转录组与代谢组联合分析揭示大豆GmERD15c参与盐胁迫下类黄酮的生物合成

doi:10.13560/j.cnki.biotech.bull.1985.2024-0373

摘要/Abstract

摘要：

【目的】大豆GmERD15c是ERD15转录因子家族成员之一，探究盐胁迫下转GmERD15c基因大豆株系的基因表达及物质代谢情况，揭示其与大豆耐盐性的关系。【方法】以转GmERD15c基因大豆株系及其受体大豆‘东农50’为材料，经盐胁迫处理后，采取根部进行转录组学和代谢组学分析。【结果】盐胁迫前后差异代谢物在类黄酮合成通路变化较大，胁迫后的代谢产物二氢山奈酚、儿茶素和槲皮素含量发生显著变化，且槲皮素含量在转GmERD15c基因株系中更高。同时，转录组学与代谢组学联合分析发现，GmERD15c基因与类黄酮生物合成相关酶关系密切。【结论】盐胁迫下，大豆GmERD15c基因可能通过调控类黄酮合成相关的酶类进而影响类黄酮的生物合成，而类黄酮可以减轻盐分引起的氧化应激，为后续揭示GmERD15c基因的耐盐机制提供了新线索。

关键词: 大豆, GmERD15, 类黄酮, 转录组学, 代谢组学

Abstract:

【Objective】 Soybean GmERD15c is one of the members of the ERD15 transcription factor family, and the gene expression and substance metabolism of soybean lines transgenic with GmERD15c gene under salt stress were explored, and the relationship between GmERD15c and soybean's tolerance to salt was revealed.【Method】 Using the transgenic soybean line and its recipient soybean ‘Dongnong 50'as the materials, and the roots after the salt stress treatment were to have analysis of transcriptomics and metabolomics.【Result】 There were significant changes in the metabolites of flavonoid synthesis pathway before and after salt stress, with significant variations in the content of dihydroquercetin, catechin, and fisetin after stress. And the content of quercetin was higher in the transgenic line of GmERD15c. Meanwhile, the combined analysis of transcriptomics and metabolomics revealed that the GmERD15c gene was closely related to the flavonoid biosynthesis-related enzymes.【Conclusion】 It is hypothesized that the soybean GmERD15c gene may affect the biosynthesis of flavonoids by regulating the enzymes related to flavonoid synthesis under salt stress, and flavonoids may reduce the oxidative stress caused by salt, which provides new clues for subsequently revealing the salt tolerance mechanism of the GmERD15c gene.

Key words: soybean, GmERD15, flavonoids, transcriptomics, metabolomics

韩乐乐, 宋文迪, 边嘉珅, 李阳, 杨双胜, 陈紫怡, 李晓薇. 转录组与代谢组联合分析揭示大豆GmERD15c参与盐胁迫下类黄酮的生物合成[J]. 生物技术通报, 2024, 40(10): 243-252.

HAN Le-le, SONG Wen-di, BIAN Jia-shen, LI Yang, YANG Shuang-sheng, CHEN Zi-yi, LI Xiao-wei. Revealing the Flavonoid Biosynthesis of Soybean GmERD15c under Salt Stress by Combined Analysis of Transcriptome and Metabolome[J]. Biotechnology Bulletin, 2024, 40(10): 243-252.

图/表 8

图1 GmERD15c在大豆不同组织中的表达情况

Fig. 1 Expressions of GmERD15c in the different tissues of soybean

图2 盐胁迫下GmERD15c在大豆根和叶中的表达情况

Fig. 2 Expressions of GmERD15c in the root and leaf of soybean under salt stress

表1 差异代谢物

Table 1 Differential metabolites

代谢物级别 Metabolite level	组别 Group	代谢物数量 Number of metabolites	上调数量 Up-regulated number	下调数量 Down-regulated number	总差异代谢物 Total differential metabolites
一级差异代谢物 Primary differential metabolites	WT-mock VS OE-mock	20 804	846	847	1 693
一级差异代谢物 Primary differential metabolites	WT-NaCl VS OE-NaCl	20 804	840	735	1 575
二级差异代谢物 Secondary differential metabolites	WT-mock VS OE-mock	402	20	15	35
二级差异代谢物 Secondary differential metabolites	WT-NaCl VS OE-NaCl	402	31	21	52

图3 转GmERD15c基因大豆类黄酮合成通路的代谢物变化 A：二级差异代谢物主要分类；B：转基因大豆代谢物KEGG分析；C：转基因大豆类黄酮化合物定量分析

Fig. 3 Metabolite variation in flavonoid synthesis pathway of soybean with transgenic GmERD15c gene A: Main classification of secondary differential metabolites. B: KEGG analysis of transgenic soybean metabolites. C: Quantitative analysis of transgenic soybean flavonoid compounds

