类受体蛋白激酶NtFERL对烟草质体色素含量的影响
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摘要
为明确烟草NtFERL(FER-like gene)的生物学功能,同源克隆NtFERL基因并进行序列分析,同时采用病毒诱导的基因沉默(Virus induced gene silence, VIGS)方法在本氏烟草中对该基因进行功能分析。结果表明:NtFERL基因位于烟草第7号染色体上,无内含子,编码序列长2 670 bp,编码889个氨基酸。NtFERL与拟南芥FERONIA基因同源性较高,在核苷酸和氨基酸水平上的一致性分别为68.8%和71.1%。沉默NtFERL后,烟草叶色泛黄。质体色素含量分析显示,除新黄质外,叶绿素a、叶绿素b、β-胡萝卜素、叶黄素均显著降低,表明NtFERL可能对烟草色素含量起着正调控的作用。
To demonstrate the biological roles of FER-like(FERL) gene in Nicotiana tabacum, NtFERL was cloned with the homologous cloning strategy and its sequence was analyzed. The function of NtFERL was characterized by virus-induced gene silence(VIGS) in N. benthamiana. The results showed that NtFERL located in chromosome 7 was found to lack intron. The CDS sequence of NtFERL was 2670 bp in length and encoded889 amino acid residues. Sequence alignment showed that at nucleotide acid and amino acid levels, NtFERL shared 68.8% and 71.1% identities with FERONIA in Arabidopsis thaliana, respectively. NtFERL-silenced tobacco plants exhibited light yellow leaves. Except for neoxanthin, the contents of chlorophyll a, chlorophyll b,β-carotene and xanthophyll were significantly reduced, which suggested that NtFERL might positively regulate the plastid pigment contents in tobacco.
To demonstrate the biological roles of FER-like(FERL) gene in Nicotiana tabacum, NtFERL was cloned with the homologous cloning strategy and its sequence was analyzed. The function of NtFERL was characterized by virus-induced gene silence(VIGS) in N. benthamiana. The results showed that NtFERL located in chromosome 7 was found to lack intron. The CDS sequence of NtFERL was 2670 bp in length and encoded889 amino acid residues. Sequence alignment showed that at nucleotide acid and amino acid levels, NtFERL shared 68.8% and 71.1% identities with FERONIA in Arabidopsis thaliana, respectively. NtFERL-silenced tobacco plants exhibited light yellow leaves. Except for neoxanthin, the contents of chlorophyll a, chlorophyll b,β-carotene and xanthophyll were significantly reduced, which suggested that NtFERL might positively regulate the plastid pigment contents in tobacco.
引文
[1]王瑞新.烟草化学[M].北京:中国农业出版社,2003:84-85.WANG Ruixin.Tobacco chemistry[M].Beijing:China Agriculture Press,2003:84-85.
[2]李亚飞,常栋,孙军伟,等.烤烟和白肋烟烟苗氮代谢的差异分析[J].烟草科技,2017,50(1):6-11.LI Yafei,CHANG Dong,SUN Junwei,et al.Difference of nitrogen metabolism between flue-cured tobacco and burley tobacco seedlings[J].Tobacco Science&Technology,2017,50(1):6-11.
[3]金闻博,戴亚,横田拓,等.烟草化学[M].北京:清华大学出版社,2000:45-46.JIN Wenbo,DAI Ya,HENG Tiantuo,et al.Tobacco chemistry[M].Beijing:Tsinghua University Press,2000:45-46.
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[5]韦凤杰,范艺宽,刘国顺,等.饼肥对烤烟叶片发育过程中质体色素降解及相关酶类活性的影响[J].作物学报,2006,32(5):766-771.WEI Fengjie,FAN Yikuan,LIU Guoshun,et al.Effect of cake fertilizer on the pigments degradation and related enzyme activities in flue-cured tobacco[J].Acta Agronomica Sinica,2006,32(5):766-771.
[6]韩锦峰,吕巧灵,杨素勤,等.饼肥种类及其与化肥配比对烤烟生长发育及产质的影响Ⅰ.对烤烟生长发育的影响[J].河南农业科学,1998(2):14-16.HAN Jinfeng,LüQiaoling,YANG Suqin,et al.Effects of cake fertilizer type and its combination with chemical fertilizers on growth,yield and quality of fluecured tobacco I.Effects of cake fertilizer on growth and development of flue-cured tobacco[J].Journal of Henan Agricultural Sciences,1998(2):14-16.
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[8]赵铭钦,刘金霞,刘国顺,等.增施不同有机物质对烤烟质体色素及其降解产物的影响[J].浙江农业科学,2008(2):243-246.ZHAO Mingqin,LIU Jinxia,LIU Guoshun,et al.Effect of different organic substance on plastid pigment and its degraded products in flue-cured tobacco[J].Journal of Zhejiang Agricultural Sciences,2008(2):243-246.
