Analysis of Endogenous Hormone Regulation Mechanism for Carotenoid Synthesis in Sarcandra glabra

doi:10.13560/j.cnki.biotech.bull.1985.2023-1047

Abstract

Abstract:

【Objective】This work aims to explore the possible regulatory mechanism of endogenous hormones on the differential accumulation of carotenoids in different organs of Sarcandra glabra. 【Method】In this study, metabolomics was used to analyze the differential distribution of endogenous hormones and carotenoid metabolites in different organs of S. glabra. Combined with transcriptomics technology, the differential enzyme genes related to carotenoids in S. glabra were excavated, and the transcription factors involved in the regulation of differential enzyme genes and their hormone-related cis-response elements were further predicted, so as to analyze the possible regulation of endogenous hormones on the differential accumulation of carotenoids. 【Result】The contents of seven endogenous hormones such as indole-3-carboxylic acid,[（-）-jasmonic acid], ethyl dihydrojasmonate, castasterone and brassinolide were high in the leaves. Consistent with the enrichment sites of 8 differential metabolites, such as canthaxanthin, echinenone, neoxanthin and nostoxanthin; and abscisic acid, gibberellin 4,5-methoxysalicylic acid, dihydrojasmonic acid and 5,6-dihydroxyindole was high in the roots, it was consistent with the accumulation site of 3 differential metabolites, abscisic aldehyde, 4,4'-aiapolycopenedial and antheraxanthin. The key transcription factor MYB106, SPL1, NAC015, ERF064, WRKY44 and BHLH116 responded to the stimulation of plant hormones such as AUX, SA, GA, ABA, and Me_ JA, and participate in the regulation of carotenoid synthesis pathway DXS, VDE, ABA2, AOG, ZEP, CYP97C1, DWARF27, CRTISO, LCYB expression of these 9 metabolic enzymes. Quantitative real-time polymerase chain reaction（RT-qPCR）results showed that the expression trends of 8 randomly selected differentially expressed genes were consistent with the transcriptome sequencing results. 【Conclusion】The 15 differential endogenous hormones and 11 differential metabolites related to carotenoids in the leaves and roots of Sarcandra glabra were screened, and 6 transcription factors were predicted to be involved in the regulation of 16 metabolic enzymes.

Key words: Sarcandra glabra, carotenoids, transcriptome, metabolome

WU Di, YOU Xiao-feng, ZHENG Yi-zheng, LIN Nan, ZHANG Yan-yan, WEI Yi-cong. Analysis of Endogenous Hormone Regulation Mechanism for Carotenoid Synthesis in Sarcandra glabra[J]. Biotechnology Bulletin, 2024, 40(5): 203-214.

