Identification of 4CL Gene Family in Glycyrrhiza uralensis and Its Expression under Drought Stress

doi:10.7668/hbnxb.20194561

Abstract

Abstract:

4-coumaric acid:coenzyme A ligase(4CL),as the main biosynthetic enzyme of flavonoids and lignin,is closely related to the formation of plant flavonoids.In order to explore the relationship between the 4CL gene family of Glycyrrhiza uralensis and the synthesis and accumulation of isoglycyrrhizin,this study determined the content of isoglycyrrhizin in G. uralensis after drought stress,and the expression characteristics and bioinformatics analysis of the 4CL gene family of G. uralensis Fisch.to understand the relationship between the accumulation characteristics of isoglycyrrhizin and the 4CL gene of G.uralensis.The results showed that isoliquiritigenin was mainly accumulated in the roots of G.uralensis,and drought stress could significantly increase the content of isoliquiritigenin in the roots of G.uralensis.The content of isoliquiritigenin after 2 h of polyethylene glycol(PEG)stress was 3.91 times that of the control group(0 h).Drought stress could induce the up regulation of Gu4CL gene in the underground part.The expression levels of Gu4CL2, Gu4CL4 and Gu4CL5 were higher after PEG stress induction,while Gu4CL2 was most significantly up-regulated after PEG stress.The expression of Gu4CL2 was similar to that of isoliquiritigenin.Bioinformatics analysis of Gu4CL gene showed that 11 genes had two conserved polypeptide motifs Box Ⅰ(SSGTTGLPKGV)and Box Ⅱ(GEICIRG),and 11 genes were distributed on 11 Scaffold fragments.Promoter cis-acting element analysis showed that Gu4CL2, Gu4CL4 and Gu4CL5 genes contained more abscisic acid and jasmonic acid response elements than other Gu4CL genes.Therefore,drought stress may induce the synthesis of jasmonic acid and abscisic acid,regulate the expression of Gu4CL2, Gu4CL4 and Gu4CL5 genes in licorice roots,and increase the accumulation of isoliquiritigenin.This study provides a research basis for optimizing the cultivation techniques of G.uralensis,improving the adaptability and quality of cultivated land in saline-alkali land,exploring the role of Gu4CL gene family in G.uralensis,and exploring the synthesis mechanism of isoliquiritigenin.

Key words: Liquorice, Gu4CL, Drought stress, Isoliquiritigenin, Bioinformatics analysis

ZHAO Bin, YAO Hua, SHI Nana, GAO Zhuanzhuan, YANG Mao, FENG Jianghua, SHEN Haitao. Identification of 4CL Gene Family in Glycyrrhiza uralensis and Its Expression under Drought Stress[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(S1): 1-10. doi: 10.7668/hbnxb.20194561.

