Transcriptome Sequencing in the Leaves of Pontederia cordata with Cadmium Exposure and Gene Mining in Phenypropanoid Pathways

doi:10.13560/j.cnki.biotech.bull.1985.2021-1151

Abstract

Abstract:

To provide theoretical base for the mechanism of phenylpropane metabolic pathway involved in Pontederia cordata against heavy metal exposure,and to analyze the transcriptome sequences in the leaves of P. cordata exposed to cadmium（Cd）,the obtained Unigenes were compared and annotated with seven databases including NR,NT,PFAM,KOG,Swiss Prot,and KEGG as well as GO,and the tendency of differentially expressed genes（DEGs）were analyzed,and DEGs involved in phenypropanoid metabolic pathway were mined. Results showed as follows. 1）A total of 221 392 unigenes were obtained,of which 170 175 unigenes were successfully annotated in the public database,and 6 506 unigenes were annotated for 31 secondary metabolic pathways. 2）A total of 168 355 were obtained after comparing the Unigenes from the leaves of P. cordata with dadabase Swiss Prot and NR. And 84 673 CDS sequences were obtained by Esacan software. 3）Trend analysis demonstrated that 20 025 DEGs were clustered into three significant profiles including two down-regulated profiles（cluster 0 covering 2 631 DEGs and cluster 1 covering 3 153 DEGs）and one up-regulated profiles（cluster 5 covering 3 733 DEGs）. 4）With Cd²⁺ the exposure,there were 26 DEGs identified in P. cordata leaves in 6 h and 48 h by further mining the transcriptome data,which encoded 3 crucial enzymes in the common pathway of phenylpropane metabolism,5 crucial enzymes related to lignin biosynthesis,and 7 crucial enzymes responsible for flavonoid biosynthesis. P. cordata is involved in the accumulation and detoxification of Cd in the leaves by regulating the biosynthesis of guaiacyl lignin and up-regulating the gene expression of cinnamate-4-hydroxylase（C4H）,anthocyanin synthase（ANS）,anthocyanin-3-O -glucosyltransferase（3GT）,caffeonyl coenzyme A-O methyltransferase（CCoAOMT）,and flavonol 3-methyltransferase（F3OMT）. The genes involved in phenylpropane metabolic pathway in the leaves of P. cordata against Cd stress is preliminarily revealed by transcriptome sequencing,which provides a theoretical basis for further research on the function of phenylpropane metabolic pathway in the resistance of the plant to heavy metal stress and related mechanisms.

Key words: cadmium, Pontederia cordata, transcriptome in leaves, phenypropanoid pathway, gene mining

XIN Jian-pan, LI Yan, ZHAO Chu, TIAN Ru-nan. Transcriptome Sequencing in the Leaves of Pontederia cordata with Cadmium Exposure and Gene Mining in Phenypropanoid Pathways[J]. Biotechnology Bulletin, 2022, 38(6): 198-210.

