钙依赖性蛋白激酶34在小麦籽粒淀粉合成的功能
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摘要
钙依赖性蛋白激酶在调控植物发育与物质合成中发挥重要功能。为探讨一个钙依赖性蛋白激酶-TaCPK34在小麦淀粉合成中的功能,利用大麦条纹花叶病介导的基因沉默技术(barley stripe mosaic virus-virus induced gene-silencing,BSMV-VIGS)将该基因沉默,并在田间条件下测定灌浆期间沉默植株籽粒内TaCPK34基因的表达量及成熟籽粒的淀粉性状。结果表明,在花后15 d、20 d、26 d和30 d,BSMV-VIGS-TaCPK34沉默植株中TaCPK34基因表达量受到显著抑制(63.8%～97.8%); BSMV-VIGS-TaCPK34沉默植株成熟籽粒淀粉含量、千粒重、粒长和粒宽均显著降低(降幅分别为6.0%、7.1%、4.4%和11.0%);扫描电镜观察发现BSMVVIGS-TaCPK34沉默植株成熟籽粒内淀粉粒排列疏松且数量减少。这些研究结果表明TaCPK34在小麦籽粒灌浆期间对籽粒淀粉的合成有重要影响。
Calcium-dependent protein kinases play important roles in regulating growth,development and biological synthesis in higher plants. In this study,the function of a wheat calcium-dependent protein kinase,TaCPK34,in regulating the starch synthesis was identified using the barley stripe mosaic virus-virus induced gene-silencing( BSMV-VIGS) technique under field conditions. Our data indicated that expression levels of TaCPK34 gene in BSMV-TaCPK34-infected wheat plants were significantly inhibited by 63. 8% ～ 97. 8% compared to those in BSMVGFP-infected wheat plants( negative control) at 15 d,20 d,26 d and 30 d after virus inoculation. Starch content,and grain weight,grain length and width in BSMV-TaCPK34-infected wheat plants were also decreased significantly.Scanning electron microscope observation showed that morphology and number of starch granules in grains of BSMV-TaCPK34-infected wheat plants became looser and fewer compared to those of BSMV-GFP-infected wheat plants.These suggested that TaCPK34 could function in regulating starch synthesis of wheat.
Calcium-dependent protein kinases play important roles in regulating growth,development and biological synthesis in higher plants. In this study,the function of a wheat calcium-dependent protein kinase,TaCPK34,in regulating the starch synthesis was identified using the barley stripe mosaic virus-virus induced gene-silencing( BSMV-VIGS) technique under field conditions. Our data indicated that expression levels of TaCPK34 gene in BSMV-TaCPK34-infected wheat plants were significantly inhibited by 63. 8% ～ 97. 8% compared to those in BSMVGFP-infected wheat plants( negative control) at 15 d,20 d,26 d and 30 d after virus inoculation. Starch content,and grain weight,grain length and width in BSMV-TaCPK34-infected wheat plants were also decreased significantly.Scanning electron microscope observation showed that morphology and number of starch granules in grains of BSMV-TaCPK34-infected wheat plants became looser and fewer compared to those of BSMV-GFP-infected wheat plants.These suggested that TaCPK34 could function in regulating starch synthesis of wheat.
引文
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[20]Ma M,Yan Y,Huang L,et al..Virus-induced gene-silencing in wheat spikes and grains and its application in functional analysis of HMW-GS-encoding genes[J].BMC Plant Biol.,2012,12(1):141.
[21]Bennypaul H S,Mutti J S,Rustgi S,et al..Virus-induced gene silencing(VIGS)of genes expressed in root,leaf,and meiotic tissues of wheat[J].Funct.Integr.Genomics,2012,12(1):143-156.
[22]Pfeifer M,Kugler K G,Sandve S R,et al..Genome interplay in the grain transcriptome of hexaploid bread wheat[J].Science,2014,345(6194):1250091.
[23]Pont C,Salse J.Wheat paleohistory created asymmetrical genomic evolution[J].Curr.Opin.Plant Biol.,2017,36:29-37.
[24]Feldman M,Levy A A,Fahima T,et al..Genomic asymmetry in allopolyploid plants:Wheat as a model[J].J.Exp.Bot.,2012,63(14):5045-5059.
[2]Batistic O,Kudla J.Analysis of calcium signaling pathways in plants[J].Biochim.Biophys.Acta,2012,1820(8):1283-1293.
[3]畅文军,付桂,陈鑫,等.番茄钙依赖性蛋白激酶基因LeCPK2在热(光)胁迫中的功能鉴定[J].基因组学与应用生物学,2011,30(4):338-345.Chang W J,Fu G,Chen X,et al..Functional characterization of a tomato calcium-dependent protein kinase gene,Le CPK2,involved in heat(light)stress[J].Genome Appl.Biol.,2011,30(4):338-345.
