过量表达辣椒CaNPR1提高烟草对青枯菌的抗性
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摘要
对水稻和拟南芥等模式植物的研究表明,NPR1(nonexpressor of pathogenesis-related genes 1)是依赖于SA通路的防御反应调节基因,但在辣椒和烟草等茄科作物中该蛋白的功能还鲜有报道。研究从辣椒cDNA文库中分离获得一个NPR1的类似物全长cDNA(CaNPR1),并获得了其超表达的转基因烟草T1代株系。研究结果表明,这些株系与其野生型植株没有明显表型差异,但却表现出较野生型植株更高的抗青枯菌侵染活性。同时,研究还发现CaNPR1的超表达还显著提高了防御相关基因的表达,表明NPR1在不同植物间具有较强的功能保守性。
The NPR1(nonexpressor of pathogenesis-related genes 1) gene plays an important role in SA-depended defense response processes in model plants such as Arabidopsis and rice.However,the function and underlying mechanism of this gene in non-model plants,including pepper(Capsicum annuum) and other Solanaceae,remain poorly understood.Here,we report on the functional characterization of CaNPR1 from pepper.Our data showed that the full-length cDNA of CaNPR1 was isolated and overexpression of CaNPR1 was construed in tobacco K326.Compared to wild type K326,CaNPR1-OE lines did not exhibit any phenotypic differences,but exhibited higher resistance to Ralstonia solanacearum inoculation.In addition,this was coupled with enhanced transcript levels of defense-related marker genes,suggesting that CaNPR1 possessed higher conservative function in cross species.
The NPR1(nonexpressor of pathogenesis-related genes 1) gene plays an important role in SA-depended defense response processes in model plants such as Arabidopsis and rice.However,the function and underlying mechanism of this gene in non-model plants,including pepper(Capsicum annuum) and other Solanaceae,remain poorly understood.Here,we report on the functional characterization of CaNPR1 from pepper.Our data showed that the full-length cDNA of CaNPR1 was isolated and overexpression of CaNPR1 was construed in tobacco K326.Compared to wild type K326,CaNPR1-OE lines did not exhibit any phenotypic differences,but exhibited higher resistance to Ralstonia solanacearum inoculation.In addition,this was coupled with enhanced transcript levels of defense-related marker genes,suggesting that CaNPR1 possessed higher conservative function in cross species.
引文
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[15]Kinkema M,Fan W,Dong X.Nuclear localization ofNPR1 is required for activation of PR gene expres-sion[J].Plant Cell,2000,12(12):2339-2350.
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[20]Oh S K,Park J M,Joung Y H,Lee S,Chung E,KimS Y,Yu S H,Choi D.A plant EPF-type zinc-fingerprotein,CaPIF1,involved in defence against patho-gens[J].Mol Plant Pathol,2005,6(3):269-285.
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[23]Chen N,Goodwin P,Hsiang T.The role of ethyleneduring the infection of Nicotiana tabacum by Colle-totrichum destructivum[J].J Exp Bot,2003,54(392):2449.
[24]Sohn S I,Kim Y H,Kim B R,Lee S Y,Lim C K,HurJ H,Lee J Y.Transgenic tobacco expressing thehrpNEP gene from Erwinia pyrifoliae triggers de-fense responses against Botrytis cinerea[J].MolCells,2007,24(2):232-239.
[25]Takahashi H,Chen Z,Du H,Liu Y,Klessig D F.De-velopment of necrosis and activation of disease re-sistance in transgenic tobacco plants with severelyreduced catalase levels[J].Plant J,1997,11(5):993-1005.
[26]Takahashi Y,Nagata T.parB:an auxin-regulatedgene encoding glutathione S-transferase[J].ProcNatl Acad Sci USA,1992,89(1):56-59.
[27]Cao H,Bowling S A,Gordon a S,Dong X.Charac-terization of an Arabidopsis mutant that is nonre-sponsive to inducers of systemic acquired resis-tance[J].Plant Cell,1994,6(11):1583-1592.
[28]Cao H,Glazebrook J,Clarke J D,Volko S,Dong X.The Arabidopsis NPR1 gene that controls systemicacquired resistance encodes a novel protein con-taining ankyrin repeats[J].Cell,1997,88(1):57-63.
[29]Spoel S H,Koornneef A,Claessens S M C,Korzeli-us J P,Van Pelt J A,Mueller M J,Buchala A J,Métraux J P,Brown R,Kazan K,et al.NPR1 modu-lates cross-talk between salicylate-and jasmonate-dependent defense pathways through a novelfunction in the cytosol[J].Plant Cell,2003,15(3):760-770.