Figures/Tables 12

References 43

[1]	张阿芳, 张庆, 代惠萍, 等. 镉胁迫对银灰杨根和叶片渗透调节物质的影响[J]. 西北林学院学报, 2018, 33(2):83-87.
	Zhang AF, Zhang Q, Dai HP, et al. Effects of cadmium stress on the osmotic adjustment substance of Populus canescens in leaves and roots[J]. J Northwest For Univ, 2018, 33(2):83-87.
[2]	余红兵, 杨知建, 肖润林, 等. 梭鱼草(Pontederia cordata)拦截沟渠中氮、磷的效果研究[J]. 农业现代化研究, 2012, 33(4):508-512.
	Yu HB, Yang ZJ, Xiao RL, et al. Research of ditch interception effect of nitrogen and phosphorus of Pontederia cordata[J]. Res Agric Mod, 2012, 33(4):508-512.
[3]	张景雯, 田如男. 四种植物对模拟的城市景观污水的净化效果[J]. 湿地科学, 2018, 16(1):85-92.
	Zhang JW, Tian RN. Purification effect of four kinds of aquatic plants on simulated urban landscape polluted water[J]. Wetl Sci, 2018, 16(1):85-92.
[4]	夏梦华, 刘铭羽, 郭宁宁, 等. 美人蕉、梭鱼草和黄菖蒲人工湿地系统对养猪废水的脱氮特征研究[J]. 生态与农村环境学报, 2020, 36(8):1080-1088.
	Xia MH, Liu MY, Guo NN, et al. Study on nitrogen removal characteristics of swine wastewater in the constructed wetland systems of Canna indica, Pontederia cordata and Iris pseudacorus[J]. J Ecol Rural Environ, 2020, 36(8):1080-1088.
[5]	许蓝心, 田如男. 梭鱼草化感物质丁二酸对微囊藻和栅藻生长及竞争的影响[J]. 生态学杂志, 2019, 38(3):770-777.
	Xu LX, Tian RN. Effects of succinic acid, an allelopathic substance of Pontederi cordata, on the growth and competition of Microcystis aeruginosa and Scenedesmus obliquus[J]. Chin J Ecol, 2019, 38(3):770-777.
[6]	赵楚, 钱燕萍, 田如男. 梭鱼草化感物质丁二酸、肉桂酸及香草酸对铜绿微囊藻生长的抑制效应[J]. 浙江农林大学学报, 2020, 37(6):1105-1111.
	Zhao C, Qian YP, Tian RN. Inhibitory effect of succinic acid, cinnamic acid and vanillic acid from Pontederia cordata on Microcystis aeruginosa[J]. J Zhejiang A F Univ, 2020, 37(6):1105-1111.
[7]	高军侠, 陶贺, 党宏斌, 等. 睡莲、梭鱼草对铜污染水体的修复效果研究[J]. 地球与环境, 2016, 44(1):96-102.
	Gao JX, Tao H, Dang HB, et al. Phytoremediation of copper-contaminated water by Nymphaea tetragona and Pontederia cordata[J]. Earth Environ, 2016, 44(1):96-102.
[8]	刘寿涛, 杨蕊嘉, 何钟响, 等. 灌溉水湿地净化系统中植物对镉的去除效果研究[J]. 灌溉排水学报, 2019, 38(10):47-54.
	Liu ST, Yang RJ, He ZX, et al. Effect of plant pond and constructed wetland system on irrigation water purification and rice cadmium control[J]. J Irrigation Drainage, 2019, 38(10):47-54.
[9]	梅金星, 张平, 彭佩钦, 等. 人工湿地系统中梭鱼草和香蒲对镉积累的动态变化[J]. 水生态学杂志, 2020, 41(2):98-104.
	Mei JX, Zhang P, Peng PQ, et al. Cadmium accumulation in Pontederia cordata and Typha orientalis in a constructed wetland[J]. J Hydroecology, 2020, 41(2):98-104.
[10]	Xin JP, Ma SS, Li Y, et al. Pontederia cordata, an ornamental aquatic macrophyte with great potential in phytoremediation of heavy-metal-contaminated wetlands[J]. Ecotoxicol Environ Saf, 2020, 203:111024.
[11]	马思思, 辛建攀, 陈宜栋, 等. 铜对梭鱼草叶片保护酶活性、抗氧化物质及非蛋白巯基肽含量的影响[J]. 草业科学, 2020, 37(3):459-468.
