小麦表皮模式建成因子基因(TaEPFL1)的克隆、定位及表达分析
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摘要
表皮模式建成因子(EPFL)基因家族的成员参与调节植物一系列生长发育过程,包括花序的结构和气孔密度等。本研究从小麦雄蕊同源转化型不育突变体HTS-1中克隆出一个EPFL基因(TaEPFL1)。测序结果表明,该基因由3个同源基因组成(TaEPFL1.1,TaEPFL1.2,TaEPFL1.3)。通过小麦缺体-四体分析(NT)表明这3个同源基因分别位于染色体6D、6B和6A上。3个同源基因的开放阅读框(ORF)长度为363 bp,编码120个氨基酸,其中TaEPFL1.2和TaEPFL1.3的ORF完全相同。聚类分析表明TaEPFL1属于植物EPFL基因家族成员,与ZmEPFL1,ObEPFL1和AtEPFL1关系较近。Real-time PCR结果表明,TaEPFL1在HTS-1雌蕊化雄蕊(PS)中的表达异常显著。由此,我们推断TaEPFL1基因过量表达可能会导致雄蕊向雌蕊或雌蕊化结构的同源转化。本研究为进一步研究TaEPFL1基因的功能和阐明了小麦雄蕊同源转化为雌蕊的分子机制提供科学依据。
Members of the epidermal pattering factor-like(EPFL) family participate in regulating a series of growth and development in plant, including the control of stomatal density and guidance of pollen tube elongation. In this work, we identified and characterized an EPFL gene(TaEFPL1) in wheat pistillody mutant HTS-1. Sequencing results indicated that this gene consisted of three homologous genes(TaEFPL1.1, TaEFPL1.2, and TaEFPL1.3).Also, the wheat nulli tetrasome analysis(NT) showed that these 3 homologous genes were located at chromosome6 D, 6 B and 6 A. The open reading frame(ORF) of the three homologous genes was 363 bp, encoding 120 amino acids. The ORF sequences of TaEPFL1.2 and TaEPF1.3 were the same. The phylogenetic analysis indicated that TaEPFL1 belonged to the EPFL family in plant, and that it was closely related to ZmEPFL1, ObEPFL1, and AtEPFL1. Real-time PCR revealed that the TaEPFL1 gene showed an abnormally high expression in pistillody stamens in HTS-1. Therefore, we deduced that the overexpression of the TaEPFL1 gene might contribute to the homeotic transformation of stamens into pistils or pistil-like structures. These data provided a basis for the futureresearch on the function of TaEPFL1 gene and the molecular mechanism of the transformation of stamen homologous to pistil in wheat.
Members of the epidermal pattering factor-like(EPFL) family participate in regulating a series of growth and development in plant, including the control of stomatal density and guidance of pollen tube elongation. In this work, we identified and characterized an EPFL gene(TaEFPL1) in wheat pistillody mutant HTS-1. Sequencing results indicated that this gene consisted of three homologous genes(TaEFPL1.1, TaEFPL1.2, and TaEFPL1.3).Also, the wheat nulli tetrasome analysis(NT) showed that these 3 homologous genes were located at chromosome6 D, 6 B and 6 A. The open reading frame(ORF) of the three homologous genes was 363 bp, encoding 120 amino acids. The ORF sequences of TaEPFL1.2 and TaEPF1.3 were the same. The phylogenetic analysis indicated that TaEPFL1 belonged to the EPFL family in plant, and that it was closely related to ZmEPFL1, ObEPFL1, and AtEPFL1. Real-time PCR revealed that the TaEPFL1 gene showed an abnormally high expression in pistillody stamens in HTS-1. Therefore, we deduced that the overexpression of the TaEPFL1 gene might contribute to the homeotic transformation of stamens into pistils or pistil-like structures. These data provided a basis for the futureresearch on the function of TaEPFL1 gene and the molecular mechanism of the transformation of stamen homologous to pistil in wheat.