[9]赵铭钦,张迪,赵进恒,等.种植密度对烤烟质体色素及其降解产物的影响[J].江苏农业学报,2010,26(4):711-715.ZHAO Mingqin,ZHANG Di,ZHAO Jinheng,et al.Effect of planting density on chromoplast pigment and its degradation products in flue-cured tobacco leaf[J].Jiangsu Journal of Agricultural Sciences,2010,26(4):711-715.
[10]Lindgren L O,St?lberg K G,H?glund A S.Seedspecific overexpression of an endogenous Arabidopsis phytoene synthase gene results in delayed germination and increased levels of carotenoids,chlorophyll,and abscisic acid[J].Plant Physiology,2003,132(2):779-785.
[11]Fraser P D,Romer S,Shipton C A,et al.Evaluation of transgenic tomato plants expressing an additional phytoene synthase in a fruit-specific manner[J].Proceedings of the National Academy of Sciences of the United States of America,2002,99(2):1092-1097.
[12]Shewmaker C K,Sheehy J A,Daley M,et al.Seedspecific overexpression of phytoene synthase:increase in carotenoids and other metabolic effects[J].The Plant Journal,1999,20(4):401-412.
[13]Tian L,Musetti V,Kim J,et al.The Arabidopsis LUT1locus encodes a member of the cytochrome P450 family that is required for carotenoidε-ring hydroxylation activity[J].Proceedings of the National Academy of Sciences of the United States of America,2004,101(1):402-407.
[14]Cunningham F X,Pogson B,Sun Z,et al.Functional analysis of the beta and epsilon lycopene cyclase enzymes of Arabidopsis reveals a mechanism for control of cyclic carotenoid formation[J].The Plant Cell,1996,8(9):1613-1626.
[15]Pogson B,McDonald K A,Truong M,et al.Arabidopsis carotenoid mutants demonstrate that lutein is not essential for photosynthesis in higher plants[J].The Plant Cell,1996,8(9):1627-1639.
[16]Yu F,Qian L C,Nibau C,et al.FERONIA receptor kinase pathway suppresses abscisic acid signaling in Arabidopsis by activating ABI2 phosphatase[J].Proceedings of the National Academy of Sciences of the United States of America,2012,109(36):14693-14698.
[17]Yu F,Li J,Huang Y,et al.FERONIA receptor kinase controls seed size in Arabidopsis thaliana[J].Molecular Plant,2014,7(5):920-922.
[18]Mao D D,Yu F,Li J,et al.FERONIA receptor kinase interacts with S-adenosylmethionine synthetase and suppresses S-adenosylmethionine production and ethylene biosynthesis in Arabidopsis[J].Plant,Cell&Environment,2015,38(12):2566-2574.
[19]Shih H W,Miller N D,Dai C,et al.The receptor-like kinase FERONIA is required for mechanical signal transduction in Arabidopsis seedlings[J].Current Biology,2014,24(16):1887-1892.
[20]Duan Q H,Kita D,Li C,et al.FERONIA receptor-like kinase regulates RHO GTPase signaling of root hair development[J].Proceedings of the National Academy of Sciences of the United States of America,2010,107(41):17821-17826.
[21]Chen J,Yu F,Liu Y,et al.FERONIA interacts with ABI2-type phosphatases to facilitate signaling cross-talk between abscisic acid and RALF peptide in Arabidopsis[J].Proceedings of the National Academy of Sciences of the United States of America,2016,113(37):E5519-E5527.
[22]Kessler S A,Shimosato-Asano H,Keinath N F,et al.Conserved molecular components for pollen tube reception and fungal invasion[J].Science,2010,330(6006):968-971.
[23]Escobar-Restrepo J M,Huck N,Kessler S,e al.The FERONIA receptor-like kinase mediates male-female interactions during pollen tube reception[J].Science,2007,317(5838):656-660.
[24]Kanaoka M M,Torii K U.FERONIA as an upstream receptor kinase for polar cell growth in plants[J].Proceedings of the National Academy of Sciences of the United States of America,2010,107(41):17461-17462.
[25]Li C,Yeh F L,Cheung A Y,et al.Glycosylphosphatidylinositol-anchored proteins as chaperones and co-receptors for FERONIA receptor kinase signaling in Arabidopsis[J/OL].eLife,2015,4:e06587(2015-06-08)[2018-07-26].https://cdn.elifesciences.org/articles/06587/elife-06587-v1.pdf.DOI:10.7554/eLife.06587.
[26]Kessler S A,Lindner H,Jones D S,et al.Functional analysis of related CrRLK1L receptor-like kinases in pollen tube reception[J].EMBO Reports,2015,16(1):107-115.
