Identification and Expression Analysis of Key Gene Families in Flavonoid Metabolism Pathway in Pigeon Pea (Cajanus cajan)
DU Ting-Ting1,*, SONG Zhi-Hua1,*, DONG Bi-Ying1, CAO Hong-Yan1, LIU Teng-Yue1, YANG Wan-Long1, QI Meng1, CHEN Ting1, WANG Meng-Ying1, MENG Dong1,2, YANG Qing1,2,**, FU Yu-Jie1,2,3,**
1 College of Forestry, Beijing Forestry University, Beijing 100083, China; 2 Beijing Advanced Innovation Center for Tree Breeding by Molecular Design, Beijing Forestry University, Beijing 100083, China; 3 Key Laboratory of Forestry Plant Ecology, Ministry of Education, Northeast Forestry University, Harbin 150040, China
Abstract：Flavonoids are important secondary metabolites in plants, and they play an important role in plant growth and resistance to adversity and stress. Studies on the function and metabolic regulation of plant flavonoids (flavonoids, flavonols, etc.) have attracted more and more attention, especially in woody plants rich in flavonoid metabolites. The research on the synergistic interaction between key enzymes in flavonoid metabolic pathway will provide guidance for the better development and utilization of plant flavonoid functional components. Pigeon pea (Cajanus cajan) is a pioneer woody legume tree growing in karst soils with high acid aluminum content, which is rich in secondary metabolites such as flavonoids. In order to identify the key pathways of flavonoid biosynthesis, this study screened and identified 8 key enzyme gene families in pigeon pea, namely phenylalanine ammonia-lyase (PAL), cinnamic acid-4-hydroxylase (C4H), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonol synthase (FLS) and dihydroflavonol 4-reductase (DFR). Bioinformatics software such as MEGA 7.0, MEME, TBtools v1.089 and PlantCare were used to analyze the phylogeny, gene structure, protein structure and cis-acting elements of the promoter, as well as the regulation of gene expression in the metabolic pathway under stress conditions. Phylogenetic analysis showed that some members of the PAL, C4H, 4CL and CHI gene families could cluster together with the genes in soybean (Glycine max), while the CHS, FLS and DFR gene families only showed sequence similarity among family members. Protein structure analysis showed that each gene family contained a special conserved domain. Analysis of promoter cis-acting elements showed that the genes containing methyl jasmonate (MeJA) response elements accounted for more than 50% of the total genes in 6 gene families. Meanwhile, in view of the habitat characteristics of pigeon pea, RNA-seq analysis was conducted on the 8 gene families under aluminum stress and methyl jasmonate treatment in the study. The results showed that CcPAL2, CcC4H1, CcCHS5~7 and CcCHS12 with the higher relative expression levels were all significantly up-regulated under aluminum stress, while CcCHI4~5 and CcFLS7 all showed a significant down-regulation trend, indicating that chalcone in pigeon pea might accumulate under aluminum stress. It was speculated that chalcone might play a role in aluminum resistance. Under MeJA treatment, CcC4H1, CcCHS5, CcCHS7, and CcCHI4 were significantly up-regulated, while CcF3H was down-regulated 0.6 times that of the control. At the same time, CcFLS1, CcFLS5, and CcFLS6 in the downstream pathway were significantly up-regulated. The above results indicated that MeJA may also promote the accumulation of naringin and flavonol, it was speculated that the substances might play an important role in the MeJA signaling pathway. The expression changes of the above flavonoid biosynthetic enzyme genes at different stages were verified by qPCR and consistent with the transcriptome sequencing results.This study reveals the expression patterns of key enzymes in flavonoid biosynthesis under different stresses and provides reference for further exploration of the regulatory role of flavonoid substances such as chalcones and flavonols in stress.
杜婷婷, 宋治华, 董碧莹, 曹红燕, 刘腾跃, 杨琬珑, 祁萌, 陈婷, 王梦莹, 孟冬, 杨清, 付玉杰. 木豆类黄酮代谢通路关键基因家族的鉴定与表达分析[J]. 农业生物技术学报, 2021, 29(12): 2289-2303.
DU Ting-Ting, SONG Zhi-Hua, DONG Bi-Ying, CAO Hong-Yan, LIU Teng-Yue, YANG Wan-Long, QI Meng, CHEN Ting, WANG Meng-Ying, MENG Dong, YANG Qing, FU Yu-Jie. Identification and Expression Analysis of Key Gene Families in Flavonoid Metabolism Pathway in Pigeon Pea (Cajanus cajan). 农业生物技术学报, 2021, 29(12): 2289-2303.
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