Mackon Enerand, Guo Yongqiang, Jeazet Dongho Epse Mackon Guibeline Charlie, Ma Yafei, Yao Yuhang, Luo Dengjie, Dai Xianggui, Zhao Neng, Lu Ying, Jandan Tahir Hussain, Liu Piqing
State Key Laboratory of Conservation and Utilization of Subtropical Agro-Bioresources, College of Life Science and Technology, Guangxi University PR China.
State Key Laboratory of Conservation and Utilization of Subtropical Agro-Bioresources, College of Agriculture, Guangxi University, Nanning 530005, PR China.
Phytochemistry. 2024 Jan;217:113896. doi: 10.1016/j.phytochem.2023.113896. Epub 2023 Oct 20.
Anthocyanins are a flavonoid compound known as one of the most important chromogenic substances. They play several functions, including health promotion and sustaining plants during adverse conditions. They are synthesized at the endoplasmic reticulum and sequestered in the vacuole. In this work, we generated knock-out lines of OsGSTU34, a glutathione transporter's tau gene family, with no transgene line and off-target through CRISPR/Cas9 mutagenesis and highlighted the loss of pigmentation in rice flowers, leaves, stems, shoots, and caryopsis. The anthocyanin quantification in the wild-type BLWT and mutant line BLG34-8 caryopsis showed that cyanidin-3-O-glucoside (C3G) and peonidin-3-O-glucoside (P3G) were almost undetectable in the mutant line. A tandem mass tag (TMT) labeling proteomic analysis was conducted to elucidate the proteomic changes in the BLWT and BLG34-8. The result revealed that 1175 proteins were altered, including 408 that were down-regulated and 767 that were upregulated. The accumulation of the OsGSTU34-related protein (Q8L576), along with several anthocyanin-related proteins, was down-regulated. The enrichment analysis showed that the down-regulated proteins were enriched in different pathways, among which the phenylpropanoid biosynthesis pathway, flavonoid biosynthesis metabolites, and anthocyanin biosynthesis pathway. Protein interaction network prediction revealed that glutathione-S-transferase (Q8L576) was connected to the proteins involved in the flavonoid and anthocyanin biosynthesis pathways, such as flavanone 3-dioxygenase 1 (Q7XM21), leucoanthocyanidin dioxygenase 1 (Q93VC3), 4-coumarate-CoA ligase 2 (Q42982), phenylalanine ammonia-lyase (P14717), chalcone synthase 1 (Q2R3A1), and 4-coumarate-CoA ligase 5 (Q6ZAC1). However, the expression of the most important anthocyanin biosynthesis gene was not altered, suggesting that only the transport mechanism was affected. Our findings highlight new insight into the anthocyanin pigmentation in black rice and provide new perspectives for future research.
花青素是一种黄酮类化合物，被认为是最重要的显色物质之一。它们具有多种功能，包括促进健康以及在不利条件下维持植物生长。花青素在内质网中合成，并被隔离在液泡中。在这项研究中，我们通过CRISPR/Cas9诱变技术获得了谷胱甘肽转运蛋白tau基因家族的OsGSTU34基因敲除系，且无转基因系和脱靶现象，并着重指出了水稻花、叶、茎、芽和颖果中色素沉着的缺失。对野生型BLWT和突变系BLG34 - 8颖果中的花青素进行定量分析表明，突变系中几乎检测不到矢车菊素 - 3 - O - 葡萄糖苷（C3G）和芍药素 - 3 - O - 葡萄糖苷（P3G）。进行了串联质谱标签（TMT）标记蛋白质组学分析，以阐明BLWT和BLG34 - 8中的蛋白质组变化。结果显示，有1175种蛋白质发生了改变，其中408种蛋白质表达下调，767种蛋白质表达上调。与OsGSTU34相关的蛋白质（Q8L576）以及几种与花青素相关的蛋白质的积累均下调。富集分析表明，下调的蛋白质在不同途径中富集，其中包括苯丙烷生物合成途径、类黄酮生物合成代谢物和花青素生物合成途径。蛋白质相互作用网络预测显示，谷胱甘肽 - S - 转移酶（Q8L576）与参与类黄酮和花青素生物合成途径的蛋白质相连，如黄烷酮3 - 双加氧酶1（Q7XM21）、无色花青素双加氧酶1（Q93VC3）、4 - 香豆酸 - CoA连接酶2（Q42982）、苯丙氨酸解氨酶（P14717）、查尔酮合酶1（Q2R3A1）和4 - 香豆酸 - CoA连接酶5（Q6ZAC1）。然而，最重要的花青素生物合成基因的表达并未改变，这表明只有转运机制受到了影响。我们的研究结果为黑米中花青素色素沉着提供了新的见解，并为未来的研究提供了新的视角。