表2 大豆转录组差异基因分析

Table 2 Transcriptome differential gene analysis in soybean

对照组Control	实验组Treat	上调表达基因Up-regulated genes	下调表达基因Down-regulated genes	差异表达基因Total DEGs
WT-mock	OE-mock	605	1 208	1 813
WT-NaCl	OE-NaCl	681	693	1 374
OE-mock	OE-NaCl	5 348	4 456	9 804
WT-mock	WT-NaCl	6 576	6 533	13 109

图4 大豆转录组差异基因Upset图 Number in each set表示每个比较组鉴定到的全部差异基因的数目。Number of each intersect表示多个比较组鉴定到的共有差异基因的数目，横坐标一个点表示该比较组鉴定到的特有差异基因的数目，横坐标多个点连线表示连线的多个比较组鉴定到的共有差异基因的数目。蓝色为1个组别，黄色为2个组别，红色为3个组别，绿色为4个组别

Fig. 4 Upset diagram of transcriptome differential genes in soybean Number in each set indicates the number of all differential genes identified in each comparison group. Number of each intersect indicates the number of common differences identified by multiple comparison groups, a point on the abscissa indicates the number of unique differences identified by the comparison group, and a line connecting multiple points on the abscissa indicates the number of common differences identified by multiple comparison groups connected by the line. Blue indicates one group, yellow indicates two groups, red indicates three groups, and green indicates four groups

表3 盐胁迫下转GmERD15c基因大豆转录组KEGG分析

Table 3 KEGG analysis of soybean transcriptome transgenic GmERD15c gene under salt stress

通路ID Pathway ID	KEGG通路 KEGG pathway	1级通路 Level 1	2级通路 Level 2	上调数量 Up_number	下调数量 Down_number	差异数量 DEG_number	总数量 Total_number
gmx00910	氮代谢	新陈代谢	能量代谢	6	2	8	44
gmx00260	甘氨酸、丝氨酸和苏氨酸代谢	新陈代谢	氨基酸代谢	4	4	8	89
gmx00940	苯丙烷类生物合成	新陈代谢	其他次生代谢产物的生物合成	8	7	15	260
gmx00565	乙醚脂质代谢	新陈代谢	脂类代谢	5	0	5	43
gmx00904	二萜类生物合成	新陈代谢	萜类化合物和聚酮类化合物的代谢	1	4	5	45
gmx00564	甘油磷脂代谢	新陈代谢	脂类代谢	9	1	10	164
gmx00592	亚麻酸代谢	新陈代谢	脂类代谢	4	2	6	72
gmx00908	玉米素生物合成	新陈代谢	萜类化合物和聚酮类化合物的代谢	3	1	4	36
gmx00350	酪氨酸代谢	新陈代谢	氨基酸代谢	3	2	5	56
gmx00270	半胱氨酸和蛋氨酸代谢	新陈代谢	氨基酸代谢	6	3	9	154
gmx00630	乙醛酸盐和二羧酸代谢	新陈代谢	碳水化合物代谢	0	6	6	87
gmx00130	泛醌和其他萜类醌生物合成	新陈代谢	辅因子和维生素的代谢	4	1	5	68
gmx00943	异黄酮生物合成	新陈代谢	其他次生代谢产物的生物合成	3	0	3	28
gmx00941	类黄酮生物合成	新陈代谢	其他次生代谢产物的生物合成	3	2	5	77

图5 转录代谢联合KEGG分析底色表示mRNA，矩形表示代谢物。蓝色表示代谢物下调，红色表示代谢物上调，绿色表示mRNA下调，黄色mRNA上调，粉色表示多差异mRNA。括号中的编号表示酶的反应类型，R07432：奎宁酸羟基肉桂酰转移酶，R01945：奎宁酸酯O-（3,4-二羟基肉桂酰基）转移酶，R01613：4-香豆酰辅酶A丙二酰转移酶 R07988：咖啡酰辅酶A丙二酰转移酶 R02444：氧化还原酶

Fig. 5 Transcriptional metabolism combined with KEGG analysis The base color indicates mRNA and the rectangle indicates metabolites. Blue indicates metabolite downregulation, red indicates metabolite upregulation, green indicates mRNA downregulation, yellow indicates mRNA upregulation, and pink indicates polydifferential mRNA.The numbers in the parentheses indicate the types of enzyme reactions. R07432: Quinate hydroxycinnamoyltransferase.R01945: Quinate ester O-（3,4-dihydroxycinnamoyl）transferase. R01613: 4-coumaroyl-CoA:malate acyltransferase. R07988: Caffeoyl-CoA:malate acyltransferase. R02444: Oxidoreductase

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