	Ma SS, Xin JP, Chen YD, et al. Effects of copper on antixoidant enzyme activities, antioxidant and non-protein thiol content in Pontederia cordata’s leaves[J]. Pratacultural Sci, 2020, 37(3):459-468.
[12]	Xin JP, Ma S, Zhao C, et al. Cadmium phytotoxicity, related physiological changes in Pontederia cordata:antioxidative, osmoregulatory substances, phytochelatins, photosynthesis, and chlorophyll fluorescence[J]. Environ Sci Pollut Res Int, 2020, 27(33):41596-41608. doi: 10.1007/s11356-020-10002-z URL
[13]	Shi X, Sun H, Chen Y, et al. Transcriptome sequencing and expression analysis of cadmium(Cd)transport and detoxification related genes in Cd-accumulating Salix integra[J]. Front Plant Sci, 2016, 7:1577.
[14]	曹继敏, 李双财, 何德. 镉胁迫后旱柳转录组变化分析[J]. 生物技术通报, 2020, 36(7):32-39. doi: 10.13560/j.cnki.biotech.bull.1985.2019-1018
	Cao JM, Li SC, He D. Transcriptome analysis of Saliz matsudana under cadmium stress[J]. Biotechnol Bull, 2020, 36(7):32-39.
[15]	Pawlak-Sprada S, Arasimowicz-Jelonek M, Podgórska M, et al. Activation of phenylpropanoid pathway in legume plants exposed to heavy metals. Part I. Effects of cadmium and lead on phenylalanine ammonia-lyase gene expression, enzyme activity and lignin content[J]. Acta Biochim Pol, 2011, 58(2):211-216. pmid: 21503278
[16]	杨欣, 徐艳红, 魏建和, 等. 几种重要植物次生代谢防御反应物质的生物合成途径及分子调控机制研究进展[J]. 生物技术通讯, 2013, 24(2):285-289.
	Yang X, Xu YH, Wei JH, et al. Advances on the biosynthesis pathways and molecular regulation mechanism of several important defensive substances in plant sec ondary metabolism[J]. Lett Biotechnol, 2013, 24(2):285-289.
[17]	耿安静, 王旭, 李秋剑, 等. 花青素介导植物抗重金属胁迫机理的研究进展[J]. 南方农业学报, 2020, 51(1):80-90.
	Geng AJ, Wang X, Li QJ, et al. Mechanism of anthocyanins-mediated resistance to heavy metals stresses in plants:a review[J]. J South Agric, 2020, 51(1):80-90.
[18]	Mhiri R, Koubaa I, Chawech R, et al. New isoflavones with antioxidant activity isolated from Cornulaca monacantha[J]. Chem Biodivers, 2020, 17(12):e2000758.
[19]	Grabherr MG, Haas BJ, Yassour M, et al. Full-length transcriptome assembly from RNA-Seq data without a reference genome[J]. Nat Biotechnol, 2011, 29(7):644-652. doi: 10.1038/nbt.1883 pmid: 21572440
[20]	Li B, Dewey CN. RSEM:accurate transcript quantification from RNA-Seq data with or without a reference genome[J]. BMC Bioinformatics, 2011, 12:323. doi: 10.1186/1471-2105-12-323 URL
[21]	Ernst J, Bar-Joseph Z. STEM:a tool for the analysis of short time series gene expression data[J]. BMC Bioinformatics, 2006, 7:191. doi: 10.1186/1471-2105-7-191 URL
[22]	魏利斌, 苗红梅, 张海洋. 芝麻发育转录组分析[J]. 中国农业科学, 2012, 45(7):1246-1256.
