Scientia Agricultura Sinica ›› 2013, Vol. 46 ›› Issue (5): 889-897.doi: 10.3864/j.issn.0578-1752.2013.05.003

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

The Expression Analysis and Screening of Interaction Protein of Mitogen-Activated Protein Kinase (CaMAPKK2) in Salt-Stress Signal Pathways of Chenopodium album

 CHEN  Sha-Sha, HE  Zhuan-Zhuan, JIANG  Sheng-Xiu, LI  Xiao-Rong, XING  Jia-Jia, 吕Xiu-Yun , LAN  Hai-Yan   

  1. College of Life Science and Technology, Xinjiang University/Xinjiang Key Laboratory of Biological Resources and Genetic Engineering, Urumqi 830046
  • Received:2012-11-14 Online:2013-03-01 Published:2013-01-16

PDF

1548

Abstract: 【Objective】The aim of the present study is to understand the mechanism of stress signal transduction to induce the response of adverse tolerance with Chenopodium album, an adverse-tolerant plant species, through analysis of the expression pattern of mitogen-activated protein kinase gene (MAPKK) under different abiotic stresses and screening of interaction proteins of MAPKK in MAPK signal transduction pathways. 【Method】 Using the total RNA from the leaf of C. album as the template, the expression patterns of MAPKK under NaCl, H2O2 and ABA were analyzed by quantitative PCR technique. The full-length cDNA sequence of MAPKK was obtained by combined reverse transcription-PCR (RT-PCR) and RACE techniques. The yeast two-hybrid technique was employed to analyze the interaction components of MAPKK in salt-stress signal pathway. 【Result】 A full length cDNA fragment of MAPKK gene from C. album included a 1 089 bp ORF, which encodes with 362 putative amino acids, named as CaMAPKK2. qPCR analysis showed that CaMAPKK2 gene was significantly induced by salt and H2O2 but not by ABA stress, at least in the present study. DPI or Na2WO4 (inhibitor of H2O2 or ABA synthesis) significantly inhibited the expression of CaMAPKK2 in C. album when exposed to 300 mmol.L-1 NaCl stress. By using the CaMAPKK2 as bait protein, five potential clones which may interact with CaMAPKK2 were obtained by the first screening of yeast two hybridization. Sequencing result indicated that one 794 bp fragment out of 5 cDNA showed correct reading frame, and shared identities of 79% and 78% in nucleotide sequence to AgTHI1 and AtTHI1 (thiazole biosynthetic enzyme) from Alnus glutinosa and Arabidopsis thaliana, respectively, while other 4 cDNA sequences had no correct reading frame. 【Conclusion】CaMAPKK2 gene was significantly induced by salt and H2O2 stress, suggesting that salt stress might induce the accumulation of H2O2 and ABA, which in turn leads to expression increasing of MAPKK. More positive clones should be acquired and the relevant experiments (e.g. Pull down, CoIP) should be performed for screening and verifying the interaction component of CaMAPKK2.

Key words: Chenopodium album , salt stress , signal transduction pathway , MAPKK , yeast two-hybrid technique , quantitative PCR

[1] Samajiova O, Plihal O, Mohamed A Y, Hirt H, Samaj J. Improvement of stress tolerance in plants by genetic manipulation of mitogen-activated protein kinase. Biotechnology Advances, 2012, doi：10.1016/j.biotechadv.2011.12.002.

[2] Jonak C, Okresz I, Bogre I, Hirt H. Complexity, crosstalk and integration of plant MAP kinase signaling. Current Opinion in Plant Biology, 2002, 15: 415-424.

[3] MAPK Group. Mitogen-activated protein kinase cascades in plants: a new nomenclature. Trends in Plant Science, 2002, 7: 301-308.

[4] Xing Y, Jia W S, Zhang J H. AtMEK1 mediates stress-induced gene expression of CAT1 catalase by triggering H2O2 production in Arabidopsis. Journal of Experimental Botany, 2007, 58: 2969-2981.

[5] Nakagami H, Soukupova H, Schikora A, Zarsky V, Hirt H. A mitogen-activated protein kinase kinase kinase mediates reactive oxygen species homeostasis in Arabidopsis. Journal Biological Chemistry, 2006, 281: 38697-38704.

[6] Teige M, Scheikl E, Eulgem T, Doczi R, Ichimura K, Shinozaki K, Dang J L, Hirt H. The MKK2 pathway mediates cold and salt stress signaling in Arabidopsis. Molecular and Cellular Biology, 2004, 15: 141-152.

[7] Matsuoka D, Nanmori T, Sato K I, Fukami Y, Kikkawa U, Yasuda T. Activation of AtMEK1, an Arabidopsis mitogen-activated protein kinase kinase, in vitro and in vivo: Analysis of active mutants expressed in E.coli and generation of the active form in stress response in seedlings. The Plant Journal, 2002, 29: 637-647.

[8] Zhang X, Zhang L, Song C P. Hydrogen peroxide is involved in abscisic acid-induced stomatal closure in Vicia faba. Plant Physiolopy, 2001, 126: 1438-1448.