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Figures/Tables 11

References 28

[17]	Tong X C, Cao A P, Wang F, Chen X F, Xie S Q, Shen H T, Jin X, Li H B. Calcium-dependent protein kinase genes in Glycyrrhiza uralensis appear to be involved in promoting the biosynthesis of glycyrrhizic acid and flavonoids under salt stress[J]. Molecules, 2019, 24(9):1837.doi:10.3390/molecules24091837.
[18]	桑雪雨. 适度干旱胁迫调控甘草次级代谢分子机制的初步研究[D]. 广州: 广东药学院, 2014.
	Sang X Y. A preliminary study on the molecular mechanism of moderate drought stress regulation on the secondary metabolism in Glycyrrhiza uralensis[D]. Guangzhou: Guangdong Pharmaceutical University, 2014.
[19]	何梦媛, 姚华, 李国治, 杨茂, 沈海涛. 甘草CHS基因家族鉴定、表达特性分析及其与甘草查尔酮A积累的关系研究[J]. 植物生理学报, 2022, 58(1):141-154.doi:10.13592/j.cnki.ppj.2021.0349.
	He M Y, Yao H, Li G Z, Yang M, Shen H T. Identification of CHS gene family and analysis of its expression characteristics in relation to the accumulation of licochalcone A in Chinese licorice(Glycyrrhiza uralensis)[J]. Plant Physiology Journal, 2022, 58(1):141-154.
[20]	Stuible H P, Kombrink E. Identification of the substrate specificity-conferring amino acid residues of 4-coumarate:coenzyme A ligase allows the rational design of mutant enzymes with new catalytic properties[J]. Journal of Biological Chemistry, 2001, 276(29):26893-26897.doi:10.1074/jbc.M100355200. pmid: 11323416
[21]	Li Z, Nair S K. Structural basis for specificity and flexibility in a plant 4-coumarate:CoA ligase[J]. Structure, 2015, 23(11):2032-2042.doi:10.1016/j.str.2015.08.012. pmid: 26412334
[22]	杨婷, 潘翔, 饶国栋, 陆海. 植物4CL基因家族结构功能与表达特性研究进展[J]. 成都大学学报(自然科学版), 2011, 30(1):4-7.doi:10.3969/j.issn.1004-5422.2011.01.002.
	Yang T, Pan X, Rao G D, Lu H. Research progress in structural function and expression characteristic of 4CL's gene family in plants[J]. Journal of Chengdu University(Natural Science Edition), 2011, 30(1):4-7.
[1]	屈谦伟. 以IGPD为靶标探究异甘草素对木糖葡萄球菌抗菌作用的研究[D]. 哈尔滨: 东北农业大学, 2020.doi:10.27010/d.cnki.gdbnu.2020.000142.
	Qu Q W. Study on the antibacterial effect of isoliquiritigenin against Staphylococcus xylosus with IGPD as the target[D]. Harbin: Northeast Agricultural University, 2020.
[2]	李媛媛, 潘新锋, 池丽俏. 异甘草素通过JAK/STAT途径治疗小鼠特应性皮炎的机制研究[J]. 河北医学, 2021, 27(5):710-715.doi:10.3969/j.issn.1006-6233.2021.05.02.
	Li Y Y, Pan X F, Chi L Q. Mechanism of isoliquiritigenin in the treatment of atopic dermatitis in mice through JAK-STAT pathway[J]. Hebei Medicine, 2021, 27(5):710-715.
[3]	邢晓旭, 张宗山, 赵莹莹, 张馨慧, 曾丹丹, 赵玉珠, 张旭. 异甘草素抗腹泻活性及其分子机制研究[J]. 东北农业大学学报, 2021, 52(11):26-33.doi:10.19720/j.cnki.issn.1005-9369.2021.11.004.
	Xing X X, Zhang Z S, Zhao Y Y, Zhang X H, Zeng D D, Zhao Y Z, Zhang X. Study on the anti-diarrhea activity of isoliquiritigenin and its molecular mechanism[J]. Journal of Northeast Agricultural University, 2021, 52(11):26-33.
[4]	祁建宏, 董芳旭. 黄酮类化合物药理作用研究进展[J]. 北京联合大学学报, 2020, 34(3):89-92. doi:10.16255/j.cnki.ldxbz.2020.03.014.
	Qi J H, Dong F X. Research progress on pharmacological effects of flavonoids[J]. Journal of Beijing Union University, 2020, 34(3):89-92. doi: 10.16255/j.cnki.ldxbz.2020.03.014.
[5]	罗睿雄, 赵志常, 高爱平, 黄建峰, 刘宽亮. 芒果4CL基因的克隆及其表达分析[J]. 华北农学报, 2016, 31(S1):57-62.doi:10.7668/hbnxb.2016.S1.010.
	Luo R X, Zhao Z C, Gao A P, Huang J F, Liu K L. Cloning and expression analysis of 4CL gene from mango[J]. Acta Agriculturae Boreali-Sinica, 2016, 31(S1):57-62.
[6]	Shi R, Sun Y H, Li Q Z, Heber S, Sederoff R, Chiang V L. Towards a systems approach for lignin biosynthesis in Populus trichocarpa:transcript abundance and specificity of the monolignol biosynthetic genes[J]. Plant & Cell Physiology, 2010, 51(1):144-163.doi:10.1093/pcp/pcp175.
[7]	Costa M A, Collins R E, Anterola A M, Cochrane F C, Davin L B, Lewis N G. An in silico assessment of gene function and organization of the phenylpropanoid pathway metabolic networks in Arabidopsis thaliana and limitations thereof[J]. Phytochemistry, 2003, 64(6):1097-1112.doi:10.1016/s0031-9422(03)00517-x.