Figures/Tables 11

References 33

[1]	秦亚秋, 黄天擎, 刘鄂湖. 草珊瑚化学成分及活性研究概述[J]. 广东化工, 2023, 50(1): 94-95, 91.
	Qin YQ, Huang TQ, Liu EH. Research progress on chemical constituents and activity of sarcandrae herba[J]. Guangdong Chem Ind, 2023, 50(1): 94-95, 91.
[2]	陈芳有, 陈志超, 罗永明. 草珊瑚化学成分及生物活性研究进展[J]. 中国中药杂志, 2022, 47(4): 872-879.
	Chen FY, Chen ZC, Luo YM. Research progress on chemical constituents and biological activities of Sarcandra glabra[J]. China J Chin Mater Med, 2022, 47(4): 872-879.
[3]	韩倩, 武晓林. 肿节风化学成分和药理作用研究进展[J]. 吉林农业, 2017(8): 63-64.
	Han Q, Wu XL. Research progress on chemical constituents and pharmacological effects of Sarcandra glabra[J]. Agric Jilin, 2017(8): 63-64.
[4]	Zeng YL, Liu JY, Zhang Q, et al. The traditional uses, phytochemistry and pharmacology of Sarcandra glabra(thunb.) nakai, a Chinese herb with potential for development: review[J]. Front Pharmacol, 2021, 12: 652926.
[5]	蔡清楼, 陈玲芳. 草珊瑚的应用开发及市场前景探析[J]. 海峡药学, 2010, 22(10): 36-38.
	Cai QL, Chen LF. Application, development and market prospect of grass coral[J]. Strait Pharm J, 2010, 22(10): 36-38.
[6]	陆晨飞, 高月霞, 黄河, 等. 植物类胡萝卜素代谢及调控研究进展[J]. 园艺学报, 2022, 49(12): 2559-2578. doi: 10.16420/j.issn.0513-353x.2021-0531
	Lu CF, Gao YX, Huang H, et al. Carotenoid metabolism and regulation in plants[J]. Acta Hortic Sin, 2022, 49(12): 2559-2578. doi: 10.16420/j.issn.0513-353x.2021-0531
[7]	朱运钦, 乔改梅, 王志强. 植物类胡萝卜素代谢调控的研究进展[J]. 分子植物育种, 2016, 14(2): 471-474.
	Zhu YQ, Qiao GM, Wang ZQ. Research advances on metabolism regulation of plant carotenoids[J]. Mol Plant Breed, 2016, 14(2): 471-474.
[8]	Nisar N, Li L, Lu S, et al. Carotenoid metabolism in plants[J]. Mol Plant, 2015, 8(1): 68-82. doi: 10.1016/j.molp.2014.12.007 pmid: 25578273
[9]	Sathasivam R, Radhakrishnan R, Kim JK, et al. An update on biosynthesis and regulation of carotenoids in plants[J]. S Afr N J Bot, 2021, 140: 290-302.
[10]	何静娟, 范燕萍. 观赏植物花色相关的类胡萝卜素组成及代谢调控研究进展[J]. 园艺学报, 2022, 49(5): 1162-1172. doi: 10.16420/j.issn.0513-353x.2021-0623
	He JJ, Fan YP. Progress in composition and metabolic regulation of carotenoids related to floral color[J]. Acta Hortic Sin, 2022, 49(5): 1162-1172. doi: 10.16420/j.issn.0513-353x.2021-0623
[11]	Lu S, Li L. Carotenoid metabolism: biosynthesis, regulation, and beyond[J]. J Integr Plant Biol, 2008, 50(7): 778-785. doi: 10.1111/j.1744-7909.2008.00708.x
[12]	Sun TH, Yuan H, Cao HB, et al. Carotenoid metabolism in plants: the role of plastids[J]. Mol Plant, 2018, 11(1): 58-74. doi: S1674-2052(17)30273-3 pmid: 28958604
[13]	Hirschberg J. Carotenoid biosynthesis in flowering plants[J]. Curr Opin Plant Biol, 2001, 4(3): 210-218. doi: 10.1016/s1369-5266(00)00163-1 pmid: 11312131
[14]	张睿, 王秀娟, 高伟. 植物激素对次生代谢产物的调控研究[J]. 中国中药杂志, 2020, 45(17): 4205-4210.
	Zhang R, Wang XJ, Gao W. Regulation mechanism of plant hormones on secondary metabolites[J]. China J Chin Mater Med, 2020, 45(17): 4205-4210.
[15]	许智宏, 薛红卫. 植物激素作用的分子机理[M]. 上海: 上海科学技术出版社, 2012.
	Xu ZH, Xue HW. Plant hormones: function and molecular mechanism[M]. Shanghai: Shanghai Scientific & Technical Publishers, 2012.
[16]	高晨曦, 黄艳, 李晶, 等. 茶树新梢内源激素与叶片色泽形成的调控关系研究[J]. 茶叶科学, 2021, 41(6): 802-812.
	Gao CX, Huang Y, Li J, et al. Research on the regulation relationship between endogenous hormones in tea shoots and leaf color formation[J]. J Tea Sci, 2021, 41(6): 802-812.
[17]	马贵芳. 外源水杨酸介导miRNA调控谷子叶酸及类胡萝卜素合成研究[D]. 太原: 山西农业大学, 2022.
	Ma GF. The researched on exogenous salicylic acid-mediated miRNA regulated folate and carotenoid metabolism in foxtail millet[D]. Taiyuan: Shanxi Agricultural University, 2022.