[9] Pei Z M, Murata Y, Schroeder J I. Calcium channels activated by hydrogen peroxide mediate abscisic acid signaling in guard cells. Nature, 2000, 406: 731-734.

[10] Jiang J, An G Y, Wang P C, Wang P T, Han J F, Jia Y B, Song C P. MAP kinase specifically mediates the ABA-induced H2O2 generation in guard cells of Vicia faba L.. Chinese Science Bulletin, 2003, 48: 181919-181926.

[11] Xing Y, Jia W, Zhang J. AtMKK1 mediates ABA-induced CAT1 expression and H2O2 production via AtMPK6-coupled signaling in Arabidopsis. The Plant Journal, 2008, 54: 440-451.

[12] Kovtun Y, Chiu W L, Tena G, Sheen J. Functional analysis of oxidative stress-activated mitogen-activated protein kinase cascade in plants. Proceedings of the National Academy of Science of the USA, 2000, 97: 2940-2945.

[13] Pitzschke A, Schikora A, Hirt H. MAPK cascade signalling networks in plant defence. Current Opinion in Plant Biology, 2009, 12: 421-426.

[14] Kumar K, Rao K P, Sharma P, Sinha A K. Differential regulation of rice mitogen activated protein kinase kinase (MKK) by abiotic stress. Plant Physiology of Biochemistry, 2008, 46: 891-897.

[15] Causier B, Davies B. Analysing protein-protein interactions with the yeast two-hybrid system. Plant Molecular Biology, 2002, 50: 855-870.

[16] Koek A, Komori M, Veenhuis M. A comparative study of peroxisomal structures in Hansenula polymorpha pex mutants. Fems Yeast Research, 2007, 7: 1126-1133.

[17] 邢福国, 张培军, 谭训刚, 徐永立. 酵母基因组的提取. 食品科学, 2007, 28(3): 210-212.

Xing F G, Zhang P J, Tan X G, Xu Y L. Extraction of the yeast genomic. Food Science, 2007, 28(3): 210-212. (in Chinese)

[18] Xu J, Li Y, Wang Y, Liu H X, Lei L, Yang H L, Liu G Q, Ren D T. Activation of MAPK kinase 9 induces ethylene and camalexin biosynthesis and enhances sensitivity to salt stress in Arabidopsis. Journal of Biology Chemistry, 2008, 283: 26996-27006.

[19] Zhang A N, Jiang M Y, Zhang J H, Tan M P, Hu X L. Mitogen-activated protein kinase is involved in abscisic acid-induced antioxidant defense and acts downstream of reactive oxygen species production in leaves of maize plants. Plant Physiology, 2006, 141: 475-487.

[20] Jiang M Y, Zhang A Y, Zhang J H, Tan M P, Hu X L. Mitogen-activated protein kinase is involved in abscisic acid-induced antioxidant defense and acts downstream of reactive oxygen species production in leaves of maize plants. Plant Physiology, 2006, 141: 475-487.

[21] Nie W F, Wang M M, Xia X J, Zhou Y H, Shi K, Chen Z X, Yu J Q. Silencing of tomato RBOH1 and MPK2 abolishes brassinosteroid- induced H2O2 generation and stress tolerance. Plant, Cell and Environment, 2012, 10: 1-15.

[22] Praekelt U, Akkermans A D, Meacock P A, Kammen A V, Bisseling T, Pawlowski K. Identification of agthi1, whose product is involved in biosynthesis of the thiamine precursor thiazole, in actinorhizal nodules of Alnus glutinosa. The Plant Journal, 1996, 10: 361-368.

[23] Machado C R, de Oliveira R L, Boiteux S, Praekelt U M, Meacock P A, Menck C F. Thi1, a thiamine biosynthetic gene in Arabidopsis thaliana, complements bacterial defects in DNA repair. Plant Molecular Biology, 1996, 31: 585-593.

[24] Jong H I, Hyoungseok L, Jitae K, Ho B K, Chung S A. Soybean MAPK, GMKK1 is dually regulated by phosphatidic acid and hydrogen peroxide and translocated to nucleus during salt stress. Molecules and Cells, 2012, 34: 271-278.

[25] Yang M, Wu Z, Fields S. Protein-peptide interactions analyzed with the yeast two-hybrid System. Nucleic Acids Research, 1995, 23: 1152-1156.