[8]	曹运鹏, 方志, 李姝妹, 闫冲冲, 丁庆庆, 程曦, 林毅, 郭宁, 蔡永萍. 砀山酥梨4CL基因家族的全基因组鉴定与分析[J]. 遗传, 2015, 37(7):711-719.doi:10.16288/j.yczz.15-069.
	Cao Y P, Fang Z, Li S M, Yan C C, Ding Q Q, Cheng X, Lin Y, Guo N, Cai Y P. Genome-wide identification and analyses of 4CL gene families in Pyrus bretschneideri Rehd[J]. Hereditas, 2015, 37(7):711-719.
[9]	Wang B, Sun W, Li Q S, Li Y, Luo H M, Song J Y, Sun C, Qian J, Zhu Y J, Hayward A, Xu H B, Chen S L. Genome-wide identification of phenolic acid biosynthetic genes in Salvia miltiorrhiza[J]. Planta, 2015, 241(3):711-725.doi:10.1007/s00425-014-2212-1. pmid: 25471478
[10]	Lindermayr C, Möllers B, Fliegmann J, Uhlmann A, Lottspeich F, Meimberg H, Ebel J. Divergent members of a soybean(Glycine max L.) 4-coumarate:coenzyme A ligase gene family[J]. European Journal of Biochemistry, 2002, 269(4):1304—1315.doi:10.1046/j.1432-1033.2002.02775.x. pmid: 11856365
[11]	Kumar A, Ellis B E. The phenylalanine ammonia-lyase gene family in raspberry.structure,expression,and evolution[J]. Plant Physiology, 2001, 127(1):230-239.doi:10.1104/pp.127.1.230. pmid: 11553751
[12]	周洵. 菘蓝中4-香豆酰辅酶A连接酶家族的功能研究[D]. 上海: 第二军医大学, 2013.doi:10.7666/d.Y2340075.
	Zhou X. Identification and characterization of a 4-coumarate:CoA ligase gene family in Isaits indigotica Fort.[D]. Shanghai: Second Military Medical University, 2013.
[13]	孙士超. 绿色棉纤维转录组分析及4CL基因家族的鉴定[D]. 石河子: 石河子大学, 2020.doi:10.27332/d.cnki.gshzu.2020.000010.
	Sun S C. Transcriptome analysis of green cotton fiber and identification of 4CL gene family[D]. Shihezi: Shihezi University, 2020.
[14]	林建中. 拟南芥4CL3基因在类黄酮合成代谢中的功能分析[D]. 长沙: 湖南大学, 2009.doi:10.7666/d.y1725482.
	Lin J Z. Functional analysis of Arabidopsis 4CL3 gene in flavonoid biosynthesis[D]. Changsha: Hunan University, 2009.
[15]	Sun H Y, Li Y, Feng S Q, Zou W H, Guo K, Fan C F, Si S L, Peng L C. Analysis of five rice 4-coumarate:coenzyme A ligase enzyme activity and stress response for potential roles in lignin and flavonoid biosynthesis in rice[J]. Biochemical and Biophysical Research Communications, 2013, 430(3):1151-1156.doi:10.1016/j.bbrc.2012.12.019. pmid: 23246835
[16]	Sun Y J, Qiao L P, Shen Y, Jiang P, Chen J C, Ye X Q. Phytochemical profile and antioxidant activity of physiological drop of Citrus fruits[J]. Journal of Food Science, 2013, 78(1):C37—C42.doi:10.1111/j.1750-3841.2012.03002.x.
[23]	Sun S C, Xiong X P, Zhang X L, Feng H J, Zhu Q H, Sun J, Li Y J. Characterization of the Gh4CL gene family reveals a role of Gh4CL7 in drought tolerance[J]. BMC Plant Biology, 2020, 20(1):125.doi:10.1186/s12870-020-2329-2.
[24]	Zhao P, Wang D D, Wang R Q, Kong N N, Zhang C, Yang C H, Wu W T, Ma H L, Chen Q. Genome-wide analysis of the potato Hsp20 gene family:identification,genomic organization and expression profiles in response to heat stress[J]. BMC Genomics, 2018, 19(1):61.doi:10.1186/s12864-018-4443-1.
[25]	Zhang C H, Ma T, Luo W C, Xu J M, Liu J Q, Wan D S. Identification of 4CL genes in desert poplars and their changes in expression in response to salt stress[J]. Genes, 2015, 6(3):901-917.doi:10.3390/genes6030901.
[26]	Gao S, Yu H N, Xu R X, Cheng A X, Lou H X. Cloning and functional characterization of a 4-coumarate CoA ligase from liverwort Plagiochasma appendiculatum[J]. Phytochemistry, 2015, 111:48-58.doi:10.1016/j.phytochem.2014.12.017.
[27]	Godoy M, Franco-Zorrilla J M, Pérez-Pérez J, Oliveros J C, Lorenzo O, Solano R. Improved protein-binding microarrays for the identification of DNA-binding specificities of transcription factors[J]. The Plant Journal, 2011, 66(4):700—711.doi:10.1111/j.1365-313X.2011.04519.x. pmid: 21284757
[28]	孙凯文. 植物激素脱落酸对茉莉酸介导的防御反应的调控[D]. 南京: 南京师范大学, 2019.doi:10.27245/d.cnki.gnjsu.2019.001050.
	Sun K W. Regulation of plant hormone abscisic acid on jasmonic acid-mediated defense response[D]. Nanjing: Nanjing Normal University, 2019.