[18]	罗秀芹, 薛晶晶, 蔡杰, 等. GA3处理对木薯类胡萝卜素生物合成的影响[J]. 热带作物学报, 2022, 43(2): 346-352. doi: 10.3969/j.issn.1000-2561.2022.02.015
	Luo XQ, Xue JJ, Cai J, et al. Effects of GA3 treatment on carotenoids biosynthesis in cassava[J]. Chin J Trop Crops, 2022, 43(2): 346-352.
[19]	徐昊, 何伟伟, 李大婧, 等. 外源脱落酸对发芽玉米籽粒中类胡萝卜素合成的影响[J]. 食品科学, 2022, 43(18): 1-8.
	Xu H, He WW, Li DJ, et al. Effect of exogenous abscisic acid on carotenoid synthesis in germinated maize kernels[J]. Food Sci, 2022, 43(18): 1-8.
[20]	Simpson K, Quiroz LF, Rodriguez-Concepción M, et al. Differential contribution of the first two enzymes of the MEP pathway to the supply of metabolic precursors for carotenoid and chlorophyll biosynthesis in carrot(Daucus carota)[J]. Front Plant Sci, 2016,7: 1344.
[21]	Ampomah-Dwamena C, Tomes S, Thrimawithana AH, et al. Overexpression of PSY1 increases fruit skin and flesh carotenoid content and reveals associated transcription factors in apple(Malus × domestica)[J]. Front Plant Sci, 2022, 13: 967143.
[22]	Srinivasan R, Babu S, Gothandam KM. Accumulation of phytoene, a colorless carotenoid by inhibition of phytoene desaturase(PDS)gene in Dunaliella salina V-101[J]. Bioresour Technol, 2017, 242: 311-318.
[23]	Sun TH, Li L. Toward the ‘golden’ era: the status in uncovering the regulatory control of carotenoid accumulation in plants[J]. Plant Sci, 2020, 290: 110331.
[24]	樊宝莲, 王晓云. 转录因子调控植物类胡萝卜素合成途径的研究进展[J]. 分子植物育种, 2021, 19(13): 4401-4408.
	Fan BL, Wang XY. Research progress of transcription factors regulating carotenoid synthesis pathway in plant[J]. Mol Plant Breed, 2021, 19(13): 4401-4408.
[25]	Zhu MK, Chen GP, Zhou S, et al. A new tomato NAC(NAM/ATAF1/2/CUC2)transcription factor, SlNAC4, functions as a positive regulator of fruit ripening and carotenoid accumulation[J]. Plant Cell Physiol, 2014, 55(1): 119-135.
[26]	Zhou D, Shen YH, Zhou P, et al. Papaya CpbHLH1/2 regulate carotenoid biosynthesis-related genes during Papaya fruit ripening[J]. Hortic Res, 2019, 6: 80.
[27]	Zhu KJ, Sun Q, Chen HY, et al. Ethylene activation of carotenoid biosynthesis by a novel transcription factor CsERF061[J]. J Exp Bot, 2021, 72(8): 3137-3154. doi: 10.1093/jxb/erab047 pmid: 33543285
[28]	Ampomah-Dwamena C, Thrimawithana AH, Dejnoprat S, et al. A kiwifruit(Actinidia deliciosa)R2R3-MYB transcription factor modulates chlorophyll and carotenoid accumulation[J]. New Phytol, 2019, 221(1): 309-325. doi: 10.1111/nph.15362 pmid: 30067292
[29]	Zhu LS, Liang SM, Chen LL, et al. Banana MaSPL16 modulates carotenoid biosynthesis during fruit ripening through activating the transcription of lycopene β-cyclase genes[J]. J Agric Food Chem, 2020, 68(5): 1286-1296.
[30]	Han YJ, Wu M, Cao LY, et al. Characterization of OfWRKY3, a transcription factor that positively regulates the carotenoid cleavage dioxygenase gene OfCCD4 in Osmanthus fragrans[J]. Plant Mol Biol, 2016, 91(4-5): 485-496.
[31]	Fu CC, Han YC, Fan ZQ, et al. The Papaya transcription factor CpNAC1 modulates carotenoid biosynthesis through activating phytoene desaturase genes CpPDS2/4 during fruit ripening[J]. J Agric Food Chem, 2016, 64(27): 5454-5463.
[32]	Fu CC, Han YC, Kuang JF, et al. Papaya CpEIN3a and CpNAC2 co-operatively regulate carotenoid biosynthesis-related genes CpPDS2/4, CpLCY-e and CpCHY-b during fruit ripening[J]. Plant Cell Physiol, 2017, 58(12): 2155-2165.
[33]	Moreno JC, Cerda A, Simpson K, et al. Increased Nicotiana tabacum fitness through positive regulation of carotenoid, gibberellin and chlorophyll pathways promoted by Daucus carota lycopene β-cyclase(Dclcyb1)expression[J]. J Exp Bot, 2016, 67(8): 2325-2338. doi: 10.1093/jxb/erw037 pmid: 26893492

