Wu Yingying, Liu Jinfeng, Sheng Xinyuan, Wang Wenqiu, Wang Tianchi, Martinez-Sanchez Marcela, Wang Songhu, Tu Meiyan, Deng Jiahui, Allan Andrew C, Atkinson Ross G, Nieuwenhuizen Niels J, Yin Xueren, Zeng Yunliu
One Belt and One Road International Joint Research Center of Horticultural Products Quality and Post-Harvest Biotechnology in Anhui Province, School of Horticulture, Anhui Agricultural University, Hefei, Anhui, China.
Anhui Province Key Laboratory of Horticultural Crop Quality Biology, School of Horticulture, Anhui Agricultural University, Hefei, Anhui, China.
Plant Biotechnol J. 2025 Jul;23(7):2554-2569. doi: 10.1111/pbi.70071. Epub 2025 Apr 4.
Changes in skin colour, as a visual cue for fruit ripeness, are important physiological markers in many crops including tomato, banana and grape. In kiwifruit, the skin remains brown during ripening, but de-greening of the pericarp occurs to reveal accumulated carotenoids and anthocyanins in gold- and red-fleshed cultivars. In this study, analysis of the inner and outer pericarp of Actinidia chinensis 'Hongyang' revealed faster chlorophyll degradation in the inner pericarp, compared with the outer pericarp. Based on transcriptome analysis, two chlorophyll degradation-related genes encoding Mg-dechelatases (AcSGR1 and AcSGR2) were more abundantly expressed in the inner pericarp, and this correlated with higher Mg-dechelatase enzyme activity in the inner pericarp than in the outer pericarp. Weighted gene co-expression network analysis identified potential regulators of AcSGR1/2. A differentially expressed NAM/ATAF/CUC transcription factor AcNAC2 was identified, which could directly interact with AcSGR1 and AcSGR2 promoters and strongly activate their expression. A closely related NAC, AcNAC3, also enhanced AcSGR1/2 expression, but was less abundantly expressed. Transient expression in tobacco confirmed that AcNAC2 and AcNAC3 promote chlorophyll degradation, and stable overexpression in kiwifruit verified that AcNAC2 acts via up-regulation of AcSGR1/2 gene expression. CRISPR-mediated knockouts of AcNAC2/3 in kiwifruit dramatically reduced expression levels of AcSGR1/2 genes in fruit, leading to significantly delayed chlorophyll degradation and de-greening. Together, these results suggest that differential chlorophyll degradation drives the differences observed in chlorophyll content between the inner and outer pericarp of kiwifruit, which is principally modulated by the transcription factor AcNAC2.
果实成熟时的皮肤颜色变化，作为一种视觉线索，是包括番茄、香蕉和葡萄在内的许多作物的重要生理指标。在猕猴桃中，果实成熟时表皮仍为褐色，但果肉会脱绿，从而在金果肉和红果肉品种中显现出积累的类胡萝卜素和花青素。在本研究中，对中华猕猴桃‘红阳’的内果皮和外果皮进行分析发现，与外果皮相比，内果皮中的叶绿素降解更快。基于转录组分析，两个编码镁脱螯合酶的叶绿素降解相关基因（AcSGR1和AcSGR2）在内果皮中表达更为丰富，这与内果皮中比外果皮更高的镁脱螯合酶活性相关。加权基因共表达网络分析确定了AcSGR1/2的潜在调控因子。鉴定出一个差异表达的NAM/ATAF/CUC转录因子AcNAC2，它可以直接与AcSGR1和AcSGR2启动子相互作用并强烈激活它们的表达。一个密切相关的NAC基因AcNAC3也增强了AcSGR1/2的表达，但表达量较低。在烟草中的瞬时表达证实AcNAC2和AcNAC3促进叶绿素降解，在猕猴桃中的稳定过表达验证了AcNAC2通过上调AcSGR1/2基因表达发挥作用。利用CRISPR技术敲除猕猴桃中的AcNAC2/3，显著降低了果实中AcSGR1/2基因的表达水平，导致叶绿素降解和脱绿明显延迟。总之，这些结果表明，叶绿素降解差异导致了猕猴桃内果皮和外果皮叶绿素含量的差异，而这主要受转录因子AcNAC2的调控。
