芝麻赤霉素合成相关基因鉴定与表达分析
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摘要
赤霉素的生物合成是多种酶促反应的过程,与植株发育、逆境响应密切相关。为分析芝麻基因组中赤霉素合成相关基因的分布、结构和活性特征,本研究以芝麻基因组序列和不同组织转录组为对象,通过生物信息学方法,从中共鉴定出32个赤霉素合成相关基因,分属7个不同的基因家族,分布在除5号染色体外芝麻所有的染色体上。不同基因家族蛋白质肽链长度存在较大差异,其中CPS家族的蛋白质序列最长,平均包含776个氨基酸残基;KAO家族蛋白序列长度的变异最大,在401～834个氨基酸之间。所有芝麻赤霉素合成相关基因编码的蛋白质预测为亲水性蛋白。与拟南芥、水稻、大豆、葡萄、高粱、番茄等物种相比,芝麻中赤霉素合成相关基因数目处于中等水平,但CPS家族中芝麻的基因数目明显多于其他物种,进化分析表明,在分析的7个物种中,芝麻与番茄具有较近的进化关系。在不同组织中,7个相关基因家族表达模式不同。KS家族基因在种子、茎尖和叶片中表达相对较高; CPS家族基因在心皮和种子中活性较强,但在茎尖和叶片中几乎没有表达; KAO和KO家族基因在不同组织中整体上具有相对较高的活性,而KS、GA3ox家族的基因在不同组织中整体表达量较低。就不同组织而言,种子中的GA合成相关基因活性整体较高;而在根尖、茎尖、叶片等与芝麻植株形态建成密切相关的3个组织中,这些基因表达以根尖较突出、其次为茎尖,在叶片中的活性整体较低。本研究为解析芝麻赤霉素合成相关基因的结构特征及其在不同组织中的作用特点提供了基因信息和理论参考。
The biosynthesis of gibberellin(GA) is a process of various enzymatic reactions and is closely related to plant development and stress response.To explore the distribution,structure and activity of sesame GA related genes,sesame genome sequence and transcriptomes from different tissues were studied based on bioinformatics methods.It showed that sesame had 32 GA related genes belonging to 7 different gene families,and these genes were distributed on all chromosomes except the chromosome 5.The protein sequence lengths of the different gene families were quite different.The CPS genes had the longest protein and contained 776 amino acid residues on average.The KAO proteins exhibited a great variation in size,varying from 401 to 834 amino acids.All the proteins encoded by GA related genes were predicted to be hydrophilic proteins.Compared with other species inclusing Arabidopsis,rice,soybean,grape,sorghum and tomato,the total number of GA related genes in sesame is not outstanding,but more CPS genes were observed in sesame.Evolution analysis indicated that sesame and tomato had a closer evolution relationship among the 7 species.In various sesame tissues,the 7 GA related gene families displayed quite different expression patterns.KS genes were highly expressed in seeds,stem tips and leaves.CPS genes were more active in pericarps and seeds,but were hardly expressed in stem tips and leaves.KAO and KO genes were constitutively and higly expressed in all tissues,while KS and GA3 ox genes showed low levels of expression.Among the different tissues,the GA related genes generally showed higher activity in seed,then in root tip and in stem tip.This study provided important genetic information and theoretical references to analyze the characteristics of GA related genes in sesame and its role in different tissues.
The biosynthesis of gibberellin(GA) is a process of various enzymatic reactions and is closely related to plant development and stress response.To explore the distribution,structure and activity of sesame GA related genes,sesame genome sequence and transcriptomes from different tissues were studied based on bioinformatics methods.It showed that sesame had 32 GA related genes belonging to 7 different gene families,and these genes were distributed on all chromosomes except the chromosome 5.The protein sequence lengths of the different gene families were quite different.The CPS genes had the longest protein and contained 776 amino acid residues on average.The KAO proteins exhibited a great variation in size,varying from 401 to 834 amino acids.All the proteins encoded by GA related genes were predicted to be hydrophilic proteins.Compared with other species inclusing Arabidopsis,rice,soybean,grape,sorghum and tomato,the total number of GA related genes in sesame is not outstanding,but more CPS genes were observed in sesame.Evolution analysis indicated that sesame and tomato had a closer evolution relationship among the 7 species.In various sesame tissues,the 7 GA related gene families displayed quite different expression patterns.KS genes were highly expressed in seeds,stem tips and leaves.CPS genes were more active in pericarps and seeds,but were hardly expressed in stem tips and leaves.KAO and KO genes were constitutively and higly expressed in all tissues,while KS and GA3 ox genes showed low levels of expression.Among the different tissues,the GA related genes generally showed higher activity in seed,then in root tip and in stem tip.This study provided important genetic information and theoretical references to analyze the characteristics of GA related genes in sesame and its role in different tissues.
