胰蛋白酶被发现具有超氧阴离子清除活性,可以显著诱导果实采后抗病性和抗逆性。先前研究发现DELLA蛋白的编码基因HuGAI1应是胰蛋白酶诱导火龙果果实抗性的关键基因,但其调控机制尚需阐明。为了研究胰蛋白酶诱导火龙果抗性的分子机制,该研究通过病毒诱导的基因沉默(virus-induced gene silencing, VIGS)技术构建HuGAI1基因的RNA沉默株,验证其在火龙果的保鲜作用,又通过转录组学分析HuGAI1调控的下游代谢途径。结果显示,胰蛋白酶处理的火龙果果实保存时间延长,HuGAI1沉默后胰蛋白酶的诱抗效果丧失,火龙果果实腐败加剧。HuGAI1沉默后的转录组学分析结果显示,HuMYB12应是与HuGAI1结合的关键转录因子,它参与了类黄酮物质的合成,且受HuGAI1-HuMYB12调控的基因主要富集在类黄酮生物合成途径中,HuCYP98A3、HuPAL1和Hu4CL3等下游多个关键酶的表达显著上调。胰蛋白酶通过上调HuGAI1-HuMYB12的表达,促进黄酮类化合物的生物合成,增强细胞抗氧化能力,延长果实保鲜时间。
Trypsin has been found to have superoxide anion scavenging activity, which significantly induced disease resistance and stress resistance in Hylocereus undatus fruits after harvest.Previous research found that HuGAI1, the gene encoding DELLA protein, should be a key gene in fruit resistance induced by trypsin, but its regulatory mechanism needs to be elucidated.In order to study the molecular mechanism of trypsin-induced resistance in H.undatus fruits, the study constructed the RNA silencing strain of HuGAI1 gene by virus-induced gene silencing (VIGS) technology to verify its effect on the preservation of H.undatus, and analyzed the downstream metabolic pathways regulated by HuGAI1 through transcriptomic analysis.The results showed that the storage time of H.undatus treated with trypsin was extended.However, the trypsin-induced resistance effect was lost after HuGAI1 was silenced, which led to increased spoilage of H.undatus fruits.The transcriptomic analysis of HuGAI1-silenced samples indicated that HuMYB12 was a key transcription factor interacting with HuGAI1, involved in flavonoid biosynthesis, and genes regulated by the HuGAI1-HuMYB12 complex were primarily enriched in the flavonoid biosynthesis pathway.The expression of several downstream key enzymes, such as HuCYP98A3, HuPAL1, and Hu4CL3, was significantly upregulated.Trypsin had extended the shelf life of the fruit by up-regulating the expression of HuGAI1-HuMYB12, promoting the biosynthesis of flavonoid compounds, and enhancing cellular antioxidant capacity.
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