	Wei LB, Miao HM, Zhang HY. Transcriptomic analysis of sesame development[J]. Sci Agric Sin, 2012, 45(7):1246-1256.
[23]	贾新平, 叶晓青, 梁丽建, 等. 基于高通量测序的海滨雀稗转录组学研究[J]. 草业学报, 2014, 23(6):242-252.
	Jia XP, Ye XQ, Liang LJ, et al. Transcriptome characteristics of Paspalum vaginatum analyzed with Illumina sequencing technology[J]. Acta Prataculturae Sin, 2014, 23(6):242-252.
[24]	王兴春, 谭河林, 陈钊, 等. 基于RNA-Seq技术的连翘转录组组装与分析及SSR分子标记的开发[J]. 中国科学:生命科学, 2015, 45(3):301-310.
	Wang XC, Tan HL, Chen Z, et al. Assembly and characterization of the transcriptome and development of SSR markers in Forsythia suspensa based on RNA-seq technology[J]. Sci Sin Vitae, 2015, 45(3):301-310. doi: 10.1360/N052014-00273 URL
[25]	袁灿, 彭芳, 钟文娟, 等. 赶黄草的转录组测序及分析[J]. 中草药, 2017, 48(21):4507-4514.
	Yuan C, Peng F, Zhong WJ, et al. De novo assembly and transcriptome characterization of Penthorum chinense[J]. Chin Tradit Herb Drugs, 2017, 48(21):4507-4514.
[26]	王继华, 罗群胜, 梅瑜, 等. 张溪香芋转录组测序及生物信息学分析[J]. 基因组学与应用生物学, 2020, 39(12):5697-5704.
	Wang JH, Luo QS, Mei Y, et al. Transcriptome sequencing and bioinformatic analysis of zhangxi taro[J]. Genom Appl Biol, 2020, 39(12):5697-5704.
[27]	韦晓霞, 王小安, 陈瑾, 等. 百香果低温胁迫转录组及茉莉酸代谢基因分析[J]. 核农学报, 2021, 35(4):815-825. doi: 10.11869/j.issn.100-8551.2021.04.0815
	Wei XX, Wang XA, Chen J, et al. Transcriptome and jasmin metabolism gene analysis of Passiflora edulia Sims under low temperature stress[J]. J Nucl Agric Sci, 2021, 35(4):815-825.
[28]	肖韵铮, 韩世明, 秦昭, 等. 滇黄精转录组测序及类黄酮合成相关基因的分析[J]. 河南农业大学学报, 2020, 54(6):931-940.
	Xiao YZ, Han SM, Qin Z, et al. Analysis of transcriptome sequencing and related genes of flavonoids biosynthesis from Polygonatum kingianum[J]. J Henan Agric Univ, 2020, 54(6):931-940.
[29]	张娜, 刘秀霞, 陈学森, 等. 基于转录组分析鉴定苹果茉莉素响应基因[J]. 植物学报, 2019, 54(6):733-743. doi: 10.11983/CBB18235
	Zhang N, Liu XX, Chen XS, et al. Identifying genes responsive to jasmonates in apple based on transcriptome analysis[J]. Chin Bull Bot, 2019, 54(6):733-743.
[30]	夏铭泽, 张雨, 余静雅, 等. 多裂骆驼蓬叶片转录组分析[J]. 广西植物, 2021, 41(4):503-513.
	Xia MZ, Zhang Y, Yu JY, et al. Transcriptome analysis for leaves of Peganum multisectum[J]. Guihaia, 2021, 41(4):503-513.
[31]	朱畇昊, 张梦佳, 李璐, 等. 夏枯草的转录组测序与次生代谢产物生物合成相关基因的挖掘[J]. 中草药, 2019, 50(5):1220-1226.
	Zhu YH, Zhang MJ, Li L, et al. Transcriptome analysis of Prunella vulgaris and identification of putative genes involved in secondary metabolism biosynthesis[J]. Chin Tradit Herb Drugs, 2019, 50(5):1220-1226.
[32]	张晓娜, 朴春兰, 董友魁, 等. 大豆根系应答重金属Cd胁迫的转录组分析[J]. 应用生态学报, 2017, 28(5):1633-1641. pmid: 29745202
	Zhang XN, Piao CL, Dong YK, et al. Transcriptome analysis of response to heavy metal Cd stress in soybean root[J]. Chin J Appl Ecol, 2017, 28(5):1633-1641. doi: 10.13287/j.1001-9332.201705.004 pmid: 29745202
