烟草MRS2/MGT基因家族的鉴定与表达分析

doi:10.11924/j.issn.1000-6850.casb2021-0793

摘要/Abstract

摘要：

为鉴定烟草基因组中的镁转运蛋白(MGT)基因,探讨MGT对烟株镁离子运输的机制,首先以水稻和拟南芥基因组中的镁转运蛋白家族基因为参考序列,应用同源序列比对方法,从烟草基因组中鉴定获得7个NtMGTs,均具有保守的Gly-Met-Asn三肽基序。再设计不同镁供应水平和光强处理的实验,结果发现NtMGTs的表达具有组织特异性且受光诱导表达。强光处理使烟株根中NtMGT1与叶片中的NtMGT2、NtMGT4、NtMGT5的表达量上升。随着镁供应浓度的上升,根系NtMGT1基因表达量提高且与根系镁含量变化趋势一致,推断NtMGT1基因主要介导烟株根系对镁的吸收;而叶片中NtMGT2、NtMGT4与NtMGT5基因表达量先上升后降低,说明3个基因属于高亲和镁离子转运系统。高温强光胁迫下,适当施镁能提高烟株地下部NtMGT1表达,促进根系吸收镁离子;提升地上部NtMGT2、NtMGT4与NtMGT5基因表达,促进镁离子的转运,保障烟株正常的生长发育。研究结果可为烟草生产上合理施镁提供理论指导。

关键词: 烟草, 镁营养, 镁转运蛋白, MRS2/MGT基因家族, 高温强光胁迫

Abstract:

To identify magnesium transporter (MGT) gene in tobacco genome and explore the mechanism of magnesium ion transport by MGT in tobacco plant, seven NtMGTs with conservative Gly-Met-Asn tripeptide motifs were identified from tobacco genome by homologous sequence alignment using the magnesium transporter family genes in rice and Arabidopsis genomes as reference sequences. The results showed that the expression of NtMGTs was tissue-specific and light-induced under different magnesium supply levels and light intensities. The expression of NtMGT1 in roots and NtMGT2, NtMGT4 and NtMGT5 in leaves of tobacco plants increased under strong light intensity. With the increase of magnesium supply, the expression of NtMGT1 gene in the root system increased, which was consistent with the change trend of magnesium content in the root system, suggesting that NtMGT1 gene mainly mediated the absorption of magnesium in the root system of tobacco plant. The expression levels of NtMGT2, NtMGT4 and NtMGT5 genes in leaves increased first and then decreased, indicating that the three genes belonged to a high affinity magnesium ion transport system. These results implied that under the stress of high temperature and strong light intensity, appropriate magnesium supply could increase the expression of NtMGT1 in the root part of tobacco plants, promote the absorption of magnesium ion by the root system, enhance the gene expression of NtMGT2, NtMGT4 and NtMGT5 in shoot part of tobacco plants, promote the transport of magnesium ion, and ensure the normal growth and development of tobacco plants. The results of this study can provide theoretical guidance for reasonable application of magnesium in tobacco production.

Key words: tobacco, magnesium supply, magnesium transporter protein, MRS2/MGT gene family, strong light intensity and high temperature stress

中图分类号:

Q94
S572

吴题, 魏晓玲, 冯常青, 黄云侠, 徐时长, 邱福祥, 郑英洁, 李文卿, 何华勤. 烟草MRS2/MGT基因家族的鉴定与表达分析[J]. 中国农学通报, 2022, 38(16): 18-26.

WU Ti, WEI Xiaoling, FENG Changqing, HUANG Yunxia, XU Shichang, QIU Fuxiang, ZHENG Yingjie, LI Wenqing, HE Huaqin. Identification and Expression Analysis of MRS2/MGT Gene Family in Tobacco[J]. Chinese Agricultural Science Bulletin, 2022, 38(16): 18-26.