引文
Abrash E.B.,and Bergmann D.C.,2010,Regional specification of stomatal production by the putative ligand CHALLAH,Development,137(3):447-455
Abrash E.B.,Davies K.A.,and Bergmann D.C.,2011,Generation of signaling specificity in Arabidopsis by spatially restricted buffering of ligand-receptor interactions,Plant Cell,23(8):2864-2879
Besshouehara K.,Wang D.R.,Furuta T.,Minami A.,Nagai K.,Gamuyao R.,Asano K.,Angeles-Shim R.B.,Shimizu Y.,Ayano M.,Komeda N.,Doi K.,Miura K.,Toda Y.,Kinoshita T.,Okuda S.,Higashiyama T.,Nomoto M.,Tada Y.,Shinohara H.,Matsubayashi Y.,Greenberg A.,Wu J.,Yasui H.,Yoshimura A.,Mori H.,Mc Couch S.R.,and Ashikari M.,2016,Loss of function at RAE2,a previously unidentified EPFL,is required for awnlessness in cultivated Asian rice,Proc.Natl.Acad.Sci.USA,113(32):8969-8974
Endo T.R.,and Gill B.S.,1996,The deletion stocks of common wheat,J.Hered.,87(4):295-307
Hama E.,Takumi S.,Ogihara Y.,and Murai K.,2004,Pistillody is caused by alterations to the class-B MADS-box gene expression pattern in alloplasmic wheats,Planta,218(5):712-720
Hara K.,Kajita R.,Torii K.U.,Bergmann D.C.,and Kakimoto T.,2007,The secretory peptide gene EPF1 enforces the stomatal one-cell-spacing rule,Genes Dev.,21(14):1720-1725
Hara K.,Yokoo T.,Kajita R.,Onishi T.,Yahata S.,Peterson K.M.,Torii K.U.,and Kakimoto T.,2009,Epidermal cell density is autoregulated via a secretory peptide,EPIDERMALPATTERNING FACTOR 2 in Arabidopsis leaves,Plant Cell Physiol.,50(6):1019-1031
Huang Y.,Tao Z.S.,Liu Q.,Wang X.F.,Yu J.Y.,Liu G.H.,and Wang H.Z.,2014,Bn EPFL6,an EPIDERMAL PATTERN-ING FACTOR-LIKE(EPFL)secreted peptide gene,is required for filament elongation in Brassica napus,Plant Mol.Biol.,85(4-5):505-517
Hunt L.,Bailey K.J.,and Gray J.E.,2010,The signalling peptide EPFL9 is a positive regulator of stomatal development,New Phytol.,186(3):609-614
Jewaria P.K.,Hara T.,Tanaka H.,Kondo T.,Betsuyaku S.,Sawa S.,Sakagami Y.,Aimoto S.,and Kakimoto T.,2013,Differential effect of peptides stomagen,EPF1 and EPF2 on activation of MAP kinase MPK6 and SPCH protein level,Plant Cell Physiol.,54(8):1253-1262
Jiang H.M.and Shi G.K.,2009,Advanced research on male sterility in wheat,Shengwu Jishu Tongbao(Biotechnology Bulletin),(10):30-33(江红梅,师光开,2009,小麦雄性不育研究进展,生物技术通报,(10):30-33)
Kimber G.,and Sears E.R.,1980,Uses of wheat aneuploids,In:Lewis W.H.,(ed.),Polyploidy,Plenum Press,New York,USA,pp.427-428
Kondo T.,Kajita R.,Miyazaki A.,Hokoyama M.,Nakamura-Miura T.,Mizuno S.,Masuda Y.,Irie K.,Tanaka Y.,Takada S.,Kakimoto T.,and Sakagami Y.,2010,Stomatal density is controlled by a mesophyll-derived signaling molecule,Plant Cell Physiol.,51(1):1-8
Livak K.J.,and Schmittgen T.D.,2001,Analysis of relative gene expression data using real time quantitative PCR and the2-ΔΔCtmethod,Methods,25(4):402-408
Marshall E.,Costa L.M.,and Gutierrez-Marcos J.,2011,Cysteine-rich peptides(CRPs)mediate diverse aspects of cell-cell communication in plant reproduction and development,J.Exp.Bot.,62(5):1677-1686
Meng X.Z.,Wang H.C.,He Y.X.,Liu Y.D.,Walker J.C.,Torii K.U.,and Zhang S.Q.,2012,A MAPK cascade downstream of ERECTA receptor-like protein kinase regulates Arabidopsis inflorescence architecture by promoting localized cell proliferation,Plant Cell,24(12):4948-4960
