百合LaMYB26基因克隆与表达分析
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摘要
本研究从亚洲百合(Lilium Asiatic hybrids)花粉败育品种‘小小的吻’中克隆获得一个MYB转录因子基因LaMYB26。序列分析显示,LaMYB26基因的开放阅读框(ORF)长762 bp,编码253个氨基酸。结构域分析显示,LaMYB26含有2个典型的SANT结构域,属于MYB转录因子家族中的R2R3-MYB型。多序列比对发现,LaMYB26与其他物种的MYB类蛋白的氨基酸序列有57.78%的相似性。理化性质、亲/疏水性和无序化分析显示, LaMYB26转录因子是亲水性蛋白且偏碱性,存在无序化区域。信号肽和跨膜结构域分析表明,其不存在信号肽和跨膜结构域。空间结构分析显示, LaMYB26有2个典型的α-螺旋串联。实时荧光定量PCR (qRT-PCR)显示, LaMYB26基因在败育时期表达量明显上调,且在花药中特异表达。结果显示, LaMYB26基因在百合花粉败育过程中可能发挥着重要作用,对LaMYB26的生物学信息学分析拟为深入研究其功能及作用机制提供参考。
LaMYB26, a MYB transcription factor, was cloned from lily(Lilium Asiatic hybrids) cultivar ‘Little Kiss’ with pollen abortion in this study. Sequence analysis revealed that the open reading frame of LaMYB26 gene is 762 bp and encodes 253 amino acids. LaMYB26 transcription factor contains two typical SANT domains and belongs to the R2 R3-MYB type in the MYB family transcription factor. Multi-sequence comparison founds that the amino acid sequence of LaMYB26 is 57.78% in similarity to those of MYB proteins in other species. The physicochemical properties, hydrophilicity/hydrophobicity and disorder analyses of LaMYB26 transcription factor demonstrates that it is a hydrophilic and alkaline protein with a disordered region. Analyses of signal peptide, transmembrane domain, and spatial structure indicate that LaMYB26 has no signal peptide and transmembrane domain, but has two typical α-helix series. Quantitative real-time PCR showed that the expression level of LaMYB26 gene was up-regulated during pollen abortion stage and was specifically expressed in lily anther. Taken together, the results suggest that LaMYB26 gene may play an important role in lily pollen abortion, and the current bioinformatic analyses of LaMYB26 would lay a foundation for further studies of its function and mechanism.
LaMYB26, a MYB transcription factor, was cloned from lily(Lilium Asiatic hybrids) cultivar ‘Little Kiss’ with pollen abortion in this study. Sequence analysis revealed that the open reading frame of LaMYB26 gene is 762 bp and encodes 253 amino acids. LaMYB26 transcription factor contains two typical SANT domains and belongs to the R2 R3-MYB type in the MYB family transcription factor. Multi-sequence comparison founds that the amino acid sequence of LaMYB26 is 57.78% in similarity to those of MYB proteins in other species. The physicochemical properties, hydrophilicity/hydrophobicity and disorder analyses of LaMYB26 transcription factor demonstrates that it is a hydrophilic and alkaline protein with a disordered region. Analyses of signal peptide, transmembrane domain, and spatial structure indicate that LaMYB26 has no signal peptide and transmembrane domain, but has two typical α-helix series. Quantitative real-time PCR showed that the expression level of LaMYB26 gene was up-regulated during pollen abortion stage and was specifically expressed in lily anther. Taken together, the results suggest that LaMYB26 gene may play an important role in lily pollen abortion, and the current bioinformatic analyses of LaMYB26 would lay a foundation for further studies of its function and mechanism.