引文
[1]张秀荣,李培武,汪雪芳,等.芝麻种子木质素组分、粗脂肪、粗蛋白含量及相关性分析[J].中国油料作物学报,2005,27(3):88-90.
[2]王蕾,黎冬华,齐小琼,等.芝麻核心种质芝麻素和芝麻酚林的关联分析[J].中国油料作物学报,2014,36(1):32-37.
[3] Moazzami A A,Kamal-Eldin A. Sesame seed is a rich source of dietary lignans[J]. J Amer Oil Chem Soc,2006,83(8):719.
[4] Wang L H,Zhang Y X,Zhu X D,et al. Development of an SSR-based genetic map in sesame and identification of quantitative trait loci associated with charcoal rot resistance[J]. Sci Rep,2017,7:8349.
[5] Li D H,Liu P,Yu J Y,et al. Genome-wide analysis of WRKY gene family in the sesame genome and identification of the WRKY genes involved in responses to abiotic stresses[J]. BMC Plant Biol,2017,17:152.
[6]丁霞,王林海,张艳欣,等.芝麻核心种质株高构成相关性状的遗传变异及关联定位[J].中国油料作物学报,2013,35(3):262-270.
[7] Kurosawa E. Experimental studies on the nature of the substance secreted by the “bakanae” fungus[J]. Nat Hist Soc Formosa,1926; 16:213-227.
[8] Yabuta T. Biochemistry of the‘bakanae’fungus of rice[J]. Agric Hort(Tokyo),1935,10:17-22.
[9] Hedden P,Kamiya Y. Gibberelin biosynthesis:enzymes,genes and their regulation[J]. Annu Rev Plant Physiol Plant Mol Biol,1997,48(1):431-460.
[10] Hedden P,Phillips A L. Gibberellin metabolism:new insights revealed by the genes[J]. Trends Plant Sci,2000,5(12):523-530.
[11] Sponsel V M,Hedden P. Gibberellin biosynthesis and inactivation[M]//Plant Hormones. Dordrecht:Springer Netherlands,2010:63-94. DOI:10. 1007/978-1-4020-2686-7_4.
[12] MacMillan J. Occurrence of gibberellins in vascular plants,fungi,and bacteria[J]. J Plant Growth Regul,2001,20(4):387-442.
[13] Davies,Peter J. Plant hormones:physiology,biochemistry and molecular biology[J]. Scientia Horticulturae,1996,66(3):267-270.
[14] Olszewski N,Sun T P,Gubler F. Gibberellin signaling:biosynthesis,catabolism,and response pathways[J].The Plant Cell,2002,14 Suppl:61-80.
[15]吴建明,陈荣发,黄杏,等.高等植物赤霉素生物合成关键组分GA20-oxidase氧化酶基因的研究进展[J].生物技术通报,2016(7):1-12.
[16] Monna L. Positional cloning of rice semidwarfing gene,sd-1:rice “green revolution gene”encodes a mutant enzyme involved in gibberellin synthesis[J]. DNA Res,2002,9(1):11-17.
[17] Chiang H H,Hwang I,Goodman H M. Isolation of the Arabidopsis GA4 locus[J]. Plant Cell,1995,7(2):195.
[18] Wuddineh W A,Mazarei M,Zhang J Y,et al. Identification and overexpression of gibberellin 2-oxidase(GA2ox)in switchgrass(Panicum virgatum L.)for improved plant architecture and reduced biomass recalcitrance[J]. Plant Biotechnol J,2015,13(5):636-647.
[19] Sakai M,Sakamoto T,Saito T,et al. Expression of novel rice gibberellin 2-oxidase gene is under homeostatic regulation by biologically active gibberellins[J]. J Plant Res,2003,116(2):161-164.
[20] Hedden P,Sponsel V. A century of gibberellin research[J]. J Plant Growth Regul,2015,34(4):740-760.
[21]王西成,任国慧,房经贵,等.葡萄赤霉素合成相关基因克隆、亚细胞定位和表达分析[J].中国农业科学,2012,45(11):2224-2231.
