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作物学报 ›› 2021, Vol. 47 ›› Issue (3): 472-480.doi: 10.3724/SP.J.1006.2021.03027

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

质膜内在蛋白ZmPIP1;1参与玉米耐旱性和光合作用的功能分析

周练, 刘朝显, 熊雨涵, 周京, 蔡一林*()   

  1. 西南大学玉米研究所 / 农业科学研究院 / 南方山地作物逆境生物学国家级培育基地, 重庆 400715
  • 收稿日期:2020-05-14 接受日期:2020-09-13 出版日期:2021-03-12 网络出版日期:2020-10-06
  • 通讯作者: 蔡一林
  • 作者简介:E-mail: zhoulianjojo@swu.edu.cn
  • 基金资助:
    国家自然科学基金项目资助(31601312)

Functional analysis of plasma membrane intrinsic protein ZmPIP1;1 involved in drought tolerance and photosynthesis in maize

ZHOU Lian, LIU Chao-Xian, XIONG Yu-Han, ZHOU Jing, CAI Yi-Lin*()   

  1. Maize Research Institute / Academy of Agricultural Sciences / State Cultivation Base of Crop Stress Biology for Southern Mountainous Land, Southwest University, Chongqing 400715, China
  • Received:2020-05-14 Accepted:2020-09-13 Published:2021-03-12 Published online:2020-10-06
  • Contact: CAI Yi-Lin
  • Supported by:
    National Natural Science Foundation of China(31601312)

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摘要：

质膜内在蛋白(plasma membrane intrinsic proteins, PIPs)是水通道蛋白主要亚家族成员之一, 在植物生长发育过程中具有重要调节功能。前期研究表明, 玉米ZmPIP1;1基因表达受到渗透和盐胁迫的强烈诱导, 但其在玉米中的生物学功能尚不明确。本研究通过玉米遗传转化获得了ZmPIP1;1超表达转基因株系, 干旱胁迫实验揭示了ZmPIP1;1超表达转基因株系较野生型具有较低的水分散失率及较强的干旱胁迫耐性。转录组测序结果表明参与ABA生物合成及其信号通路相关基因的表达水平发生了显著变化。在田间正常生长条件下, ZmPIP1;1超表达转基因植株与野生型在生长发育过程中没有明显差异, 但转基因玉米株系具有较高的光合效率, 粒宽和百粒重增加, 玉米单果穗的产量提高。此外, 通过荧光双分子互补实验观察到ZmPIP1;1和ZmPIP2;6蛋白在玉米叶肉细胞原生质体的细胞质膜和叶绿体膜上存在互作, 并且可能导致了ZmPIP蛋白的重定位。该研究为ZmPIP1;1分子机制的解析奠定了重要基础, 为玉米高光效分子设计育种开辟了新的途径。

关键词: 玉米, ZmPIP, 干旱胁迫, 光合作用, 重定位

Abstract:

Plasma membrane intrinsic protein (PIP) is one of the main subfamily of aquaporin regulates diverse physiology functions during plant growth and development. In previous research, the expression of ZmPIP1;1 was induced by osmosis or salt stress. However, the biological function of ZmPIP1;1 was still unclear in maize. In this study, ZmPIP1;1 overexpressed transgenic plants were obtained and exhibited less water loss rate and enhanced drought tolerance compared to wild type. Transcriptome sequencing indicated significant changes in the expression levels of genes involved in ABA biosynthesis and its signaling pathways. Photosynthetic activity, kernel width and kernel weight was increased in ZmPIP1;1 overexpression maize, but not growth under normal condition compared with wild type. Moreover, interaction between ZmPIP1;1 and ZmPIP2;6 was observed by bimolecular fluorescence complementation (BiFC) experiment, resulting in re-localized on plasma membrane and chloroplast in maize mesophyll protoplast. Our study laid an important foundation for understanding the molecular mechanism of ZmPIP1;1, and provided a new method of molecular breeding for high photosynthetic efficiency.

Key words: maize, ZmPIP, drought stress, photosynthesis, re-localization

图1

ZmPIP1;1超表达转基因玉米的检测 A: ZmPIP1;1超表达载体的T-DNA序列示意图。B: ZmPIP1;1超表达T1代转基因株系的PCR检测; 1~5: 独立的转基因株系; MK: DNA marker; PC: 质粒正对照; WT: 野生型。C: 4个独立的ZmPIP1;1超表达T3代转基因株系的qRT-PCR分析。"

图2

ZmPIP1;1超表达转基因植株在干旱处理下的表型以及水分散失率 A: 野生型和ZmPIP1;1超表达转基因玉米干旱处理15 d的表型。B: 土培干旱处理后不同基因型植株与正常条件下相比的相对地上部分生物量, 数值为4个生物学重复的平均值, 误差线为标准误。C: 野生型和ZmPIP1;1超表达转基因株系叶片水分散失率的检测。*表示转基因玉米叶片的水分散失率相比野生型在P < 0.01水平差异显著。"