引文
Albert NW,Griffiths AG,Cousins GR,et al(2014).Anthocyanin leaf markings are regulated by a family of R2R3-MYB genes in the genus Trifolium.New Phytol,205(2):882-893
Boyer LA,Latek RR,Peterson CL(2004).The SANT domain:a unique histone-tail-binding module.Nat Rev Mol Cell Biol,5(2):158-163
Chen Q,Tang HR,Dong XL,et al(2009).Progress in the study of plant Myb transcription factors.Genom Appl Biol,28(2):365-372(in Chinese with English abstract)[陈清,汤浩茹,董晓莉等(2009).植物MYB转录因子的研究进展.基因组学与应用生物学,28(2):365-372]
Dawson J,S?zen E,Vizir I,et al(1999).Characterization and genetic mapping of a mutation(ms35)which prevents anther dehiscence in Arabidopsis thaliana by affecting secondary wall thickening in the endothecium.New Phytol,144(2):213-222
Ding AQ,Li SC,Liu QC,et al(2018).Cloning and expression analysis of Rh ATAF1 gene in Rosa hybrida.Plant Physiol J,54(11):1711-1718(in Chinese with English abstract)[丁爱琴,李绍翠,刘庆超等(2018).月季Rh ATAF1基因的克隆及表达特性分析.植物生理学报,54(11):1711-1718]
Dubos C,Stracke R,Grotewold E,et al(2010).MYB transcription factors in Arabidopsis.Trends Plant Sci,15(10):570-581
Fan JT,Jiang WX,Xing JH,et al(2014).Structure and function of the 22nd subfamily in Arabidopsis R2R3-MYBfamily.Hereditas Beijing,36(10):985-994(in Chinese with English abstract)[樊锦涛,蒋琛茜,邢继红等(2014).拟南芥R2R3-MYB家族第22亚族的结构与功能.遗传,36(10):985-994]
Feng DD(2008).Fine mapping of Arabidopsis thaliana gene controlling microspores development and Cloning of male sterility gene promoters from Arabidopsis(dissertation).Shanghai:Shanghai Normal University(in Chinese with English abstract)[冯丹丹(2008).一个控制拟南芥小孢子发育基因的定位和雄性不育基因启动子的克隆(学位论文).上海:上海师范大学]
Jenkins ES,Paul W,Craze M,et al(2010).Dehiscence-related expression of an Arabidopsis thaliana gene encoding a polygalacturonase in transgenic plants of Brassica napus.Plant Cell Environ,22(2):159-167
Kazama T,Itabashi E,Fujii S,et al(2016).Mitochondrial ORF79 levels determine pollen abortion in cytoplasmic male sterile rice.Plant J,85(6):707-716
Kranz HD,Denekamp M,Greco R,et al(1998).Towards functional characterisation of the members of the R2R3-MYB gene family from Arabidopsis thaliana.Plant J,16(2):263-276
Li YH,He SQ,Meng Y,et al(2018).Isolation and identification of the fungal pathogens causing root and bulb rot disease on edible lily(Lillium brownie var.viridulum)of Changping district in Beijing.J Beijing Univ Agr,33(2):32-37(in Chinese with English abstract)[李一寒,何思琦,孟嫣等(2018).北京昌平区食用百合根及鳞茎腐烂的病原菌分离与鉴定.北京农学院学报,33(2):32-37]
Liu Q,Zhang GY,Chen SY(2000).The structure and regulation of plant transcription factors.Chin Sci Bull,45(14):1465-1474[刘强,张贵友,陈受宜(2000).植物转录因子的结构与调控作用.科学通报,45(14):1465-1474]
Nelson MR,Band LR,Dyson RJ,et al(2012).A biomechanical model of anther opening reveals the roles of dehydration and secondary thickening.New Phytol,196(4):1030-1037
Paz-Ares J,Ghosal D,Wienand U,et al(1987).The regulatory c1 locus of Zea mays encodes a protein with homology to myb proto-oncogene products and with structural similarities to transcriptional activators.EMBO J,6(12):3553-3558
Riechmann JL,Heard J,Martin G,et al(2000).Arabidopsis transcription factors:genome-wide comparative analysis among eukaryotes.Science,290(5499):2105-2110
Steiner-Lange S,Unte US,Eckstein L,et al(2003).Disruption of Arabidopsis thaliana MYB26 results in male sterility due to non-dehiscent anthers.Plant J,34(4):519-528