[26] James P, Halladay J, Craig E A. Genonic libraries and a host strain designed for highly efficient two-hybrid Selection in yeast. Genetics, 1996, 144: 1425-1436.
[1] HU YaLi,NIE JingZhi,WU Xia,PAN Jiao,CAO Shan,YUE Jiao,LUO DengJie,WANG CaiJin,LI ZengQiang,ZHANG Hui,WU QiJing,CHEN Peng. Effect of Salicylic Acid Priming on Salt Tolerance of Kenaf Seedlings [J]. Scientia Agricultura Sinica, 2022, 55(14): 2696-2708.
[2] ZHU ChunYan,SONG JiaWei,BAI TianLiang,WANG Na,MA ShuaiGuo,PU ZhengFei,DONG Yan,LÜ JianDong,LI Jie,TIAN RongRong,LUO ChengKe,ZHANG YinXia,MA TianLi,LI PeiFu,TIAN Lei. Effects of NaCl Stress on the Chlorophyll Fluorescence Characteristics of Seedlings of Japonica Rice Germplasm with Different Salt Tolerances [J]. Scientia Agricultura Sinica, 2022, 55(13): 2509-2525.
[3] LI XiaoJing,ZHANG SiYu,LIU Di,YUAN XiaoWei,LI XingSheng,SHI YanXia,XIE XueWen,LI Lei,FAN TengFei,LI BaoJu,CHAI ALi. Establishment and Application of Rapid Quantitative Detection of Viable Plasmodiophora brassicae by PMAxx-qPCR Method [J]. Scientia Agricultura Sinica, 2022, 55(10): 1938-1948.
[4] LIU Chuang,GAO Zhen,YAO YuXin,DU YuanPeng. Functional Identification of Grape Potassium Ion Transporter VviHKT1;7 Under Salt Stress [J]. Scientia Agricultura Sinica, 2021, 54(9): 1952-1963.
[5] ZHANG GuiYun,ZHU JingWen,SUN MingFa,YAN GuoHong,LIU Kai,WAN BaiJie,DAI JinYing,ZHU GuoYong. Analysis of Differential Metabolites in Grains of Rice Cultivar Changbai 10 Under Salt Stress [J]. Scientia Agricultura Sinica, 2021, 54(4): 675-683.
[6] WANG Jie,WU XiaoYu,YANG Liu,DUAN QiaoHong,HUANG JiaBao. Genome-Wide Identification and Expression Analysis of ACA Gene Family in Brassica rapa [J]. Scientia Agricultura Sinica, 2021, 54(22): 4851-4868.
[7] SHAO MeiQi,ZHAO WeiSong,SU ZhenHe,DONG LiHong,GUO QingGang,MA Ping. Effect of Bacillus subtilis NCD-2 on the Growth of Tomato and the Microbial Community Structure of Rhizosphere Soil Under Salt Stress [J]. Scientia Agricultura Sinica, 2021, 54(21): 4573-4584.
[8] CHEN Yuan,CAI He,LI Li,WANG LinJie,ZHONG Tao,ZHANG HongPing. Alternative Splicing of TNNT3 and Its Effect on the Differentiation of MuSCs in Goat [J]. Scientia Agricultura Sinica, 2021, 54(20): 4466-4477.
[9] WANG Na,ZHAO ZiBo,GAO Qiong,HE ShouPu,MA ChenHui,PENG Zhen,DU XiongMing. Cloning and Functional Analysis of Salt Stress Response Gene GhPEAMT1 in Upland Cotton [J]. Scientia Agricultura Sinica, 2021, 54(2): 248-260.
[10] KONG YaLi,ZHU ChunQuan,CAO XiaoChuang,ZHU LianFeng,JIN QianYu,HONG XiaoZhi,ZHANG JunHua. Research Progress of Soil Microbial Mechanisms in Mediating Plant Salt Resistance [J]. Scientia Agricultura Sinica, 2021, 54(10): 2073-2083.
[11] LI Hui,HAN ZhanPin,HE LiXia,YANG YaLing,YOU ShuYan,DENG Lin,WANG ChunGuo. Cloning and Functional Analysis of BraERF023a Under Salt and Drought Stresses in Cauliflower (Brassica oleracea L. var. botrytis) [J]. Scientia Agricultura Sinica, 2021, 54(1): 152-163.
[12] ShuJun MENG,XueHai ZHANG,QiYue WANG,Wen ZHANG,Li HUANG,Dong DING,JiHua TANG. Identification of miRNAs and tRFs in Response to Salt Stress in Rice Roots [J]. Scientia Agricultura Sinica, 2020, 53(4): 669-682.
[13] ZHOU Lian,XIONG YuHan,HONG XiangDe,ZHOU Jing,LIU ChaoXian,WANG JiuGuang,WANG GuoQiang,CAI YiLin. Functional Characterization of a Maize Plasma Membrane Intrinsic Protein ZmPIP2;6 Responses to Osmotic, Salt and Drought Stress [J]. Scientia Agricultura Sinica, 2020, 53(3): 461-473.
[14] SI XuYang,JIA XiaoWei,ZHANG HongYan,JIA YangYang,TIAN ShiJun,ZHANG Ke,PAN YanYun. Genomic Profiling and Expression Analysis of Phosphatidylinositol- specific PLC Gene Families Among Chinese Spring Wheat [J]. Scientia Agricultura Sinica, 2020, 53(24): 4969-4981.
[15] HAO ShuLin,CHEN HongWei,LIAO FangLi,LI Li,LIU ChangYan,LIU LiangJun,WAN ZhengHuang,SHA AiHua. Analysis of F-Box Gene Family Based on Salt-Stressed Transcriptome Sequencing in Vicia faba L. [J]. Scientia Agricultura Sinica, 2020, 53(17): 3443-3454.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
京ICP备09089781号-30
Copyright 2018 © Editorial office of Scientia Agricultura Sinica
Tel: 86-010-82106279    E-mail: zgnykx@mail.caas.net.cn
Supported by Beijing Magtech Co.Ltd