[33]	热依麦阿依·阿布都艾尼, 陈静, 陈芸, 等. 盐胁迫下棉花根系的转录组分析及耐盐基因筛选[J]. 华南师范大学学报:自然科学版, 2020, 52(5):85-92.
	Reyimaiayi A, Chen J, Chen Y, et al. Transcriptome analysis and salt tolerance gene screening of cotton root under salt stress[J]. J South China Norm Univ:Nat Sci Ed, 2020, 52(5):85-92.
[34]	曹昆, 屈文言, 徐洪伟, 等. 冷驯化下的牛皮杜鹃转录组变化分析[J]. 吉林师范大学学报:自然科学版, 2021, 42(1):92-98.
	Cao K, Qu WY, Xu HW, et al. Analysis of the transcriptome changes of Rhododendron chrysanthum Pall under cold acclimation[J]. J Jilin Norm Univ:Nat Sci Ed, 2021, 42(1):92-98.
[35]	Chang A, Lim MH, Lee SW, et al. Tomato phenylalanine ammonia-lyase gene family, highly redundant but strongly underutilized[J]. J Biol Chem, 2008, 283(48):33591-33601. doi: 10.1074/jbc.M804428200 URL
[36]	刘畅, 俸婷婷, 刘雄伟, 等. 苗药八爪金龙转录组测序与次生代谢产物合成相关基因的挖掘[J]. 中草药, 2021, 52(5):1434-1447.
	Liu C, Feng TT, Liu XW, et al. Transcriptome analysis and identification of related genes involved in secondary metabolism biosynthesis in Ardisia crispa[J]. Chin Tradit Herb Drugs, 2021, 52(5):1434-1447.
[37]	Yang YJ, Cheng LM, Liu ZH. Rapid effect of cadmium on lignin biosynthesis in soybean roots[J]. Plant Sci, 2007, 172(3):632-639. doi: 10.1016/j.plantsci.2006.11.018 URL
[38]	Kováčik J, Klejdus B. Dynamics of phenolic acids and lignin accumulation in metal-treated Matricaria chamomilla roots[J]. Plant Cell Rep, 2008, 27(3):605-615. doi: 10.1007/s00299-007-0490-9 URL
[39]	董娟娥, 张康健, 梁宗锁. 植物次生代谢与调控[M]. 杨凌: 西北农林科技大学出版社, 2009.
	Dong JE, Zhang KJ, Liang ZS. Plant secondary metabolism and its regulation[M]. Yangling: Northwest A & F University Press, 2009.
[40]	邓灿辉, 唐蜻, 戴志刚, 等. 不同叶用黄麻种质对重金属吸附的差异及其机制分析[J]. 农业资源与环境学报, 2020, 37(3):438-444.
	Deng CH, Tang Q, Dai ZG, et al. Differential adsorption performance and mechanism of leaf-used jute germplasm for heavy metal removal[J]. J Agric Resour Environ, 2020, 37(3):438-444.
[41]	李丹丹, 梁宗锁, 普布卓玛, 等. 干旱胁迫对紫花苜蓿黄酮类化合物含量及其合成途径关键酶活性的影响[J]. 西北植物学报, 2020, 40(8):1380-1388.
	Li DD, Liang ZS, Pu B, et al. Flavonoids contents and flavonoids synthetic key enzyme activities in alfalfa under drought stress[J]. Acta Bot Boreali Occidentalia Sin, 2020, 40(8):1380-1388.
[42]	Albert NW, Lewis DH, Zhang H, et al. Light-induced vegetative anthocyanin pigmentation in Petunia[J]. J Exp Bot, 2009, 60(7):2191-2202. doi: 10.1093/jxb/erp097 pmid: 19380423
[43]	郑雪良, 刘春荣, 王登亮, 等. 胡柚小青果的黄酮类化合物及抗氧化活性研究[J]. 浙江农业学报, 2015, 27(7):1185-1191.
	Zheng XL, Liu CR, Wang DL, et al. Flavonoids contents and antioxidant activities in Citrus paradisi cv. Changshan Huyou during development[J]. Acta Agric Zhejiangensis, 2015, 27(7):1185-1191.