图3

保守序列的聚类同源组(COG)的功能分类 T和G柱分别代表“信号传导机制”和“碳水化合物运输和代谢”。"

图4

玉米ABA生物合成与信号通路的关键基因的表达分析 正常和干旱胁迫条件下野生型和ZmPIP1;1超表达株系叶中(A)参与ABA合成的ZmNCED9-1、ZmNCED9-2、ZmABA1和ZmAAO3基因; (B)参与ABA信号通路的ZmABF1、ZmPYL2、ZmABI5和ZmSnRK3基因的表达。所有数据为3个生物学重复, 误差线代表SD。*表示转基因株系与野生型的基因表达水平在P < 0.01时差异显著。ZmACT1的表达作为内参。"

图5

野生型和ZmPIP1;1超表达转基因玉米植株的AN和Ci 所有数据为平均值±标准误(n = 4); *表示转基因玉米植株的AN和Ci相比野生型在P < 0.01水平差异显著。"

图6

ZmPIP1;1超表达转基因玉米籽粒的表型鉴定 野生型和ZmPIP1;1转基因玉米籽粒表型测定(A)和照片(B)。*表示转基因玉米籽粒的十粒宽、每穗粒重和百粒重相比野生型在P < 0.01水平差异显著。标尺为1 cm。"

图7

玉米原生质体中ZmPIP1;1与ZmPIP2;6的BiFC实验观察 pCAMBIA-ZmPIP2;6-GFP、pDOE-ZmPIP1;1-ZmPIP2;6和pDOE- ZmPIP2;6-ZmPIP1;1载体与mCherry标记的质膜标记(mCherry- PM; CD3-1007)共定位。标尺为20 μm。"