Stracke R,Werber M,Weisshaar B(2001).The R2R3-MYBgene family in Arabidopsis thaliana.Curr Opin Plant Biol,4(5):447-456
Takos AM,JafféFW,Jacob SR,et al(2006).Light-induced expression of a MYB gene regulates anthocyanin biosynthesis in red apples.Plant Physiol,142(3):1216-1232
Tohge T,Nishiyama Y,Hirai MY,et al(2005).Functional genomics by integrated analysis of metabolome and transcriptome of Arabidopsis plants over-expressing an MYBtranscription factor.Plant J,42(2):218-235
Uimari A,Strommer J(1997).MYB26:a MYB-like protein of pea flowers with affinity for promoters of phenylpropanoid genes.Plant J,12(6):1273-1284
Wang K(2018).Arabidopsis transcription factor MS188regulates the expression of sporopollenin synthesis gene(dissertation).Shanghai:Shanghai Normal University(in Chinese with English abstract)[王科(2018).拟南芥转录因子MS188调控孢粉素合成基因表达的研究(学位论文).上海:上海师范大学]
Wang LQ,Wu MJ,Wang XQ,et al(2018).Determination of pollen quantity and features of pollen dispersal for 95lily resources.J Nanjing Agr Univ,41(6):1018-1028(in Chinese with English abstract)[王蓝青,吴美娇,王雪倩等(2018).95份百合种质资源花粉量的测定与散粉特性分析.南京农业大学学报,41(6):1018-1028]
Yang C,Song J,Ferguson AC,et al(2017).Transcription factor MYB26 is key to spatial specificity in anther secondary thickening formation.Plant Physiol,75:333-350
Yang ZR,Wang XC,Li XM,et al(2004).Advance on the study of transcription factors in higher plants.Hereditas Beijing,26(3):403-408(in Chinese with English abstract)[杨致荣,王兴春,李西明等(2004).高等植物转录因子的研究进展.遗传,26(3):403-408]
Zhang F,Liu X,Zuo K,et al(2011).Molecular cloning and expression analysis of a novel SANT/MYB gene from Gossypium barbadense.Mol Biol Rep,38(4):2329-2336
Zhang J,Wu J,Zeng JY(2005).Study on stability of anther pigment of Lilium siberia.Food Sci,26(1):88-90(in Chinese with English abstract)[张继,吴建,曾家豫(2005).西伯利亚百合花药色素稳定性的研究.食品科学,26(1):88-90]
Zhang M,Miao YJ,Li YF,et al(2016).Determination of the critical period of floral induction for hybrid plants between Lilium×formolongi and qriental hybrid lily.Acta Hortic Sin,43(4):735-742(in Chinese with English abstract)[张盟,苗英靖,李玉帆等(2016).新铁炮百合与东方百合杂种成花诱导关键时期的确定.园艺学报,43(4):735-742]
Zhang S,Fang ZJ,Zhu J,et al(2010).OsMYB103 is required for rice anther development by regulating tapetum development and exine formation.Chin Sci Bull,55(29):3288-3297
Zhao XN,Li JC,Zhang P(2018).Several problems in the process of artificial emasculation of cotton.Mod Rural Sci Technol,(9):100(in Chinese)[赵香娜,李记臣,张鹏(2018).棉花人工去雄杂交过程中的几个问题.现代农村科技,(9):100]
Zhao XY,Wang WH(2017).Market expansion,strengths combination,cooperation and win-win situation:the status quo,existing problems and development prospects of China’s lily industry.China Flowers Hortic,(13):10-13(in Chinese)[赵祥云,王文和(2017).拓展市场强强联手合作共赢--我国百合产业现状、存在问题和发展前景.中国花卉园艺,(13):10-13]
Zhao XH,Yin DS,Zhang DX,et al(2009).Breeding of an innovative resource of anther degraded male sterile lily“No Powder White”.Liaoning Agr Sci,(1):61(in Chinese)[赵兴华,印东生,张道旭等(2009).花药退化型雄性不育百合创新资源“无粉白”的选育.辽宁农业科学,(1):61]
Zhuang Y,Sun XM,Wang XY,et al(2010).The influence of ambient air pollutants on outpatient visits for allergic disease and pollinosis.Chin J Prev Med,44(12):1121-1127(in Chinese with English abstract)[庄严,孙新民,王学艳等(2010).大气污染对变应性疾病及花粉症就诊人次的影响.中华预防医学杂志,44(12):1121-1127]
Zuo R,Xu ML,Chai GH,et al(2012).Function and regulation mechanism of plant MYB transcription factors.Chin Bull Life Sci,24(10):1133-1140(in Chinese with English abstract)[左然,徐美玲,柴国华等(2012).植物MYB转录因子功能及调控机制研究进展.生命科学,24(10):1133-1140]