基因ID Gene ID	上游引物序列(5'-3') Upstream primer sequence(5'-3')	下游引物序列(5'-3') Downstream primer sequence(5'-3')
Glyur000051s00003417.1	AATTCGTCTTCACCTTCCTC	TGTGTGATGATGAGTTTCGT
Glyur001041s00027454.1	TTAGATGCTGCTGTCATACC	ACTCCAGGTACCAACTATCA
Glyur001000s00030117.1	CAATGTGGAGGCTAATACGA	GCAAGTACCCATCAGAATCA
Glyur000100s00008376	CTGATGCAGAGCTTCAAATCGAG	TTCGGAAGGAAGGTTGAGGTAAG

基因ID Gene ID	上游引物序列(5'-3') Upstream primer sequence(5'-3')	下游引物序列(5'-3') Downstream primer sequence(5'-3')
Glyur000051s00003417.1	AATTCGTCTTCACCTTCCTC	TGTGTGATGATGAGTTTCGT
Glyur001041s00027454.1	TTAGATGCTGCTGTCATACC	ACTCCAGGTACCAACTATCA
Glyur001000s00030117.1	CAATGTGGAGGCTAATACGA	GCAAGTACCCATCAGAATCA
Glyur000100s00008376	CTGATGCAGAGCTTCAAATCGAG	TTCGGAAGGAAGGTTGAGGTAAG

基因 Gene	地上部分表达量Aboveground part expression level					地下部分表达量Underground part expression level
基因 Gene	0 h	2 h	6 h	12 h	24 h	0 h	2 h	6 h	12 h	24 h
Gu4CL1	9.26±3.08b	8.59±2.55b	4.66±0.92b	13.72±4.02b	43.79±23.22a	0.82±0.74ab	0.85±0.32ab	0.52±0.38b	0.85±0.12ab	1.61±0.69a
Gu4CL2	36.58±13.22ab	46.83±12.04ab	23.09±13.52b	36.93±11.23ab	63.23±30.27a	6.41±4.38c	56.17±14.06a	19.89±8.03bc	34.03±3.09b	33.60±15.57b
Gu4CL3	36.64±10.85ab	30.14±8.68ab	41.21±3.50a	26.91±7.34bc	15.32±2.43c	21.27±5.66a	22.13±9.76a	20.57±3.15a	17.88±1.53ab	9.18±3.34b
Gu4CL4	6.87±3.91a	10.57±2.06a	5.32±2.31a	9.03±2.71a	8.79±7.11a	36.95±8.83b	124.52±66.94a	62.49±14.25ab	59.81±16.80ab	55.44±37.15ab
Gu4CL5	14.77±5.17c	38.94±14.93a	15.80±4.80c	5.61±1.88c	48.45±23.02a	2.53±1.15c	17.99±6.00a	4.93±3.35b	4.98±2.13b	2.42±1.24b
Gu4CL6	1.66±0.31b	2.63±0.90ab	2.19±0.81b	4.60±1.04a	3.71±2.19ab	5.15±0.17a	14.88±8.00a	9.93±2.72a	13.23±6.07a	6.76±5.10a
Gu4CL7	0.04±0.05a	0.00±0.00a	0.01±0.02a	0.04±0.04a	0.36±0.62a	0.93±0.29a	0.32±0.32a	0.52±0.32a	0.45±0.32a	0.52±0.37a
Gu4CL8	3.30±2.70a	3.47±1.69a	2.78±0.99a	7.81±4.93a	4.03±2.33a	0.38±0.32a	0.36±0.09a	0.34±0.18a	0.49±0.25a	0.86±0.55a
Gu4CL9	0.46±0.38ab	0.27±0.08b	0.35±0.22b	1.04±0.47a	0.80±0.30ab	0.40±0.40a	0.06±0.05a	0.19±0.13a	0.28±0.21a	0.20±0.22a
Gu4CL10	1.21±0.58ab	0.32±0.07b	1.40±1.52ab	5.35±5.30a	0.49±0.50b	0.25±0.22a	0.00±0.00b	0.00±0.00b	0.00±0.00b	0.00±0.00b
Gu4CL11	2.73±0.76b	2.06±0.34b	2.54±1.57b	5.74±1.90a	2.58±1.91b	0.78±1.08a	0.39±0.54a	0.62±0.17a	0.08±0.09a	0.33±0.45a