图/表 9

参考文献 26

[1]	邓超, 王能如, 王东胜,等. 烟草镁素营养研究进展[J]. 安徽农业科学杂志, 2008(19):8123-8126.
[2]	关广晟. 烟草镁吸收积累规律与调控研究[D]. 湖南农业大学, 2007.
[3]	MENGUTAY M, CEYLAN Y, KUTMAN U B, et al. Adequate magnesium nutrition mitigates adverse effects of heat stress on maize and wheat[J]. Plant & soil, 2013, 368(1-2):57-72.
[4]	CAKMAK I. Magnesium in crop production, food quality and human health[J]. Plant & soil, 2013, 368(1):1-4.
[5]	HMIEL S P, SNAVELY M D, FLORER J B, et al. Magnesium transport in Salmonella typhimurium: characterization of magnesium influx and cloning of a transport gene[J]. Journal of bacteriology, 1986, 168(3):1444-1450. doi: 10.1128/jb.168.3.1444-1450.1986 URL
[6]	SNAVELY M D, FLORER J B, MILLER C G, et al. Magnesium transport in Salmonella typhimurium: 28Mg²⁺transport by the CorA, MgtA, and MgtB systems[J]. Journal of bacteriology, 1989, 171(9):4761-4766. doi: 10.1128/jb.171.9.4761-4766.1989 URL
[7]	SMITH R L, THOMPSON L J, MAGUIRE M E. Cloning and characterization of MgtE, a putative new class of Mg²⁺ transporter from Bacillus firmusOF4[J]. Journal of bacteriology, 1995, 177(5):1233-1238. doi: 10.1128/jb.177.5.1233-1238.1995 URL
[8]	王勇军. 甘蔗MGT基因家族鉴定与缺镁胁迫的转录组动力学研究[D]. 福州: 福建农林大学, 2019.
[9]	SAITO T, KOBAYASHI N I, TANOI K, et al. Expression and functional analysis of the CorA-MRS2-ALR-type magnesium transporter family in rice[J]. Plant and cell physiology, 2013, 54(10):1673-1683. doi: 10.1093/pcp/pct112 URL
[10]	LI H Y, DU H M, HUANG K F, et al. Identification, and functional and expression analyses of the CorA/MRS2/MGT-type magnesium transporter family in maize[J]. Plant and cell physiology, 2016, 57(6):1153-1168. doi: 10.1093/pcp/pcw064 URL
[11]	ZHANG L, WEN A, WU X Q, et al. Molecular identification of the magnesium transport gene family in Brassica napus[J]. Plant physiology and biochemistry, 2019, 136(6):204-214. doi: 10.1016/j.plaphy.2019.01.017 URL
[12]	阳江华, 秦云霞, 方永军,等. 巴西橡胶树镁离子转运蛋白基因HbMGT10的克隆及表达分析[J]. 热带作物学报, 2016, 37(12):2353.
[13]	LI L, TUTONE A F, DRUMMOND R, et al. A novel family of magnesium transport genes in Arabidopsis[J]. The plant cell, 2001, 13(12):2761-2775. doi: 10.1105/tpc.010352 URL
[14]	范才银. 不同施镁水平对烤烟生长发育和产量及质量的影响研究[D]. 长沙: 湖南农业大学, 2007.
[15]	CHEN Z C, YAMAJI N, MOTOYAMA R, et al. Up-regulation of a magnesium transporter gene OsMGT1 is required for conferring aluminum tolerance in rice[J]. Plant physiology, 2012, 159(4):1624-1633. doi: 10.1104/pp.112.199778 URL
[16]	CHEN Z C, YAMAJI N, HORIE T, et al. A magnesium transporter OsMGT1 plays a critical role in salt tolerance in rice[J]. Plant physiology, 2017, 174(3):1837-1849. doi: 10.1104/pp.17.00532 URL
[17]	WOFFELMAN C. DNAMAN for Windows, Version 5.2.10[Z]. Lynon Biosoft, Institute of Molecular Plant Sciences, Netherlands: Leiden University, 2004.
[18]	KUMAR S, STECHER G, TAMURA K. MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets[J]. Molecular biology and evolution, 2016, 33(7):1870-1874. doi: 10.1093/molbev/msw054 URL
[19]	LALITHA S. Primer premier 5[J]. Biotech software and internet report, 2000, 1(6):270-272. doi: 10.1089/152791600459894 URL
[20]	WATERS B M. Moving magnesium in plant cells[J]. New phytologist, 2011, 190(3):510-513. doi: 10.1111/j.1469-8137.2011.03724.x URL
[21]	SHIU S H, KARLOWSKI W M, PAN R S, et al. Comparative analysis of the receptor-like kinase family in Arabidopsis and rice[J]. The plant cell, 2004, 16(5):1220-1234. doi: 10.1105/tpc.020834 URL
[22]	MARTNEZ M, ABRAHAM Z, CARBONERO P, et al. Comparative phylogenetic analysis of cystatin gene families from Arabidopsis, rice and barley[J]. Molecular genetics and genomics, 2005, 273(5):423-432. doi: 10.1007/s00438-005-1147-4 URL
[23]	GARDNER R C. Genes for magnesium transport[J]. Current opinion in plant biology, 2003, 6(3):263-267. doi: 10.1016/S1369-5266(03)00032-3 URL
[24]	许花, 杨泽峰, 顾世梁. 基因组水平上拟南芥和水稻mrs2基因家族的比较系统发生分析[J]. 中国农业科技导报, 2007, 9(6):56-63.
[25]	GEBERT M, MESCHENMOSER K, SVIDOVA S, et al. A root-expressed magnesium transporter of the MRS2/MGT gene family in Arabidopsis thaliana allows for growth in low-Mg²⁺environments[J]. The plant cell, 2009, 21(12):4018-4030. doi: 10.1105/tpc.109.070557 URL
[26]	DRUMMOND R, TUTONE A, LI Y C, et al. A putative magnesium transporter AtMRS2-11 is localized to the plant chloroplast envelope membrane system[J]. Plant science, 2006, 170(1):78-89. doi: 10.1016/j.plantsci.2005.08.018 URL