Murai K.,Miyamae M.,Kato H.,Takumi S.,and Ogihara Y.,2003,WAP1,a wheat APETALA1 homolog,plays a central role in the phase transition from vegetative to reproductive growth,Plant Cell Physiol.,44(12):1255-1265
Murai K.,Takumi S.,Koga H.,and Ogihara Y.,2002,Pistillody,homeotic transformation of stamens into pistil-like structures,caused by nuclear-cytoplasm interaction in wheat,Plant J.,29(2):169-181
Peng Z.S.,Yang Z.J.,Ouyang Z.M.and Yang H.,2013,Characterization of a novel pistillody mutant in common wheat,Aust.J.Crop Sci.,7(1):159-164
Sugano S.S.,Shimada T.,Imai Y.,Okawa K.,Tamai A.,Mori M.,and Hara-Nishimura I.,2010,Stomagen positively regulates stomatal density in Arabidopsis,Nature,463(7278):241-244
Takatsuji H.,Nakamura N.,and Katsumoto Y.,1994,A new family of zinc finger proteins in petunia:structure,DNA sequence recognition,and floral organ-specific expression,Plant Cell,6(7):947-958
Uchida N.,Lee J.S.,Horst R.J.,Lai H.H.,Kajita R.,Kakimoto T.,Tasaka M.,and Torii K.U.,2012,Regulation of inflorescence architecture by intertissue layer ligand-receptor communication between endodermis and phloem,Proc.Natl.A-cad.Sci.USA,109(16):6337-6342
Uchida N.,and Tasaka M.,2013,Regulation of plant vascular stem cells by endodermis-derived EPFL-family peptide hormones and phloem-expressed ERECTA-family receptor kinases,J.Exp.Bot.,64(17):5335-5343
Yamada K.,Saraike T.,Shitsukawa N.,Hirabayashi C.,Takumi S.,and Murai K.,2009,Class D and B(sister)MADS-box genes are associated with ectopic ovule formation in the pistil-like stamens of alloplasmic wheat(Triticum aestivum L.),Plant Mol.Biol.,71(1-2):1-14
Yang Z.J.,Peng Z.S.,Wei S.H.,Liao M.L.,Yu Y.,and Jang Z.Y.,2015,Pistillody mutant reveals key insights into stamen and pistil development in wheat(Triticum aestivum L.),BMC Genomics,16(1):211-220
Abrash E.B.,Davies K.A.,and Bergmann D.C.,2011,Generation of signaling specificity in Arabidopsis by spatially restricted buffering of ligand-receptor interactions,Plant Cell,23(8):2864-2879
Besshouehara K.,Wang D.R.,Furuta T.,Minami A.,Nagai K.,Gamuyao R.,Asano K.,Angeles-Shim R.B.,Shimizu Y.,Ayano M.,Komeda N.,Doi K.,Miura K.,Toda Y.,Kinoshita T.,Okuda S.,Higashiyama T.,Nomoto M.,Tada Y.,Shinohara H.,Matsubayashi Y.,Greenberg A.,Wu J.,Yasui H.,Yoshimura A.,Mori H.,Mc Couch S.R.,and Ashikari M.,2016,Loss of function at RAE2,a previously unidentified EPFL,is required for awnlessness in cultivated Asian rice,Proc.Natl.Acad.Sci.USA,113(32):8969-8974
Endo T.R.,and Gill B.S.,1996,The deletion stocks of common wheat,J.Hered.,87(4):295-307
Hama E.,Takumi S.,Ogihara Y.,and Murai K.,2004,Pistillody is caused by alterations to the class-B MADS-box gene expression pattern in alloplasmic wheats,Planta,218(5):712-720
Hara K.,Kajita R.,Torii K.U.,Bergmann D.C.,and Kakimoto T.,2007,The secretory peptide gene EPF1 enforces the stomatal one-cell-spacing rule,Genes Dev.,21(14):1720-1725
Hara K.,Yokoo T.,Kajita R.,Onishi T.,Yahata S.,Peterson K.M.,Torii K.U.,and Kakimoto T.,2009,Epidermal cell density is autoregulated via a secretory peptide,EPIDERMALPATTERNING FACTOR 2 in Arabidopsis leaves,Plant Cell Physiol.,50(6):1019-1031
Huang Y.,Tao Z.S.,Liu Q.,Wang X.F.,Yu J.Y.,Liu G.H.,and Wang H.Z.,2014,Bn EPFL6,an EPIDERMAL PATTERN-ING FACTOR-LIKE(EPFL)secreted peptide gene,is required for filament elongation in Brassica napus,Plant Mol.Biol.,85(4-5):505-517