[1] Yu C. China’s water crisis needs more than words. Nature, 2011,470:307.
pmid: 21331001
[2] Long S P, Zhu X G, Naidu S L, Ort D R. Can improvement in photosynthesis increase crop yields? Plant Cell Environ, 2006,29:315-330.
doi: 10.1111/j.1365-3040.2005.01493.x pmid: 17080588
[3] Nowicka B, Ciura J, Szymanska R, Kruk J. Improving photosynthesis, plant productivity and abiotic stress tolerance- current trends and future perspectives. J Plant Physiol, 2018,231:415-433.
[4] Chaumont F, Moshelion M, Daniels M J. Regulation of plant aquaporin activity. Biol Cell, 2005,97:749-764.
[5] Maurel C. Plant aquaporins: novel functions and regulation properties. FEBS Lett, 2007,581:2227-2236.
pmid: 17382935
[6] Kaldenhoff R, Ribas-Carbo M, Sans J F, Lovisolo C, Heckwolf M, Uehlein N. Aquaporins and plant water balance. Plant Cell Environ, 2008,31:658-666.
pmid: 18266903
[7] Chaumont F, Tyerman S D. Aquaporins: highly regulated channels controlling plant water relations. Plant Physiol, 2014,164:1600-1618.
pmid: 24449709
[8] Chaumont F, Barrieu F, Wojcik E, Chrispeels M J, Jung R. Aquaporins constitute a large and highly divergent protein family in maize. Plant Physiol, 2001,125:1206-1215.
[9] Johanson U, Karlsson M, Johansson I, Gustavsson S, Sjovall S, Fraysse L, Weig A R, Kjellbom P. The complete set of genes encoding major intrinsic proteins inArabidopsis provides a framework for a new nomenclature for major intrinsic proteins in plants. Plant Physiol, 2001,126:1358-1369.
doi: 10.1104/pp.126.4.1358 pmid: 11500536
[10] Ishibashi K. Aquaporin superfamily with unusual npa boxes: S-aquaporins (superfamily, sip-like and subcellular-aquaporins). Cell Mol Biol (Noisy-le-grand), 2006,52:20-27.
[11] Maurel C, Verdoucq L, Luu D T, Santoni V. Plant aquaporins: membrane channels with multiple integrated functions. Annu Rev Plant Biol, 2008,59:595-624.
pmid: 18444909
[12] Danielson J A, Johanson U. Unexpected complexity of the aquaporin gene family in the moss Physcomitrella patens. BMC Plant Biol, 2008,8:45-59.
doi: 10.1186/1471-2229-8-45 pmid: 18430224
[13] Bienert G P, Bienert M D, Jahn T P, Boutry M, Chaumont F. Solanaceae XIPs are plasma membrane aquaporins that facilitate the transport of many uncharged substrates. Plant J, 2011,66:306-317.
pmid: 21241387
[14] Kammerloher W, Fischer U, Piechottka G P, Schaffner A R. Water channels in the plant plasma membrane cloned by immunoselection from a mammalian expression system. Plant J, 1994,6:187-199.
doi: 10.1046/j.1365-313x.1994.6020187.x pmid: 7920711
[15] Murata K, Mitsuoka K, Hirai T, Walz T, Agre P, Heymann J B, Engel A, Fujiyoshi Y. Structural determinants of water permeation through aquaporin-1. Nature, 2000,407:599-605.
[16] Fetter K, Van Wilder V, Moshelion M, Chaumont F. Interactions between plasma membrane aquaporins modulate their water channel activity. Plant Cell, 2004,16:215-228.
pmid: 14671024
[17] Zelazny E, Borst J W, Muylaert M, Batoko H, Hemminga M A, Chaumont F. FRET imaging in living maize cells reveals that plasma membrane aquaporins interact to regulate their subcellular localization. Proc Natl Acad Sci USA, 2007,104:12359-12364.
[18] Vandeleur R K, Mayo G, Shelden M C, Gilliham M, Kaiser B N, Tyerman S D. The role of plasma membrane intrinsic protein aquaporins in water transport through roots: diurnal and drought stress responses reveal different strategies between isohydric and anisohydric cultivars of grapevine. Plant Physiol, 2009,149:445-460.
pmid: 18987216
[19] Chen W, Yin X, Wang L, Tian J, Yang R, Liu D, Yu Z, Ma N, Gao J. Involvement of rose aquaporin RhPIP1;1 in ethylene-regulated petal expansion through interaction with RhPIP2;1. Plant Mol Biol, 2013,83:219-233.
[20] Yaneff A, Sigaut L, Marquez M, Alleva K, Pietrasanta L I, Amodeo G. Heteromerization of PIP aquaporins affects their intrinsic permeability. Proc Natl Acad Sci USA, 2014,111:231-236.
[21] Afzal Z, Howton T C, Sun Y, Mukhtar M S. The roles of aquaporins in plant stress responses. J Dev Biol, 2016,4:9-30.
[22] Alexandersson E, Danielson J A, Rade J, Moparthi V K, Fontes M, Kjellbom P, Johanson U. Transcriptional regulation of aquaporins in accessions ofArabidopsis in response to drought stress. Plant J, 2010,61:650-660.
[23] Bae E K, Lee H, Lee J S, Noh E W. Drought, salt and wounding stress induce the expression of the plasma membrane intrinsic protein 1 gene in poplar (Populus alba × P. tremula var. glandulosa). Gene, 2011,483:43-48.
[24] Peng Y, Lin W, Cai W, Arora R. Overexpression of aPanax ginseng tonoplast aquaporin alters salt tolerance, drought tolerance and cold acclimation ability in transgenic Arabidopsis plants. Planta, 2007,226:729-740.
[25] Zhou S, Hu W, Deng X, Ma Z, Chen L, Huang C, Wang C, Wang J, He Y, Yang G, He G. Overexpression of the wheat aquaporin gene,TaAQP7, enhances drought tolerance in transgenic tobacco. PLoS One, 2012,7:e52439.
pmid: 23285044