Boyer LA,Latek RR,Peterson CL(2004).The SANT domain:a unique histone-tail-binding module.Nat Rev Mol Cell Biol,5(2):158-163
Chen Q,Tang HR,Dong XL,et al(2009).Progress in the study of plant Myb transcription factors.Genom Appl Biol,28(2):365-372(in Chinese with English abstract)[陈清,汤浩茹,董晓莉等(2009).植物MYB转录因子的研究进展.基因组学与应用生物学,28(2):365-372]
Dawson J,S?zen E,Vizir I,et al(1999).Characterization and genetic mapping of a mutation(ms35)which prevents anther dehiscence in Arabidopsis thaliana by affecting secondary wall thickening in the endothecium.New Phytol,144(2):213-222
Ding AQ,Li SC,Liu QC,et al(2018).Cloning and expression analysis of Rh ATAF1 gene in Rosa hybrida.Plant Physiol J,54(11):1711-1718(in Chinese with English abstract)[丁爱琴,李绍翠,刘庆超等(2018).月季Rh ATAF1基因的克隆及表达特性分析.植物生理学报,54(11):1711-1718]
Dubos C,Stracke R,Grotewold E,et al(2010).MYB transcription factors in Arabidopsis.Trends Plant Sci,15(10):570-581
Fan JT,Jiang WX,Xing JH,et al(2014).Structure and function of the 22nd subfamily in Arabidopsis R2R3-MYBfamily.Hereditas Beijing,36(10):985-994(in Chinese with English abstract)[樊锦涛,蒋琛茜,邢继红等(2014).拟南芥R2R3-MYB家族第22亚族的结构与功能.遗传,36(10):985-994]
Feng DD(2008).Fine mapping of Arabidopsis thaliana gene controlling microspores development and Cloning of male sterility gene promoters from Arabidopsis(dissertation).Shanghai:Shanghai Normal University(in Chinese with English abstract)[冯丹丹(2008).一个控制拟南芥小孢子发育基因的定位和雄性不育基因启动子的克隆(学位论文).上海:上海师范大学]
Jenkins ES,Paul W,Craze M,et al(2010).Dehiscence-related expression of an Arabidopsis thaliana gene encoding a polygalacturonase in transgenic plants of Brassica napus.Plant Cell Environ,22(2):159-167
Kazama T,Itabashi E,Fujii S,et al(2016).Mitochondrial ORF79 levels determine pollen abortion in cytoplasmic male sterile rice.Plant J,85(6):707-716
Kranz HD,Denekamp M,Greco R,et al(1998).Towards functional characterisation of the members of the R2R3-MYB gene family from Arabidopsis thaliana.Plant J,16(2):263-276
Li YH,He SQ,Meng Y,et al(2018).Isolation and identification of the fungal pathogens causing root and bulb rot disease on edible lily(Lillium brownie var.viridulum)of Changping district in Beijing.J Beijing Univ Agr,33(2):32-37(in Chinese with English abstract)[李一寒,何思琦,孟嫣等(2018).北京昌平区食用百合根及鳞茎腐烂的病原菌分离与鉴定.北京农学院学报,33(2):32-37]
Liu Q,Zhang GY,Chen SY(2000).The structure and regulation of plant transcription factors.Chin Sci Bull,45(14):1465-1474[刘强,张贵友,陈受宜(2000).植物转录因子的结构与调控作用.科学通报,45(14):1465-1474]
Nelson MR,Band LR,Dyson RJ,et al(2012).A biomechanical model of anther opening reveals the roles of dehydration and secondary thickening.New Phytol,196(4):1030-1037
Paz-Ares J,Ghosal D,Wienand U,et al(1987).The regulatory c1 locus of Zea mays encodes a protein with homology to myb proto-oncogene products and with structural similarities to transcriptional activators.EMBO J,6(12):3553-3558
Riechmann JL,Heard J,Martin G,et al(2000).Arabidopsis transcription factors:genome-wide comparative analysis among eukaryotes.Science,290(5499):2105-2110
Steiner-Lange S,Unte US,Eckstein L,et al(2003).Disruption of Arabidopsis thaliana MYB26 results in male sterility due to non-dehiscent anthers.Plant J,34(4):519-528
Stracke R,Werber M,Weisshaar B(2001).The R2R3-MYBgene family in Arabidopsis thaliana.Curr Opin Plant Biol,4(5):447-456