基因 Gene	地上部分表达量Aboveground part expression level					地下部分表达量Underground part expression level
基因 Gene	0 h	2 h	6 h	12 h	24 h	0 h	2 h	6 h	12 h	24 h
Gu4CL1	9.26±3.08b	8.59±2.55b	4.66±0.92b	13.72±4.02b	43.79±23.22a	0.82±0.74ab	0.85±0.32ab	0.52±0.38b	0.85±0.12ab	1.61±0.69a
Gu4CL2	36.58±13.22ab	46.83±12.04ab	23.09±13.52b	36.93±11.23ab	63.23±30.27a	6.41±4.38c	56.17±14.06a	19.89±8.03bc	34.03±3.09b	33.60±15.57b
Gu4CL3	36.64±10.85ab	30.14±8.68ab	41.21±3.50a	26.91±7.34bc	15.32±2.43c	21.27±5.66a	22.13±9.76a	20.57±3.15a	17.88±1.53ab	9.18±3.34b
Gu4CL4	6.87±3.91a	10.57±2.06a	5.32±2.31a	9.03±2.71a	8.79±7.11a	36.95±8.83b	124.52±66.94a	62.49±14.25ab	59.81±16.80ab	55.44±37.15ab
Gu4CL5	14.77±5.17c	38.94±14.93a	15.80±4.80c	5.61±1.88c	48.45±23.02a	2.53±1.15c	17.99±6.00a	4.93±3.35b	4.98±2.13b	2.42±1.24b
Gu4CL6	1.66±0.31b	2.63±0.90ab	2.19±0.81b	4.60±1.04a	3.71±2.19ab	5.15±0.17a	14.88±8.00a	9.93±2.72a	13.23±6.07a	6.76±5.10a
Gu4CL7	0.04±0.05a	0.00±0.00a	0.01±0.02a	0.04±0.04a	0.36±0.62a	0.93±0.29a	0.32±0.32a	0.52±0.32a	0.45±0.32a	0.52±0.37a
Gu4CL8	3.30±2.70a	3.47±1.69a	2.78±0.99a	7.81±4.93a	4.03±2.33a	0.38±0.32a	0.36±0.09a	0.34±0.18a	0.49±0.25a	0.86±0.55a
Gu4CL9	0.46±0.38ab	0.27±0.08b	0.35±0.22b	1.04±0.47a	0.80±0.30ab	0.40±0.40a	0.06±0.05a	0.19±0.13a	0.28±0.21a	0.20±0.22a
Gu4CL10	1.21±0.58ab	0.32±0.07b	1.40±1.52ab	5.35±5.30a	0.49±0.50b	0.25±0.22a	0.00±0.00b	0.00±0.00b	0.00±0.00b	0.00±0.00b
Gu4CL11	2.73±0.76b	2.06±0.34b	2.54±1.57b	5.74±1.90a	2.58±1.91b	0.78±1.08a	0.39±0.54a	0.62±0.17a	0.08±0.09a	0.33±0.45a

蛋白名称 Protein name	氨基酸数量/aa Number of amino acid	分子量/ku Molecular weight	等电点 pI	不稳定指数 Instability index	脂肪系数 Aliphatic index	亲疏水性 GRAVY	亚细胞定位 Subcellular location
Gu4CL1	556	60.865 43	6.91	38.65	101.78	0.201	质膜
Gu4CL2	588	64.016 18	5.60	38.64	95.14	-0.081	叶绿体
Gu4CL3	575	62.282 92	6.61	45.45	95.01	-0.015	质膜
Gu4CL4	525	56.811 20	5.48	41.18	93.81	0.063	质膜
Gu4CL5	526	56.783 97	6.11	33.75	96.62	-0.006	质膜
Gu4CL6	542	59.158 83	6.06	29.74	98.80	0.020	质膜
Gu4CL7	519	56.729 70	5.70	31.76	101.19	0.093	质膜
Gu4CL8	614	67.857 14	9.05	40.09	97.75	0.005	质膜
Gu4CL9	570	61.790 69	6.15	44.15	100.23	0.092	质膜
Gu4CL10	420	46.314 00	6.71	30.45	93.05	-0.011	质膜
Gu4CL11	553	60.075 32	6.37	45.16	98.92	0.052	内质网

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