	Min length/bp	Max length/bp	Mean length/bp	N₅₀	N₉₀
Transcripts	201	25 638	1 542	2 554	746
Unigenes	201	25 638	1 753	2 609	883

	Min length/bp	Max length/bp	Mean length/bp	N₅₀	N₉₀
Transcripts	201	25 638	1 542	2 554	746
Unigenes	201	25 638	1 753	2 609	883

数据库 Annotated database	Unigenes数量 Number of unigenes	占比 Percentage/%
Annotated in NR	158 229	71.47
Annotated in NT	107 290	48.46
Annotated in KO	67 629	30.54
Annotated in SwissProt	121 487	54.87
Annotated in PFAM	98 391	44.44
Annotated in GO	98 391	44.44
Annotated in KOG	49 597	22.40
Annotated in all databases	22 729	10.26
Annotated in at least one database	170 175	76.86
Total unigenes	221 392	100

数据库 Annotated database	Unigenes数量 Number of unigenes	占比 Percentage/%
Annotated in NR	158 229	71.47
Annotated in NT	107 290	48.46
Annotated in KO	67 629	30.54
Annotated in SwissProt	121 487	54.87
Annotated in PFAM	98 391	44.44
Annotated in GO	98 391	44.44
Annotated in KOG	49 597	22.40
Annotated in all databases	22 729	10.26
Annotated in at least one database	170 175	76.86
Total unigenes	221 392	100

序号 No.	通路 Pathway	通路_ID Pathway_ID	Unigenes数量 Numbers of unigenes
1	苯丙素生物合成Phenylpropanoid biosynthesis	ko00940	696
2	缬氨酸、亮氨酸和异亮氨酸降解Valine,leucine and isoleucine degradation	ko00280	621
3	萜类化合物骨架生物合成Terpenoid backbone biosynthesis	ko00900	597
4	类胡萝卜素生物合成Carotenoid biosynthesis	ko00906	488
5	酪氨酸代谢Tyrosine metabolism	ko00350	426
6	泛醌及其他萜类醌生物合成Ubiquinone and other terpenoid-quinone biosynthesis	ko00130	370
7	苯丙氨酸、酪氨酸和色氨酸生物合成Phenylalanine,tyrosine and tryptophan biosynthesis	ko00400	346
8	烟酸和烟酰胺代谢Nicotinate and nicotinamide metabolism	ko00760	326
9	苯丙烷代谢Phenylalanine metabolism	ko00360	291
10	色氨酸代谢Tryptophan metabolism	ko00380	269
11	类固醇生物合成Steroid biosynthesis	ko00100	235
12	类黄酮生物合成Flavonoid biosynthesis	ko00941	222
13	异喹啉生物碱生物合成Isoquinoline alkaloid biosynthesis	ko00950	216
14	莨菪烷、哌啶和吡啶生物碱生物合成Tropane,piperidine and pyridine alkaloid biosynthesis	ko00960	197
15	玉米素生物合成Zeatin biosynthesis	ko00908	197
16	二苯乙烯、二芳基庚烷和姜酚生物合成Stilbenoid,diarylheptanoid and gingerol biosynthesis	ko00945	155
17	亮氨酸代谢Linoleic acid metabolism	ko00591	143
18	缬氨酸、亮氨酸和异亮氨酸生物合成Valine,leucine and isoleucine biosynthesis	ko00290	139
19	柠檬烯和蒎烯降解Limonene and pinene degradation	ko00903	128
20	油菜素内酯生物合成Brassinosteroid biosynthesis	ko00905	97
21	二萜生物合成Diterpenoid biosynthesis	ko00904	83
22	黄酮和黄酮醇生物合成Flavone and flavonol biosynthesis	ko00944	55
23	芥子油苷生物合成Glucosinolate biosynthesis	ko00966	58
24	单萜的生物合成Monoterpenoid biosynthesis	ko00902	48
25	倍半萜和三萜生物合成Sesquiterpenoid and triterpenoid biosynthesis	ko00909	44
26	咖啡因代谢Caffeine metabolism	ko00232	28
27	花青素生物合成Anthocyanin biosynthesis	ko00942	14
29	甜菜红碱生物合成Betalain biosynthesis	ko00965	10
30	异黄酮生物合成Isoflavonoid biosynthesis	ko00943	6
31	吲哚生物碱生物合成Indole alkaloid biosynthesis	ko00901	1