基因 Gene	正向引物 Forward primer（5'-3'）	反向引物Reverse primer（5'-3'）
CAC	TCCGACAAATTGGAGGTTGC	TGCTGCTGACAACAATCACG
DXS	GCCTTGAACGGACTCTTAGAC	AGGTGGAGAGCATCCTTGTT
PSY	GGCGTCCTTGTTGTTGTACC	ATGAGTGCTCTAGTGTAGGCTATG
NCED	CTCCAAGGCGTGTTCTATCG	CTCCAAGGCGTGTTCTATCG
VDE	CACGACAACGAGTACCTTCAC	TGACAGCACCACCATATCCA
PDS	ACGAGTACCAGGTGTTGTTCTA	AATCAGTTTGCCAGCATTCATTG
LYCB	GGAAGATATACAGGAGAGGATGGT	TCTGAGGAAGCACTGGAAGT
ZEP	GCTTGGAGGCAGAGTATTGAC	GCAGCCATTCTAGCCATTCC
CYP97C1	GATGAGAGCACAACAAGAGGTT	CGCCTTATTAACACTGGTGGAT

基因 Gene	正向引物 Forward primer（5'-3'）	反向引物Reverse primer（5'-3'）
CAC	TCCGACAAATTGGAGGTTGC	TGCTGCTGACAACAATCACG
DXS	GCCTTGAACGGACTCTTAGAC	AGGTGGAGAGCATCCTTGTT
PSY	GGCGTCCTTGTTGTTGTACC	ATGAGTGCTCTAGTGTAGGCTATG
NCED	CTCCAAGGCGTGTTCTATCG	CTCCAAGGCGTGTTCTATCG
VDE	CACGACAACGAGTACCTTCAC	TGACAGCACCACCATATCCA
PDS	ACGAGTACCAGGTGTTGTTCTA	AATCAGTTTGCCAGCATTCATTG
LYCB	GGAAGATATACAGGAGAGGATGGT	TCTGAGGAAGCACTGGAAGT
ZEP	GCTTGGAGGCAGAGTATTGAC	GCAGCCATTCTAGCCATTCC
CYP97C1	GATGAGAGCACAACAAGAGGTT	CGCCTTATTAACACTGGTGGAT