[4]De Falco T A,Bender K W,Snedden W A.Breaking the code:Ca2+sensors in plant signalling[J].Biochem.J.,2009,425:27-40.
[5]Simeunovic A,Mair A,Wurzinger B,et al..Know where your clients are:Subcellular localization and targets of calciumdependent protein kinases[J].J.Exp.Bot.,2016,67(13),3855-3872.
[6]Li A L,Zhu Y F,Tan X M,et al..Evolutionary and functional study of the CDPK gene family in wheat(Triticum aestivum L.)[J].Plant Mol.Biol.,2008,66(4):429-443.
[7]Xu J,Tian Y S,Peng R H,et al..At CPK6,a functionally redundant and positive regulator involved in salt/drought stress tolerance in Arabidopsis[J].Planta,2010,231(6):1251-1260.
[8]Ma S Y,Wu W H.At CPK23 functions in Arabidopsis responses to drought and salt stresses[J].Plant Mol.Biol.,2007,65(4):511-518.
[9]Kang G Z,Xu W,Liu G Q,et al..Comprehensive analysis of the transcription of starch synthesis genes and the transcription factor RSR1 in wheat(Triticum aestivum)endosperm[J].Genome,2013,56(2):115-122.
[10]Li G Z,Wu Y F,Liu G Y,et al..Large-scale proteomics combined with transgenic experiments demonstrates an important role of jasmonic acid in potassium deficiency response in wheat and rice[J].Mol.Cell.Proteomics,2017,16(11):1889-1905.
[11]Sandberg A,Lindell G,Kllstrm B N,et al..Tumor proteomics by multivariate analysis on individual pathway data for characterization of vulvar cancer phenotypes[J].Mol.Cell Proteomics,2012,11(7):M112.016998.
[12]Ma H Z,Liu G Q,Li C W,et al..Identification of the TaBTF3 gene in wheat(Triticum aestivum L.)and the effect of its silencing on wheat chloroplast,mitochondria and mesophyll cell development[J].Biochem.Biophys.Res.Commun.,2012,426(4):608-614.
[13]Liu G Y,Wu Y F,Xu M J,et al..Virus-induced gene silencing identifies an important role of the TaRSR1transcription factor in starch synthesis in bread wheat[J].Int.J.Mol.Sci.,2016,17(10):1557.
[14]Kang G Z,Wang Y H,Liu C,et al..Difference in AGPase subunits could be associated with starch accumulation in grains between two wheat cultivars[J].Plant Growth Regul.,2010,61(1):61-66.
[15]Grimault A,Gendrot G,Chaignon S,et al..Role of B3 domain transcription factors of the AFL family in maize kernel filling[J].Plant Sci.,2015,236:116-125.
[16]Li H,Xiao Q L,Zhang C X,et al..Identification and characterization of transcription factor Zm EREB94 involved in starch synthesis in maize[J].J.Plant Physiol.,2017,216:11-16.
[17]Geng J,Li L,Lv Q,et al..TaGW2-6A allelic variation contributes to grain size possibly by regulating the expression of cytokinins and starch-related genes in wheat[J].Planta,2017,246(6):1153-1163.
[18]Mao L,Wan J,Ling H Q.Wheat functional genomics research in China:A decade of development[J].Crop J.,2018,6:1-6.
[19]叶兴国,徐惠君,杜丽璞,等.小麦规模化转基因技术体系构建及其应用[J].中国农业科学,2014,47(21):4155-4171.Ye X G,Xu H J,Du L P,et al..Establishment and application of large-scale transformation systems in wheat[J].Sci.Agric.Sin.,2014,47(21):4155-4171.
[20]Ma M,Yan Y,Huang L,et al..Virus-induced gene-silencing in wheat spikes and grains and its application in functional analysis of HMW-GS-encoding genes[J].BMC Plant Biol.,2012,12(1):141.
[21]Bennypaul H S,Mutti J S,Rustgi S,et al..Virus-induced gene silencing(VIGS)of genes expressed in root,leaf,and meiotic tissues of wheat[J].Funct.Integr.Genomics,2012,12(1):143-156.
[22]Pfeifer M,Kugler K G,Sandve S R,et al..Genome interplay in the grain transcriptome of hexaploid bread wheat[J].Science,2014,345(6194):1250091.
[23]Pont C,Salse J.Wheat paleohistory created asymmetrical genomic evolution[J].Curr.Opin.Plant Biol.,2017,36:29-37.
[24]Feldman M,Levy A A,Fahima T,et al..Genomic asymmetry in allopolyploid plants:Wheat as a model[J].J.Exp.Bot.,2012,63(14):5045-5059.