Hunt L.,Bailey K.J.,and Gray J.E.,2010,The signalling peptide EPFL9 is a positive regulator of stomatal development,New Phytol.,186(3):609-614
Jewaria P.K.,Hara T.,Tanaka H.,Kondo T.,Betsuyaku S.,Sawa S.,Sakagami Y.,Aimoto S.,and Kakimoto T.,2013,Differential effect of peptides stomagen,EPF1 and EPF2 on activation of MAP kinase MPK6 and SPCH protein level,Plant Cell Physiol.,54(8):1253-1262
Jiang H.M.and Shi G.K.,2009,Advanced research on male sterility in wheat,Shengwu Jishu Tongbao(Biotechnology Bulletin),(10):30-33(江红梅,师光开,2009,小麦雄性不育研究进展,生物技术通报,(10):30-33)
Kimber G.,and Sears E.R.,1980,Uses of wheat aneuploids,In:Lewis W.H.,(ed.),Polyploidy,Plenum Press,New York,USA,pp.427-428
Kondo T.,Kajita R.,Miyazaki A.,Hokoyama M.,Nakamura-Miura T.,Mizuno S.,Masuda Y.,Irie K.,Tanaka Y.,Takada S.,Kakimoto T.,and Sakagami Y.,2010,Stomatal density is controlled by a mesophyll-derived signaling molecule,Plant Cell Physiol.,51(1):1-8
Livak K.J.,and Schmittgen T.D.,2001,Analysis of relative gene expression data using real time quantitative PCR and the2-ΔΔCtmethod,Methods,25(4):402-408
Marshall E.,Costa L.M.,and Gutierrez-Marcos J.,2011,Cysteine-rich peptides(CRPs)mediate diverse aspects of cell-cell communication in plant reproduction and development,J.Exp.Bot.,62(5):1677-1686
Meng X.Z.,Wang H.C.,He Y.X.,Liu Y.D.,Walker J.C.,Torii K.U.,and Zhang S.Q.,2012,A MAPK cascade downstream of ERECTA receptor-like protein kinase regulates Arabidopsis inflorescence architecture by promoting localized cell proliferation,Plant Cell,24(12):4948-4960
Murai K.,Miyamae M.,Kato H.,Takumi S.,and Ogihara Y.,2003,WAP1,a wheat APETALA1 homolog,plays a central role in the phase transition from vegetative to reproductive growth,Plant Cell Physiol.,44(12):1255-1265
Murai K.,Takumi S.,Koga H.,and Ogihara Y.,2002,Pistillody,homeotic transformation of stamens into pistil-like structures,caused by nuclear-cytoplasm interaction in wheat,Plant J.,29(2):169-181
Peng Z.S.,Yang Z.J.,Ouyang Z.M.and Yang H.,2013,Characterization of a novel pistillody mutant in common wheat,Aust.J.Crop Sci.,7(1):159-164
Sugano S.S.,Shimada T.,Imai Y.,Okawa K.,Tamai A.,Mori M.,and Hara-Nishimura I.,2010,Stomagen positively regulates stomatal density in Arabidopsis,Nature,463(7278):241-244
Takatsuji H.,Nakamura N.,and Katsumoto Y.,1994,A new family of zinc finger proteins in petunia:structure,DNA sequence recognition,and floral organ-specific expression,Plant Cell,6(7):947-958
Uchida N.,Lee J.S.,Horst R.J.,Lai H.H.,Kajita R.,Kakimoto T.,Tasaka M.,and Torii K.U.,2012,Regulation of inflorescence architecture by intertissue layer ligand-receptor communication between endodermis and phloem,Proc.Natl.A-cad.Sci.USA,109(16):6337-6342
Uchida N.,and Tasaka M.,2013,Regulation of plant vascular stem cells by endodermis-derived EPFL-family peptide hormones and phloem-expressed ERECTA-family receptor kinases,J.Exp.Bot.,64(17):5335-5343
Yamada K.,Saraike T.,Shitsukawa N.,Hirabayashi C.,Takumi S.,and Murai K.,2009,Class D and B(sister)MADS-box genes are associated with ectopic ovule formation in the pistil-like stamens of alloplasmic wheat(Triticum aestivum L.),Plant Mol.Biol.,71(1-2):1-14
Yang Z.J.,Peng Z.S.,Wei S.H.,Liao M.L.,Yu Y.,and Jang Z.Y.,2015,Pistillody mutant reveals key insights into stamen and pistil development in wheat(Triticum aestivum L.),BMC Genomics,16(1):211-220