化合物 Compound	分子式 Formula	离子模式 ESM	质荷比 m/z	保留时间 Retention/min	二级鉴定分数 MS2 score
吲哚-3-羧酸 Indole-3-carboxylic acid	C₉H₇NO₂	-	160.04	1.84	43.60
吲哚 Indole	C₈H₇N	-	152.01	1.88	63.40
吲哚-5-羧酸 Indole-5-carboxylic acid	C₉H₇NO₂	-	160.04	2.05	33.60
3-甲基羟基吲哚 3-Methyldioxyindole	C₉H₉NO₂	-	162.06	2.27	38.70
5,6-二羟基吲哚 5,6-Dihydroxyindole	C₈H₇NO₂	-	297.09	2.67	27.00
吲哚-3-甲醛 Indole-3-carboxaldehyde	C₉H₇N_O	+	110.04	3.88	0.00
5,6-吲哚醌-2-羧酸 5,6-Indolequinone-2-carboxylic acid	C₉H₅NO₄	+	383.05	1.81	22.00
1-萘乙酸乙酯 Ethyl 1-naphthylacetic acid	C₁₄H₁₄O₂	+	215.11	3.60	34.30
赤霉素3 Gibberellin A3	C₁₉H₂₂O₆	-	391.15	3.16	74.30
赤霉素4 Gibberellin A4	C₁₉H₂₄O₅	-	663.29	6.78	78.10
赤霉素36 Gibberellin A36	C₂₀H₂₆O₆	+	345.17	7.76	66.80
邻β-d-葡糖基玉米素 O-Beta-D-glucosylzeatin	C₁₆H₂₃N₅O₆	-	402.13	3.65	74.30
油菜素内酯 Brassinolide	C₂₈H₄₈O₆	+	445.34	10.88	53.40
二氢茉莉酸 Dihydrojasmonic acid	C₁₂H₂₀O₃	-	233.12	4.42	36.50
（-）-茉莉酸（-）-Jasmonic acid	C₁₂H₁₈O₃	+	175.11	4.99	66.60
茉莉酸 Jasmonic acid	C₁₂H₁₈O₃	+	175.11	7.34	69.00
5-甲氧基水杨酸 5-Methoxysalicylic acid	C₈H₈O₄	-	167.04	1.35	86.90
赤霉素20 Gibberellin A20	C₁₉H₂₄O₅	-	353.15	7.05	47.80
赤霉素8 Gibberellin A8	C₁₉H₂₄O₇	+	387.13	4.17	66.10
赤霉素19 Gibberellin A19	C₂₀H₂₆O₆	+	385.17	6.84	70.20
赤霉素9 Gibberellin A9	C₁₉H₂₄O₄	+	317.18	8.95	42.20
（+）-脱落酸（+）-Abscisic acid	C₁₅H₂₀O₄	+	247.13	5.70	81.20
油菜素甾酮 Castasterone	C₂₈H₄₈O₅	+	447.34	10.77	60.70
褪黑素 Melatonin	C₁₃H₁₆N₂O₂	-	277.12	2.80	29.50
对水杨酸 P-salicylic acid	C₇H₆O₃	-	137.03	1.56	28.90
茉莉酸甲酯 Methyl jasmonate	C₁₃H₂₀O₃	+	207.14	3.00	45.30
二氢茉莉酮酸甲酯 Methyl dihydrojasmonate	C₁₃H₂₂O₃	-	225.15	4.17	25.80
2-萘乙酸 2-Naphthylacetic acid	C₁₂H₁₀O₂	+	187.08	8.05	33.90
玉米素 Zeatin	C₁₀H₁₃N₅O	+	220.12	1.57	26.50
玉米素核苷 cis-Zeatin riboside	C₁₅H₂₁N₅O₅	+	374.15	3.43	78.90
3-吲哚丙烯酸 3-Indoleacrylic acid	C₁₁H₉NO₂	-	208.03	3.07	43.60

