键作用，本项目拟以GAPC2为突破口，通过基因克隆、亚细胞定位、基因沉默和过表达技术获得转基因植株并对其进行果实成熟度、GAPC氧化程度、代谢组以及转录组相关分析，系统阐明ABA和蔗糖协同调控GAPC2促进草莓果实成熟的作用机制，以期拓宽对草莓果实成熟调控机理的认识以及草莓冬季产量的增加和农民增收。

草莓是研究非呼吸跃变的典型材料。在生产上，“冬草莓”售价较高，因此其冬季产量与农户种植效益密切相关。ABA和蔗糖是调控草莓果实成熟的两个关键因素。我们前期研究发现，田间喷施ABA和蔗糖能有效缩短草莓果实成熟期至少4~12天，因此是一种提高草莓冬季产量的潜在方法。转录组进一步分析发现，仅有7个基因显著地特异响应ABA和蔗糖混合处理，其中4个都是糖酵解途径关键酶，尤以GAPC2对处理响应变化最为明显。由此说明，糖酵解参与了ABA和蔗糖协同调控草莓果实成熟的进程。为进一步揭示糖酵解在草莓果实成熟中的关键作用，本项目拟以GAPC2为突破口，通过基因克隆、亚细胞定位、基因沉默和过表达技术获得转基因植株并对其进行果实成熟度、GAPC氧化程度、代谢组以及转录组相关分析，系统阐明ABA和蔗糖协同调控GAPC2促进草莓果实成熟的作用机制，以期拓宽对草莓果实成熟调控机理的认识以及草莓冬季产量的增加和农民增收。研究结果表明：FaGAPC2转录本在果实中显示出高转录本水平。FaGAPC2的瞬时过表达延迟了果实成熟，而RNAi干扰促进了果实成熟并影响了果实花青素和蔗糖水平。FaGAPC2表达模式的变化也影响了几个糖酵解相关和成熟相关基因的表达，如CEL1、CEL2、SS、ANS、MYB5、NCED1、ABI1、ALDO、PK和G6PDH，以及H2O2水平和还原型谷胱甘肽(GSH)/谷胱甘肽二硫化物(GSSG)氧化还原态势。同时，代谢组学实验表明，FaGAPC2的瞬时过表达导致花青素、黄酮类化合物、有机酸、氨基酸及其衍生物的减少。此外，脱落酸（ABA）和蔗糖处理诱导大量H2O2的产生并抑制草莓果实中FaGAPC2的表达。这些结果表明，FaGAPC2是ABA和蔗糖介导的果实成熟的负调节因子，可通过氧化应激进行草莓果实成熟的调节。