[22] Plackett A R G,Thomas S G,Wilson Z A,et al. Gibberellin control of stamen development:a fertile field[J]. Trends Plant Sci,2011,16(10):568-578.
[23]肖景华.番茄GA 20-氧化酶和GA 2-氧化酶基因的克隆与功能分析[D].武汉:华中农业大学,2006.
[24] Schomburg F M. Overexpression of a novel class of gibberellin 2-oxidases decreases gibberellin levels and creates dwarf plants[J]. Plant Cell,2003,15(1):151-163.
[25] Helliwell C A. The CYP88A cytochrome P450,entkaurenoic acid oxidase,catalyzes three steps of the gibberellin biosynthesis pathway[J]. Proc Natl Acad Sci,2001,98(4):2065-2070.
[26] Helliwell C A,Sullivan J A,Mould R M,et al. A plastid envelope location of Arabidopsis ent-kaurene oxidase links the plastid and endoplasmic reticulum steps of the gibberellin biosynthesis pathway[J]. Plant J,2001,28(2):201-208.
[27]朱晓凤,黎冬华,王林海,等.矮秆与高秆芝麻株高建成中内源激素含量变化比较分析[J].中国油料作物学报,2015,37(1):83-89.
[28] Wang L H,Yu S,Tong C B,et al. Genome sequencing of the high oil crop sesame provides insight into oil biosynthesis[J]. Genome Biol,2014,15(2):39.
[29] Chou K C,Shen H B. Cell-PLoc 2. 0:an improved package of web-servers for predicting subcellular localization of proteins in various organisms[J]. Nat Sci,2010,2(10):1090-1103.
[30] Deng W K,Wang Y B,Liu Z X,et al. Hem I:A toolkit for illustrating heatmaps[J]. Plos One,2014,9(11):e111988. DOI:10. 1371/journal. pone. 0111988.
[31] Lewis D F V,Watson E,Lake B G. Evolution of the cytochrome P450 superfamily:sequence alignments and pharmacogenetics[J]. Mutat Res Mutat Res,1998,410(3):245-270.
[32] Yu J Y,Wang L H,Guo H,et al. Genome evolutionary dynamics followed by diversifying selection explains the complexity of the Sesamum indicum genome[J]. Bmc Genom,2017,18:257.
[33] The Tomato Genome Consortium. The tomato genome sequence provides insights into fleshy fruit evolution[J].Nature,2012,485(7400):635-641.
[34] Han F M,Zhu B G. Evolutionary analysis of three gibberellin oxidase genesin rice,Arabidopsis,and soybean[J]. Gene,2011,473(1):23-35.
[35]王增辉.黄瓜赤霉素氧化酶基因GA20ox、GA2ox、GA3ox的生物信息学及GA20ox1功能分析[D].泰安:山东农业大学,2014.
[36] Smith V A,Knatt C J,Gaskin P,et al. The distribution of gibberellins in vegetative tissues of Pisum sativum L.:I. biological and biochemical consequences of the le mutation[J]. Plant Physiol,1992,99(2):368-371.
[37] Fleet C M. Overexpression of At CPS and At KS in Arabidopsis confers increased ent-kaurene production but no increase in bioactive gibberellins[J]. Plant Physiol,2003,132(2):830-839.
[38] Mitchum M G,Yamaguchi S,Hanada A,et al. Distinct and overlapping roles of two gibberellin 3-oxidases in Arabidopsis development[J]. Plant J,2006,45(5):804-818.
[39] Zhao H J,Dong J L,Wang T. Function and expression analysis of gibberellin oxidases in apple[J]. Plant Mol Biol Rep,2010,28(2):231-238.
[40] Wu K,Li L,Gage D A,et al. Molecular cloning and photoperiod-regulated expression of gibberellin 20-oxidase from the long-day plant spinach[J]. Plant Physiol,1996,110(2):547-554.
[41] Zhao X Y,Zhu D F,Zhou B,et al. Over-expression of the At GA2ox8 gene decreases the biomass accumulation and lignification in rapeseed(Brassica napus L.)[J]. J Zhejiang Univ Sci B,2010,11(7):471-481.
[42] Zhou B,Peng D,Lin J Z,et al. Heterologous expression of a gibberellin 2-oxidase gene from Arabidopsis thaliana enhanced the photosynthesis capacity in Brassica napus L[J]. J Plant Biol,2011,54(1):23-32.