[27]Schmittgen T D,Livak K J.Analyzing real-time PCRdata by the comparative CTmethod[J].Nature Protocols,2008,3(6):1101-1108.
[28]YC/T 382-2010烟草及烟草制品质体色素的测定高效液相色谱法[S].YC/T 382-2010 Tobacco and tobacco productsDetermination of plastid pigments-High performance liquid chromatography method[S].
[29]Schallus T,Jaeckh C,Feher K,et al.Malectin:a novel carbohydrate-binding protein of the endoplasmic reticulum and a candidate player in the early steps of protein N-glycosylation[J].Molecular Biology of the Cell,2008,19(8):3404-3414.
[30]Shiu S H,Bleecker A B.Receptor-like kinases from Arabidopsis form a monophyletic gene family related to animal receptor kinases[J].Proceedings of the National Academy of Sciences of the United States of America,2001,98(19):10763-10768.
[31]Park H,Kreunen S S,Cuttriss A J,et al.Identification of the carotenoid isomerase provides insight into carotenoid biosynthesis,prolamellar body formation,and photomorphogenesis[J].The Plant Cell,2002,14(2):321-332.
[2]李亚飞,常栋,孙军伟,等.烤烟和白肋烟烟苗氮代谢的差异分析[J].烟草科技,2017,50(1):6-11.LI Yafei,CHANG Dong,SUN Junwei,et al.Difference of nitrogen metabolism between flue-cured tobacco and burley tobacco seedlings[J].Tobacco Science&Technology,2017,50(1):6-11.
[3]金闻博,戴亚,横田拓,等.烟草化学[M].北京:清华大学出版社,2000:45-46.JIN Wenbo,DAI Ya,HENG Tiantuo,et al.Tobacco chemistry[M].Beijing:Tsinghua University Press,2000:45-46.
[4]Tso T C.Physiology and biochemistry of tobacco plants[M].Stroudsburg,PA:Dowden,Hutchinson&Ross Inc.,1972:209-217.
[5]韦凤杰,范艺宽,刘国顺,等.饼肥对烤烟叶片发育过程中质体色素降解及相关酶类活性的影响[J].作物学报,2006,32(5):766-771.WEI Fengjie,FAN Yikuan,LIU Guoshun,et al.Effect of cake fertilizer on the pigments degradation and related enzyme activities in flue-cured tobacco[J].Acta Agronomica Sinica,2006,32(5):766-771.
[6]韩锦峰,吕巧灵,杨素勤,等.饼肥种类及其与化肥配比对烤烟生长发育及产质的影响Ⅰ.对烤烟生长发育的影响[J].河南农业科学,1998(2):14-16.HAN Jinfeng,LüQiaoling,YANG Suqin,et al.Effects of cake fertilizer type and its combination with chemical fertilizers on growth,yield and quality of fluecured tobacco I.Effects of cake fertilizer on growth and development of flue-cured tobacco[J].Journal of Henan Agricultural Sciences,1998(2):14-16.
[7]韩锦峰,朱大恒,林学梧,等.多胺对烟株生长发育及烤烟产质的生理效应[J].中国烟草,1992(1):1-7.HAN Jinfeng,ZHU Daheng,LIN Xuewu,et al.Physiological effects of polyamine on the growth and development of tobacco,the quality and yield of the leaves[J].China Tobacco,1992(1):1-7.
[8]赵铭钦,刘金霞,刘国顺,等.增施不同有机物质对烤烟质体色素及其降解产物的影响[J].浙江农业科学,2008(2):243-246.ZHAO Mingqin,LIU Jinxia,LIU Guoshun,et al.Effect of different organic substance on plastid pigment and its degraded products in flue-cured tobacco[J].Journal of Zhejiang Agricultural Sciences,2008(2):243-246.
[9]赵铭钦,张迪,赵进恒,等.种植密度对烤烟质体色素及其降解产物的影响[J].江苏农业学报,2010,26(4):711-715.ZHAO Mingqin,ZHANG Di,ZHAO Jinheng,et al.Effect of planting density on chromoplast pigment and its degradation products in flue-cured tobacco leaf[J].Jiangsu Journal of Agricultural Sciences,2010,26(4):711-715.
[10]Lindgren L O,St?lberg K G,H?glund A S.Seedspecific overexpression of an endogenous Arabidopsis phytoene synthase gene results in delayed germination and increased levels of carotenoids,chlorophyll,and abscisic acid[J].Plant Physiology,2003,132(2):779-785.
[11]Fraser P D,Romer S,Shipton C A,et al.Evaluation of transgenic tomato plants expressing an additional phytoene synthase in a fruit-specific manner[J].Proceedings of the National Academy of Sciences of the United States of America,2002,99(2):1092-1097.