Takos AM,JafféFW,Jacob SR,et al(2006).Light-induced expression of a MYB gene regulates anthocyanin biosynthesis in red apples.Plant Physiol,142(3):1216-1232
Tohge T,Nishiyama Y,Hirai MY,et al(2005).Functional genomics by integrated analysis of metabolome and transcriptome of Arabidopsis plants over-expressing an MYBtranscription factor.Plant J,42(2):218-235
Uimari A,Strommer J(1997).MYB26:a MYB-like protein of pea flowers with affinity for promoters of phenylpropanoid genes.Plant J,12(6):1273-1284
Wang K(2018).Arabidopsis transcription factor MS188regulates the expression of sporopollenin synthesis gene(dissertation).Shanghai:Shanghai Normal University(in Chinese with English abstract)[王科(2018).拟南芥转录因子MS188调控孢粉素合成基因表达的研究(学位论文).上海:上海师范大学]
Wang LQ,Wu MJ,Wang XQ,et al(2018).Determination of pollen quantity and features of pollen dispersal for 95lily resources.J Nanjing Agr Univ,41(6):1018-1028(in Chinese with English abstract)[王蓝青,吴美娇,王雪倩等(2018).95份百合种质资源花粉量的测定与散粉特性分析.南京农业大学学报,41(6):1018-1028]
Yang C,Song J,Ferguson AC,et al(2017).Transcription factor MYB26 is key to spatial specificity in anther secondary thickening formation.Plant Physiol,75:333-350
Yang ZR,Wang XC,Li XM,et al(2004).Advance on the study of transcription factors in higher plants.Hereditas Beijing,26(3):403-408(in Chinese with English abstract)[杨致荣,王兴春,李西明等(2004).高等植物转录因子的研究进展.遗传,26(3):403-408]
Zhang F,Liu X,Zuo K,et al(2011).Molecular cloning and expression analysis of a novel SANT/MYB gene from Gossypium barbadense.Mol Biol Rep,38(4):2329-2336
Zhang J,Wu J,Zeng JY(2005).Study on stability of anther pigment of Lilium siberia.Food Sci,26(1):88-90(in Chinese with English abstract)[张继,吴建,曾家豫(2005).西伯利亚百合花药色素稳定性的研究.食品科学,26(1):88-90]
Zhang M,Miao YJ,Li YF,et al(2016).Determination of the critical period of floral induction for hybrid plants between Lilium×formolongi and qriental hybrid lily.Acta Hortic Sin,43(4):735-742(in Chinese with English abstract)[张盟,苗英靖,李玉帆等(2016).新铁炮百合与东方百合杂种成花诱导关键时期的确定.园艺学报,43(4):735-742]
Zhang S,Fang ZJ,Zhu J,et al(2010).OsMYB103 is required for rice anther development by regulating tapetum development and exine formation.Chin Sci Bull,55(29):3288-3297
Zhao XN,Li JC,Zhang P(2018).Several problems in the process of artificial emasculation of cotton.Mod Rural Sci Technol,(9):100(in Chinese)[赵香娜,李记臣,张鹏(2018).棉花人工去雄杂交过程中的几个问题.现代农村科技,(9):100]
Zhao XY,Wang WH(2017).Market expansion,strengths combination,cooperation and win-win situation:the status quo,existing problems and development prospects of China’s lily industry.China Flowers Hortic,(13):10-13(in Chinese)[赵祥云,王文和(2017).拓展市场强强联手合作共赢--我国百合产业现状、存在问题和发展前景.中国花卉园艺,(13):10-13]
Zhao XH,Yin DS,Zhang DX,et al(2009).Breeding of an innovative resource of anther degraded male sterile lily“No Powder White”.Liaoning Agr Sci,(1):61(in Chinese)[赵兴华,印东生,张道旭等(2009).花药退化型雄性不育百合创新资源“无粉白”的选育.辽宁农业科学,(1):61]
Zhuang Y,Sun XM,Wang XY,et al(2010).The influence of ambient air pollutants on outpatient visits for allergic disease and pollinosis.Chin J Prev Med,44(12):1121-1127(in Chinese with English abstract)[庄严,孙新民,王学艳等(2010).大气污染对变应性疾病及花粉症就诊人次的影响.中华预防医学杂志,44(12):1121-1127]
Zuo R,Xu ML,Chai GH,et al(2012).Function and regulation mechanism of plant MYB transcription factors.Chin Bull Life Sci,24(10):1133-1140(in Chinese with English abstract)[左然,徐美玲,柴国华等(2012).植物MYB转录因子功能及调控机制研究进展.生命科学,24(10):1133-1140]