[43] Zhou B,Lin J Z,Peng W S,et al. Dwarfism in Brassica napus L. induced by the over-expression of a gibberellin 2-oxidase gene from Arabidopsis thaliana[J].Mol Breeding,2012,29(1):115-127.
[2]王蕾,黎冬华,齐小琼,等.芝麻核心种质芝麻素和芝麻酚林的关联分析[J].中国油料作物学报,2014,36(1):32-37.
[3] Moazzami A A,Kamal-Eldin A. Sesame seed is a rich source of dietary lignans[J]. J Amer Oil Chem Soc,2006,83(8):719.
[4] Wang L H,Zhang Y X,Zhu X D,et al. Development of an SSR-based genetic map in sesame and identification of quantitative trait loci associated with charcoal rot resistance[J]. Sci Rep,2017,7:8349.
[5] Li D H,Liu P,Yu J Y,et al. Genome-wide analysis of WRKY gene family in the sesame genome and identification of the WRKY genes involved in responses to abiotic stresses[J]. BMC Plant Biol,2017,17:152.
[6]丁霞,王林海,张艳欣,等.芝麻核心种质株高构成相关性状的遗传变异及关联定位[J].中国油料作物学报,2013,35(3):262-270.
[7] Kurosawa E. Experimental studies on the nature of the substance secreted by the “bakanae” fungus[J]. Nat Hist Soc Formosa,1926; 16:213-227.
[8] Yabuta T. Biochemistry of the‘bakanae’fungus of rice[J]. Agric Hort(Tokyo),1935,10:17-22.
[9] Hedden P,Kamiya Y. Gibberelin biosynthesis:enzymes,genes and their regulation[J]. Annu Rev Plant Physiol Plant Mol Biol,1997,48(1):431-460.
[10] Hedden P,Phillips A L. Gibberellin metabolism:new insights revealed by the genes[J]. Trends Plant Sci,2000,5(12):523-530.
[11] Sponsel V M,Hedden P. Gibberellin biosynthesis and inactivation[M]//Plant Hormones. Dordrecht:Springer Netherlands,2010:63-94. DOI:10. 1007/978-1-4020-2686-7_4.
[12] MacMillan J. Occurrence of gibberellins in vascular plants,fungi,and bacteria[J]. J Plant Growth Regul,2001,20(4):387-442.
[13] Davies,Peter J. Plant hormones:physiology,biochemistry and molecular biology[J]. Scientia Horticulturae,1996,66(3):267-270.
[14] Olszewski N,Sun T P,Gubler F. Gibberellin signaling:biosynthesis,catabolism,and response pathways[J].The Plant Cell,2002,14 Suppl:61-80.
[15]吴建明,陈荣发,黄杏,等.高等植物赤霉素生物合成关键组分GA20-oxidase氧化酶基因的研究进展[J].生物技术通报,2016(7):1-12.
[16] Monna L. Positional cloning of rice semidwarfing gene,sd-1:rice “green revolution gene”encodes a mutant enzyme involved in gibberellin synthesis[J]. DNA Res,2002,9(1):11-17.
[17] Chiang H H,Hwang I,Goodman H M. Isolation of the Arabidopsis GA4 locus[J]. Plant Cell,1995,7(2):195.
[18] Wuddineh W A,Mazarei M,Zhang J Y,et al. Identification and overexpression of gibberellin 2-oxidase(GA2ox)in switchgrass(Panicum virgatum L.)for improved plant architecture and reduced biomass recalcitrance[J]. Plant Biotechnol J,2015,13(5):636-647.
[19] Sakai M,Sakamoto T,Saito T,et al. Expression of novel rice gibberellin 2-oxidase gene is under homeostatic regulation by biologically active gibberellins[J]. J Plant Res,2003,116(2):161-164.
[20] Hedden P,Sponsel V. A century of gibberellin research[J]. J Plant Growth Regul,2015,34(4):740-760.
[21]王西成,任国慧,房经贵,等.葡萄赤霉素合成相关基因克隆、亚细胞定位和表达分析[J].中国农业科学,2012,45(11):2224-2231.
[22] Plackett A R G,Thomas S G,Wilson Z A,et al. Gibberellin control of stamen development:a fertile field[J]. Trends Plant Sci,2011,16(10):568-578.