[30]Després C,Delong C,Glaze S,Liu E,Fobert P R.The Arabidopsis NPR1/NIM1 protein enhances theDNA binding activity of a subgroup of the TGAfamily of bZIP transcription factors[J].Plant Cell,2000,12(2):279-290.
[31]Nandi A,Kachroo P,Fukushige H,Hildebrand D F,Klessig D F,Shah J.Ethylene and jasmonic acidsignaling affect the NPR1-independent expressionof defense genes without impacting resistance toPseudomonas syringae and Peronospora parasiti-ca in the Arabidopsis ssi1 mutant[J].Mol PlantMicrobe Interact,2003,16(7):588-599.
[32]Chern M,Canlas P E,Fitzgerald H A,Ronald P C.Rice NRR,a negative regulator of disease resis-tance,interacts with Arabidopsis NPR1 and riceNH1[J].Plant J,2005,43(5):623-635.
[2]Ma W,Smigel A,Tsai Y C,Braam J,Berkowitz GA.Innate immunity signaling:cytosolic Ca2+eleva-tion is linked to downstream nitric oxide generationthrough the action of calmodulin or a calmodulin-like protein[J].Plant Physiol,2008,148(2):818-828.
[3]Makandar R,Nalam V,Chaturvedi R,Jeannotte R,Sparks A A,Shah J.Involvement of salicylate andjasmonate signaling pathways in Arabidopsis inter-action with Fusarium graminearum[J].Mol PlantMicrobe Interact,2010,23(7):861-870.
[4]Han L,Li G J,Yang K Y,Mao G,Wang R,Liu Y,Zhang S.Mitogen-activated protein kinase 3 and 6regulate Botrytis cinerea-induced ethylene produc-tion in Arabidopsis[J].Plant J,2010,64(1):114-127.
[5]Larkindale J,Knight M R.Protection against heatstress-induced oxidative damage in Arabidopsisinvolves calcium,abscisic acid,ethylene,and sali-cylic acid[J].Plant Physiol,2002,128(2):682-695.
[6]Liu H T,Liu Y Y,Pan Q H,Yang H R,Zhan J C,Huang W D.Novel interrelationship between sali-cylic acid,abscisic acid,and PIP2-specific phos-pholipase C in heat acclimation-induced thermoto-lerance in pea leaves[J].J Exp Bot,2006,57(12):3337-3347.
[7]Nakashita H,Yasuda M,Nitta T,Asami T,FujiokaS,Arai Y,Sekimata K,Takatsuto S,Yamaguchi I,Yoshida S.Brassinosteroid functions in a broadrange of disease resistance in tobacco and rice[J].Plant J,2003,33(5):887-898.
[8]Jonak C,Okresz L,Bogre L,Hirt H.Complexity,cross talk and integration of plant MAP kinase sig-nalling[J].Curr Opin Plant Biol,2002,5(5):415-424.
[9]Rushton P J,Somssich I E,Ringler P,Shen Q J.WRKY transcription factors[J].Trends Plant Sci,2010,15(5):247-258.
[10]Wu Y,Deng Z,Lai J,Zhang Y,Yang C,Yin B,ZhaoQ,Zhang L,Li Y.Dual function of Arabidopsis AT-AF1 in abiotic and biotic stress responses[J].CellRes,2009,19(11):1279-1290.
[11]Xia N,Zhang G,Liu X Y,Deng L,Cai G L,ZhangY,Wang X J,Zhao J,Huang L L,Kang Z S.Characterization of a novel wheat NAC transcrip-tion factor gene involved in defense response a-gainst stripe rust pathogen infection and abioticstresses[J].Mol Biol Rep,2010,37(8):3703-3712.
[12]Tsutsui T,Kato W,Asada Y,Sako K,Sato T,Sono-da Y,Kidokoro S,Yamaguchi-Shinozaki K,TamaokiM,Arakawa K.DEAR1,a transcriptional repressorof DREB protein that mediates plant defense andfreezing stress responses in Arabidopsis[J].JPlant Res,2009,122(6):633-643.
[13]On珘ate-Sánchez L,Anderson J P,Young J,Singh KB.AtERF14,a member of the ERF family of tran-scription factors,plays a nonredundant role in plantdefense[J].Plant Physiol,2007,143(1):400-409.
[14]Mou Z,Fan W,Dong X.Inducers of plant systemicacquired resistance regulate NPR1 functionthrough redox changes[J].Cell,2003,113(7):935-944.