[12]Shewmaker C K,Sheehy J A,Daley M,et al.Seedspecific overexpression of phytoene synthase:increase in carotenoids and other metabolic effects[J].The Plant Journal,1999,20(4):401-412.
[13]Tian L,Musetti V,Kim J,et al.The Arabidopsis LUT1locus encodes a member of the cytochrome P450 family that is required for carotenoidε-ring hydroxylation activity[J].Proceedings of the National Academy of Sciences of the United States of America,2004,101(1):402-407.
[14]Cunningham F X,Pogson B,Sun Z,et al.Functional analysis of the beta and epsilon lycopene cyclase enzymes of Arabidopsis reveals a mechanism for control of cyclic carotenoid formation[J].The Plant Cell,1996,8(9):1613-1626.
[15]Pogson B,McDonald K A,Truong M,et al.Arabidopsis carotenoid mutants demonstrate that lutein is not essential for photosynthesis in higher plants[J].The Plant Cell,1996,8(9):1627-1639.
[16]Yu F,Qian L C,Nibau C,et al.FERONIA receptor kinase pathway suppresses abscisic acid signaling in Arabidopsis by activating ABI2 phosphatase[J].Proceedings of the National Academy of Sciences of the United States of America,2012,109(36):14693-14698.
[17]Yu F,Li J,Huang Y,et al.FERONIA receptor kinase controls seed size in Arabidopsis thaliana[J].Molecular Plant,2014,7(5):920-922.
[18]Mao D D,Yu F,Li J,et al.FERONIA receptor kinase interacts with S-adenosylmethionine synthetase and suppresses S-adenosylmethionine production and ethylene biosynthesis in Arabidopsis[J].Plant,Cell&Environment,2015,38(12):2566-2574.
[19]Shih H W,Miller N D,Dai C,et al.The receptor-like kinase FERONIA is required for mechanical signal transduction in Arabidopsis seedlings[J].Current Biology,2014,24(16):1887-1892.
[20]Duan Q H,Kita D,Li C,et al.FERONIA receptor-like kinase regulates RHO GTPase signaling of root hair development[J].Proceedings of the National Academy of Sciences of the United States of America,2010,107(41):17821-17826.
[21]Chen J,Yu F,Liu Y,et al.FERONIA interacts with ABI2-type phosphatases to facilitate signaling cross-talk between abscisic acid and RALF peptide in Arabidopsis[J].Proceedings of the National Academy of Sciences of the United States of America,2016,113(37):E5519-E5527.
[22]Kessler S A,Shimosato-Asano H,Keinath N F,et al.Conserved molecular components for pollen tube reception and fungal invasion[J].Science,2010,330(6006):968-971.
[23]Escobar-Restrepo J M,Huck N,Kessler S,e al.The FERONIA receptor-like kinase mediates male-female interactions during pollen tube reception[J].Science,2007,317(5838):656-660.
[24]Kanaoka M M,Torii K U.FERONIA as an upstream receptor kinase for polar cell growth in plants[J].Proceedings of the National Academy of Sciences of the United States of America,2010,107(41):17461-17462.
[25]Li C,Yeh F L,Cheung A Y,et al.Glycosylphosphatidylinositol-anchored proteins as chaperones and co-receptors for FERONIA receptor kinase signaling in Arabidopsis[J/OL].eLife,2015,4:e06587(2015-06-08)[2018-07-26].https://cdn.elifesciences.org/articles/06587/elife-06587-v1.pdf.DOI:10.7554/eLife.06587.
[26]Kessler S A,Lindner H,Jones D S,et al.Functional analysis of related CrRLK1L receptor-like kinases in pollen tube reception[J].EMBO Reports,2015,16(1):107-115.
[27]Schmittgen T D,Livak K J.Analyzing real-time PCRdata by the comparative CTmethod[J].Nature Protocols,2008,3(6):1101-1108.
[28]YC/T 382-2010烟草及烟草制品质体色素的测定高效液相色谱法[S].YC/T 382-2010 Tobacco and tobacco productsDetermination of plastid pigments-High performance liquid chromatography method[S].
[29]Schallus T,Jaeckh C,Feher K,et al.Malectin:a novel carbohydrate-binding protein of the endoplasmic reticulum and a candidate player in the early steps of protein N-glycosylation[J].Molecular Biology of the Cell,2008,19(8):3404-3414.
[30]Shiu S H,Bleecker A B.Receptor-like kinases from Arabidopsis form a monophyletic gene family related to animal receptor kinases[J].Proceedings of the National Academy of Sciences of the United States of America,2001,98(19):10763-10768.
[31]Park H,Kreunen S S,Cuttriss A J,et al.Identification of the carotenoid isomerase provides insight into carotenoid biosynthesis,prolamellar body formation,and photomorphogenesis[J].The Plant Cell,2002,14(2):321-332.