[23]肖景华.番茄GA 20-氧化酶和GA 2-氧化酶基因的克隆与功能分析[D].武汉:华中农业大学,2006.
[24] Schomburg F M. Overexpression of a novel class of gibberellin 2-oxidases decreases gibberellin levels and creates dwarf plants[J]. Plant Cell,2003,15(1):151-163.
[25] Helliwell C A. The CYP88A cytochrome P450,entkaurenoic acid oxidase,catalyzes three steps of the gibberellin biosynthesis pathway[J]. Proc Natl Acad Sci,2001,98(4):2065-2070.
[26] Helliwell C A,Sullivan J A,Mould R M,et al. A plastid envelope location of Arabidopsis ent-kaurene oxidase links the plastid and endoplasmic reticulum steps of the gibberellin biosynthesis pathway[J]. Plant J,2001,28(2):201-208.
[27]朱晓凤,黎冬华,王林海,等.矮秆与高秆芝麻株高建成中内源激素含量变化比较分析[J].中国油料作物学报,2015,37(1):83-89.
[28] Wang L H,Yu S,Tong C B,et al. Genome sequencing of the high oil crop sesame provides insight into oil biosynthesis[J]. Genome Biol,2014,15(2):39.
[29] Chou K C,Shen H B. Cell-PLoc 2. 0:an improved package of web-servers for predicting subcellular localization of proteins in various organisms[J]. Nat Sci,2010,2(10):1090-1103.
[30] Deng W K,Wang Y B,Liu Z X,et al. Hem I:A toolkit for illustrating heatmaps[J]. Plos One,2014,9(11):e111988. DOI:10. 1371/journal. pone. 0111988.
[31] Lewis D F V,Watson E,Lake B G. Evolution of the cytochrome P450 superfamily:sequence alignments and pharmacogenetics[J]. Mutat Res Mutat Res,1998,410(3):245-270.
[32] Yu J Y,Wang L H,Guo H,et al. Genome evolutionary dynamics followed by diversifying selection explains the complexity of the Sesamum indicum genome[J]. Bmc Genom,2017,18:257.
[33] The Tomato Genome Consortium. The tomato genome sequence provides insights into fleshy fruit evolution[J].Nature,2012,485(7400):635-641.
[34] Han F M,Zhu B G. Evolutionary analysis of three gibberellin oxidase genesin rice,Arabidopsis,and soybean[J]. Gene,2011,473(1):23-35.
[35]王增辉.黄瓜赤霉素氧化酶基因GA20ox、GA2ox、GA3ox的生物信息学及GA20ox1功能分析[D].泰安:山东农业大学,2014.
[36] Smith V A,Knatt C J,Gaskin P,et al. The distribution of gibberellins in vegetative tissues of Pisum sativum L.:I. biological and biochemical consequences of the le mutation[J]. Plant Physiol,1992,99(2):368-371.
[37] Fleet C M. Overexpression of At CPS and At KS in Arabidopsis confers increased ent-kaurene production but no increase in bioactive gibberellins[J]. Plant Physiol,2003,132(2):830-839.
[38] Mitchum M G,Yamaguchi S,Hanada A,et al. Distinct and overlapping roles of two gibberellin 3-oxidases in Arabidopsis development[J]. Plant J,2006,45(5):804-818.
[39] Zhao H J,Dong J L,Wang T. Function and expression analysis of gibberellin oxidases in apple[J]. Plant Mol Biol Rep,2010,28(2):231-238.
[40] Wu K,Li L,Gage D A,et al. Molecular cloning and photoperiod-regulated expression of gibberellin 20-oxidase from the long-day plant spinach[J]. Plant Physiol,1996,110(2):547-554.
[41] Zhao X Y,Zhu D F,Zhou B,et al. Over-expression of the At GA2ox8 gene decreases the biomass accumulation and lignification in rapeseed(Brassica napus L.)[J]. J Zhejiang Univ Sci B,2010,11(7):471-481.
[42] Zhou B,Peng D,Lin J Z,et al. Heterologous expression of a gibberellin 2-oxidase gene from Arabidopsis thaliana enhanced the photosynthesis capacity in Brassica napus L[J]. J Plant Biol,2011,54(1):23-32.
[43] Zhou B,Lin J Z,Peng W S,et al. Dwarfism in Brassica napus L. induced by the over-expression of a gibberellin 2-oxidase gene from Arabidopsis thaliana[J].Mol Breeding,2012,29(1):115-127.
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