化合物 Compound	分子式 Formula	离子模式 ESM	质荷比 m/z	保留时间 Retention/min	二级鉴定分数 MS2 score
吲哚-3-羧酸 Indole-3-carboxylic acid	C₉H₇NO₂	-	160.04	1.84	43.60
吲哚 Indole	C₈H₇N	-	152.01	1.88	63.40
吲哚-5-羧酸 Indole-5-carboxylic acid	C₉H₇NO₂	-	160.04	2.05	33.60
3-甲基羟基吲哚 3-Methyldioxyindole	C₉H₉NO₂	-	162.06	2.27	38.70
5,6-二羟基吲哚 5,6-Dihydroxyindole	C₈H₇NO₂	-	297.09	2.67	27.00
吲哚-3-甲醛 Indole-3-carboxaldehyde	C₉H₇N_O	+	110.04	3.88	0.00
5,6-吲哚醌-2-羧酸 5,6-Indolequinone-2-carboxylic acid	C₉H₅NO₄	+	383.05	1.81	22.00
1-萘乙酸乙酯 Ethyl 1-naphthylacetic acid	C₁₄H₁₄O₂	+	215.11	3.60	34.30
赤霉素3 Gibberellin A3	C₁₉H₂₂O₆	-	391.15	3.16	74.30
赤霉素4 Gibberellin A4	C₁₉H₂₄O₅	-	663.29	6.78	78.10
赤霉素36 Gibberellin A36	C₂₀H₂₆O₆	+	345.17	7.76	66.80
邻β-d-葡糖基玉米素 O-Beta-D-glucosylzeatin	C₁₆H₂₃N₅O₆	-	402.13	3.65	74.30
油菜素内酯 Brassinolide	C₂₈H₄₈O₆	+	445.34	10.88	53.40
二氢茉莉酸 Dihydrojasmonic acid	C₁₂H₂₀O₃	-	233.12	4.42	36.50
（-）-茉莉酸（-）-Jasmonic acid	C₁₂H₁₈O₃	+	175.11	4.99	66.60
茉莉酸 Jasmonic acid	C₁₂H₁₈O₃	+	175.11	7.34	69.00
5-甲氧基水杨酸 5-Methoxysalicylic acid	C₈H₈O₄	-	167.04	1.35	86.90
赤霉素20 Gibberellin A20	C₁₉H₂₄O₅	-	353.15	7.05	47.80
赤霉素8 Gibberellin A8	C₁₉H₂₄O₇	+	387.13	4.17	66.10
赤霉素19 Gibberellin A19	C₂₀H₂₆O₆	+	385.17	6.84	70.20
赤霉素9 Gibberellin A9	C₁₉H₂₄O₄	+	317.18	8.95	42.20
（+）-脱落酸（+）-Abscisic acid	C₁₅H₂₀O₄	+	247.13	5.70	81.20
油菜素甾酮 Castasterone	C₂₈H₄₈O₅	+	447.34	10.77	60.70
褪黑素 Melatonin	C₁₃H₁₆N₂O₂	-	277.12	2.80	29.50
对水杨酸 P-salicylic acid	C₇H₆O₃	-	137.03	1.56	28.90
茉莉酸甲酯 Methyl jasmonate	C₁₃H₂₀O₃	+	207.14	3.00	45.30
二氢茉莉酮酸甲酯 Methyl dihydrojasmonate	C₁₃H₂₂O₃	-	225.15	4.17	25.80
2-萘乙酸 2-Naphthylacetic acid	C₁₂H₁₀O₂	+	187.08	8.05	33.90
玉米素 Zeatin	C₁₀H₁₃N₅O	+	220.12	1.57	26.50
玉米素核苷 cis-Zeatin riboside	C₁₅H₂₁N₅O₅	+	374.15	3.43	78.90
3-吲哚丙烯酸 3-Indoleacrylic acid	C₁₁H₉NO₂	-	208.03	3.07	43.60

代谢酶Enzyme name	缩写Abbreviation	KO 编码KO ID	unigene数量Number of unigene
1-脱氧-D-木酮糖5-磷酸合酶1-Deoxy-D-xylulose 5-phosphate synthase	DXS	K01662	3
15-cis-phytoene合成酶15-cis-Phytoene synthase	PSY	K02291	1
15 - cis - Phytoene去饱和酶15-cis-Phytoene desaturase	PDS	K02293	1
前番茄红素异构酶Prolycopene isomerase	CRTISO	K09835	1
番茄红素β-环化酶Lycopene beta-cyclase	LCYB	K06443	1
番茄红素环化酶Lycopene epsilon-cyclase	LCYE	K06444	1
胡萝卜素羟基Beta-ring hydroxylase	CHYB	K15747	2
类胡萝卜素ε-羟化酶Carotenoid epsilon hydroxylase	CYP97C1	K09837	1
β -胡萝卜素异构酶Beta-carotene isomerase	DWARF27	K17911	3
内酯合成酶Carlactone synthase	CCD8	K17913	1
玉米黄质环氧化酶Zeaxanthin epoxidase	ZEP	K09838	3
紫黄质脱环氧化酶Violaxanthin de-epoxidase	VDE	K09839	1
9-顺式-环氧类胡萝卜素二加氧酶9-cis-Epoxycarotenoid dioxygenase	NCED	K09840	2
黄氧素脱氢酶Xanthoxin dehydrogenase	ABA2	K09841	3
（+）- 脱落酸8’羟化酶（+）-Abscisic acid 8’-hydroxylase	CYP707A	K09843	1
脱落酸β-葡萄糖基转移酶Abscisate beta-glucosyltransferase	AOG	K14595	1