基因名	上游引物(5'-3')	下游引物(5'-3')
NtMGT1	GGTTGCCATCTTCGGGGTT	AGCGCTTATTTCCCTCATAGGCTC
NtMGT2	GCAGCCGATATACTCCGACCAAAC	ATGACCGTATTCCCGTCCCCTTC
NtMGT4	GTGGATCAGGTGCAGCCAGTTG	TGCCCGAAGTAAGCCTCCAATAAC
NtMGT5	ATGGGTGGTAATCATTAGTGCTATC	CTTGGATGCTGTATGGTTCCTTTC
β-actin	GATCTTGCTGGTCGTGATCT	ACTTCCGGACATCTGAACCT

基因名	上游引物(5'-3')	下游引物(5'-3')
NtMGT1	GGTTGCCATCTTCGGGGTT	AGCGCTTATTTCCCTCATAGGCTC
NtMGT2	GCAGCCGATATACTCCGACCAAAC	ATGACCGTATTCCCGTCCCCTTC
NtMGT4	GTGGATCAGGTGCAGCCAGTTG	TGCCCGAAGTAAGCCTCCAATAAC
NtMGT5	ATGGGTGGTAATCATTAGTGCTATC	CTTGGATGCTGTATGGTTCCTTTC
β-actin	GATCTTGCTGGTCGTGATCT	ACTTCCGGACATCTGAACCT

基因名称	基因编号	转录编号	蛋白编号
NtMGT1	107824590	XM_016651392.1	XP_016506878.1
NtMGT2	107797716	XM_016620625.1	XP_016476111.1
NtMGT3	107775786	XM_016595567.1	XP_016451053.1
NtMGT4	107765702	XM_016584374.1	XP_016439860.1
NtMGT5-1	107778350	XM_016598596.1	XP_016454083.1
NtMGT5-2	107778350	XM_016598597.1	XP_016454082.1
NtMGT6	107825361	XM_016652209.1	XP_016507695.1
NtMGT7	107791732	XM_016613852.1	XP_016469338.1

基因名称	基因编号	转录编号	蛋白编号
NtMGT1	107824590	XM_016651392.1	XP_016506878.1
NtMGT2	107797716	XM_016620625.1	XP_016476111.1
NtMGT3	107775786	XM_016595567.1	XP_016451053.1
NtMGT4	107765702	XM_016584374.1	XP_016439860.1
NtMGT5-1	107778350	XM_016598596.1	XP_016454083.1
NtMGT5-2	107778350	XM_016598597.1	XP_016454082.1
NtMGT6	107825361	XM_016652209.1	XP_016507695.1
NtMGT7	107791732	XM_016613852.1	XP_016469338.1

基因	基因编号	氨基酸数目	分子量/kDa	结构域	等电位点	膜螺旋结构数	亚细胞定位
NtMGT1	107824590	445	50.54	Mrs2_Mfm1p-like	5.20	2	Plasma membrane
NtMGT2	107797716	499	54.95	Mrs2_Mfm1p-like	4.83	2	Chloroplast
NtMGT3	107775786	499	54.97	Mrs2_Mfm1p-like	4.83	2	Chloroplast
NtMGT4	107765702	393	44.42	Mrs2_Mfm1p-like	5.10	3	Plasma membrane
NtMGT5	107778350	393	44.42	Mrs2_Mfm1p-like	5.10	3	Plasma membrane
NtMGT6	107825361	372	41.71	Mrs2_Mfm1p-like	4.87	2	Chloroplast
NtMGT7	107791732	442	49.84	Mrs2_Mfm1p-like	6.09	2	Plasma membrane