[26] Sreedharan S, Shekhawat U K, Ganapathi T R. Transgenic banana plants overexpressing a native plasma membrane aquaporin MusaPIP1;2 display high tolerance levels to different abiotic stresses. Plant Biotechnol J, 2013,11:942-952.
[27] Zhou L, Zhou J, Xiong Y, Liu C, Wang J, Wang G, Cai Y. Overexpression of a maize plasma membrane intrinsic protein ZmPIP1;1 confers drought and salt tolerance in Arabidopsis. PLoS One, 2018,13:e0198639.
[28] Uehlein N, Otto B, Hanson D T, Fischer M, McDowell N, Kaldenhoff R. Function ofNicotiana tabacum aquaporins as chloroplast gas pores challenges the concept of membrane CO2 permeability. Plant Cell, 2008,20:648-657.
[29] Hanba Y T, Shibasaka M, Hayashi Y, Hayakawa T, Kasamo K, Terashima I, Katsuhara M. Overexpression of the barley aquaporin HvPIP2;1 increases internal CO2 conductance and CO2 assimilation in the leaves of transgenic rice plants. Plant Cell Physiol, 2004,45:521-529.
doi: 10.1093/pcp/pch070 pmid: 15169933
[30] Heckwolf M, Pater D, Hanson D T, Kaldenhoff R. TheArabidopsis thaliana aquaporin AtPIP1;2 is a physiologically relevant CO2 transport facilitator. Plant J, 2011,67:795-804.
pmid: 21564354
[31] 周练, 熊雨涵, 洪祥德, 周京, 刘朝显, 王久光, 王国强, 蔡一林. 玉米质膜内在蛋白ZmPIP2;6响应渗透、盐和干旱胁迫的功能鉴定. 中国农业科学, 53:461-473.
Zhou L, Xiong Y H, Hong X D, Zhou J, Liu C X, Wang J G, Wang G Q, Cai Y L. Functional characterization of a maize plasma membrane intrinsic protein ZmPIP2;6 responses to osmotic, salt and drought stress. Sci Agric Sin, 2020,53:461-473 (in Chinese with English abstract).
[32] Gookin T E, Assmann S M. Significant reduction of BiFC non-specific assembly facilitates in planta assessment of heterotrimeric G-protein interactors. Plant J, 2014,80:553-567.
[33] Nelson B K, Cai X, Nebenfuhr A. A multicolored set ofin vivo organelle markers for co-localization studies in Arabidopsis and other plants. Plant J, 2007,51:1126-1136.
doi: 10.1111/j.1365-313X.2007.03212.x pmid: 17666025
[34] Bart R, Chern M, Park C J, Bartley L, Ronald P C. A novel system for gene silencing using siRNAs in rice leaf and stem-derived protoplasts. Plant Methods, 2006,2:13-21.
[35] Nakashima K, Yamaguchi-Shinozaki K. ABA signaling in stress-response and seed development. Plant Cell Rep, 2013,32:959-970.
pmid: 23535869
[36] Wang Z Y, Xiong L, Li W, Zhu J K, Zhu J. The plant cuticle is required for osmotic stress regulation of abscisic acid biosynthesis and osmotic stress tolerance in Arabidopsis. Plant Cell, 2011,23:1971-1984.
doi: 10.1105/tpc.110.081943 pmid: 21610183
[37] Lee S C, Lan W, Buchanan B B, Luan S. A protein kinase-phosphatase pair interacts with an ion channel to regulate ABA signaling in plant guard cells. Proc Natl Acad Sci USA, 2009,106:21419-21424.
[38] Chater C, Peng K, Movahedi M, Dunn J A, Walker H J, Liang Y K, McLachlan D H, Casson S, Isner J C, Wilson I, Neill S J, Hedrich R, Gray J E, Hetherington A M. Elevated CO2-induced responses in stomata require ABA and ABA signaling. Curr Biol, 2015,25:2709-2716.
[39] Sorrentino G, Haworth M, Wahbi S, Mahmood T, Zuomin S, Centritto M. Abscisic acid induces rapid reductions in mesophyll conductance to carbon dioxide. PLoS One, 2016,11:e0148554.
[40] Flexas J, Ribas-Carbo M, Hanson D T, Bota J, Otto B, Cifre J, McDowell N, Medrano H, Kaldenhoff R. Tobacco aquaporin NtAQP1 is involved in mesophyll conductance to CO2 in vivo. Plant J, 2006,48:427-439.
pmid: 17010114
[41] Ding L, Gao L, Liu W, Wang M, Gu M, Ren B, Xu G, Shen Q, Guo S. Aquaporin plays an important role in mediating chloroplastic CO2 concentration under high-N supply in rice (Oryza sativa) plants. Physiol Plant, 2016,156:215-226.
[42] Heinen R B, Bienert G P, Cohen D, Chevalier A S, Uehlein N, Hachez C, Kaldenhoff R, Le Thiec D, Chaumont F. Expression and characterization of plasma membrane aquaporins in stomatal complexes of Zea mays. Plant Mol Biol, 2014,86:335-350.
doi: 10.1007/s11103-014-0232-7 pmid: 25082269
[43] Lin W, Peng Y, Li G, Arora R, Tang Z, Su W, Cai W. Isolation and functional characterization ofPgTIP1, a hormone-autotrophic cells-specific tonoplast aquaporin in ginseng. J Exp Bot, 2007,58:947-956.
pmid: 17237160
[44] Zhou L, Wang C, Liu R, Han Q, Vandeleur R K, Du J, Tyerman S D, Shou H. Constitutive overexpression of soybean plasma membrane intrinsic protein GmPIP1;6 confers salt tolerance. BMC Plant Biol, 2014,14:181-193.
pmid: 24998596
[45] Patrick J W, Zhang W, Tyerman S D, Offler C E, Walker N A. Role of membrane transport in phloem translocation of assimilates and water. Funct Plant Biol, 2001,28:697-709.
[46] Liu F, Vantoai T, Moy L P, Bock G, Linford L D, Quackenbush J. Global transcription profiling reveals comprehensive insights into hypoxic response inArabidopsis. Plant Physiol, 2005,137:1115-1129.
doi: 10.1104/pp.104.055475 pmid: 15734912
[47] Zhou Y, Setz N, Niemietz C, Qu H, Offler C E, Tyerman S D, Patrick J W. Aquaporins and unloading of phloem-imported water in coats of developing bean seeds. Plant Cell Environ, 2007,30:1566-1577.
doi: 10.1111/j.1365-3040.2007.01732.x pmid: 17927694
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