[15]Kinkema M,Fan W,Dong X.Nuclear localization ofNPR1 is required for activation of PR gene expres-sion[J].Plant Cell,2000,12(12):2339-2350.
[16]Ristaino J B,Johnston S A.Ecologically based ap-proaches to management of Phytophthora blighton bell pepper[J].Plant Dis,1999,83(12):1080-1089.
[17]Hwang B K,Kim C H.Phytophthora blight of pep-per and its control in Korea[J].Plant Dis,1995,79(3):221-227.
[18]Kelman A.The relationship of pathogenicity inPseudomonas solanacearum to colony appear-ance on a tetrazolium medium[J].Phytopatholo-gy,1954,44(12):693-695.
[19]Jung J,Won S Y,Suh S C,Kim H R,Wing R,JeongY,Hwang I,Kim M.The barley ERF-type transcrip-tion factor HvRAF confers enhanced pathogen re-sistance and salt tolerance in Arabidopsis[J].Planta,2007,225(3):575-588.
[20]Oh S K,Park J M,Joung Y H,Lee S,Chung E,KimS Y,Yu S H,Choi D.A plant EPF-type zinc-fingerprotein,CaPIF1,involved in defence against patho-gens[J].Mol Plant Pathol,2005,6(3):269-285.
[21]Livak K J,Schmittgen T D.Analysis of relativegene expression data using real-time quantitativePCR and the 2(-Delta Delta C(T))method[J].Methods,2001,25(4):402-408.
[22]Bae H,Roberts D P,Lim H S,Strem M D,Park SC,Ryu C M,Melnick R L,Bailey B A.EndophyticTrichoderma isolates from tropical environmentsdelay disease onset and induce resistance againstPhytophthora capsici in hot pepper using multiplemechanisms[J].Mol Plant Microbe Interact,2011,24(3):336-351.
[23]Chen N,Goodwin P,Hsiang T.The role of ethyleneduring the infection of Nicotiana tabacum by Colle-totrichum destructivum[J].J Exp Bot,2003,54(392):2449.
[24]Sohn S I,Kim Y H,Kim B R,Lee S Y,Lim C K,HurJ H,Lee J Y.Transgenic tobacco expressing thehrpNEP gene from Erwinia pyrifoliae triggers de-fense responses against Botrytis cinerea[J].MolCells,2007,24(2):232-239.
[25]Takahashi H,Chen Z,Du H,Liu Y,Klessig D F.De-velopment of necrosis and activation of disease re-sistance in transgenic tobacco plants with severelyreduced catalase levels[J].Plant J,1997,11(5):993-1005.
[26]Takahashi Y,Nagata T.parB:an auxin-regulatedgene encoding glutathione S-transferase[J].ProcNatl Acad Sci USA,1992,89(1):56-59.
[27]Cao H,Bowling S A,Gordon a S,Dong X.Charac-terization of an Arabidopsis mutant that is nonre-sponsive to inducers of systemic acquired resis-tance[J].Plant Cell,1994,6(11):1583-1592.
[28]Cao H,Glazebrook J,Clarke J D,Volko S,Dong X.The Arabidopsis NPR1 gene that controls systemicacquired resistance encodes a novel protein con-taining ankyrin repeats[J].Cell,1997,88(1):57-63.
[29]Spoel S H,Koornneef A,Claessens S M C,Korzeli-us J P,Van Pelt J A,Mueller M J,Buchala A J,Métraux J P,Brown R,Kazan K,et al.NPR1 modu-lates cross-talk between salicylate-and jasmonate-dependent defense pathways through a novelfunction in the cytosol[J].Plant Cell,2003,15(3):760-770.
[30]Després C,Delong C,Glaze S,Liu E,Fobert P R.The Arabidopsis NPR1/NIM1 protein enhances theDNA binding activity of a subgroup of the TGAfamily of bZIP transcription factors[J].Plant Cell,2000,12(2):279-290.
[31]Nandi A,Kachroo P,Fukushige H,Hildebrand D F,Klessig D F,Shah J.Ethylene and jasmonic acidsignaling affect the NPR1-independent expressionof defense genes without impacting resistance toPseudomonas syringae and Peronospora parasiti-ca in the Arabidopsis ssi1 mutant[J].Mol PlantMicrobe Interact,2003,16(7):588-599.
[32]Chern M,Canlas P E,Fitzgerald H A,Ronald P C.Rice NRR,a negative regulator of disease resis-tance,interacts with Arabidopsis NPR1 and riceNH1[J].Plant J,2005,43(5):623-635.