脱落酸调节长春花中单萜吲哚生物碱的生物合成
|
摘要
利用病毒诱导的基因沉默技术沉默长春花幼苗中ABA生物合成途径基因八氢番茄红素脱氢酶(Cr PDS)的表达,控制内源ABA的含量,并对其进行外源ABA处理,从而研究脱落酸(abscisic acid, ABA)对长春花中单萜吲哚生物碱(monoterpenoid indole alkaloids, MIAs)生物合成的影响。Real-time PCR结果显示,在ABA处理后,单萜吲哚生物碱生物合成基因CrTDC、CrNMT和CrD4H表达量显著上升。进一步实验表明,ABA生物合成途径基因CrPDS的沉默能够降低内源ABA水平,并通过下调单萜吲哚生物碱生物合成途径基因降低长春质碱和文朵灵的积累。外源ABA处理能够恢复这些基因的正常表达,由此确定单萜吲哚生物碱生物合成基因CrTDC、CrNMT和CrD4H能够响应ABA信号,并且在ABA介导的长春花单萜吲哚生物碱生物合成调控中发挥重要作用。
In order to examine the effect of abscisic acid(ABA) on monoterpenoid indole alkaloids(MIAs) biosynthesis in C. roseus, the expression of ABA biosynthesis gene CrPDS in C. roseus seedlings was silenced by virus induced gene silencing, the content of endogenous ABA was controlled and exogenous ABA treatment was applied.Real-time PCR results showed that the expression levels of three MIA biosynthesis genes, CrTDC, CrNMT, and CrD4 H, obviously increased after ABA treatment. Further experiments showed that silencing of ABA biosynthesis gene CrPDS could reduce endogenous ABA levels. The accumulation of catharanthine and vindoline was decreased through down-regulating MIA biosynthesis genes. Normal expression of these genes could be restored by exogenous ABA treatment, which helped us identify that MIA biosynthesis gene, CrTDC, CrNMT, and CrD4 H, could respond to ABA signal and play an important role in ABA-mediated regulation of MIAs biosynthesis in C. roseus.
In order to examine the effect of abscisic acid(ABA) on monoterpenoid indole alkaloids(MIAs) biosynthesis in C. roseus, the expression of ABA biosynthesis gene CrPDS in C. roseus seedlings was silenced by virus induced gene silencing, the content of endogenous ABA was controlled and exogenous ABA treatment was applied.Real-time PCR results showed that the expression levels of three MIA biosynthesis genes, CrTDC, CrNMT, and CrD4 H, obviously increased after ABA treatment. Further experiments showed that silencing of ABA biosynthesis gene CrPDS could reduce endogenous ABA levels. The accumulation of catharanthine and vindoline was decreased through down-regulating MIA biosynthesis genes. Normal expression of these genes could be restored by exogenous ABA treatment, which helped us identify that MIA biosynthesis gene, CrTDC, CrNMT, and CrD4 H, could respond to ABA signal and play an important role in ABA-mediated regulation of MIAs biosynthesis in C. roseus.
引文
De Luca V.,Salim V.,Thamm A.,Masada S.A.,and Yu F.,2014,Making iridoids/secoiridoids and monoterpenoid indole alkaloids:progress on pathway elucidation,Curr.Opin.Plant Biol.,19:35-42
El-Sayed M.,and Verpoorte R.,2004,Growth,metabolic profiling and enzymes activities of Catharanthus roseus seedlings treated with plant growth regulators,Plant Growth Regul.,44(1):53-58
Niu Y.L.,Jiang X.M.,and Xu X.Y.,2016,Cloning and VIGSvector construction of GRAS transcription factors family gene SIGLD1 from tomato,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),35(8):2155-2160(牛义岭,姜秀明,许向阳,2016,番茄GRAS转录因子家族基因SIGLD1的克隆及VIGS载体构建,基因组学与应用生物学,35(8):2155-2160)
Pan Q.F.,Chen Y.,Wang Q.,Yuan F.,Xing S.H.,Tian Y.S.,Zhao J.Y.,Sun X.F.,and Tang K.X.,2010,Effect of plant growth regulators on the biosynthesis of vinblastine,vindoline and catharanthine in Catharanthus roseus,Plant Growth Regul.,60(2):133-141
Pan Q.F.,Mustafa N.R.,Tang K.X.,Choi Y.H.,and Verpoorte R.,2016,Monoterpenoid indole alkaloids biosynthesis and its regulation in Catharanthus roseus:a literature review from genes to metabolites,Phytochem.Rev.,15(2):221-250
Papon N.,Bremer J.,Vansiri A.,Andreu F.,Rideau M.,and Creche J.,2005,Cytokinin and ethylene control indole alkaloid production at the level of the MEP/terpenoid pathways in Catharanthus roseus suspension cells,Planta Med.,71(6):572-574
Pasquali G.,Goddijn O.J.M.,De Waal A.,Verpoorte R.,Schilperoort R.A.,Hoge J.H.C.,and Memelink J.,1992,Coordinated regulation of two indole alkaloid biosynthetic genes from Catharanthus roseus by auxin and elicitors,Plant Mol.Biol.,18(6):1121-1131
Pre M.,Atallah M.,Champion A.,De Vos M.,Pieterse C.M.J.,and Memelink J.,2008,The AP2/ERF domain transcription factor ORA59 integrates jasmonic acid and ethylene signals in plant defense,Plant Physiol.,147(3):1347-1357
Qu Y.,Easson M.L.A.E.,Froese J.,Simionescu R.,Hudlicky T.,and De Luca V.,2015,Completion of the seven-step pathway from tabersonine to the anticancer drug precursor vindoline and its assembly in yeast,Proc.Natl.Acad.Sci.USA,112(19):6224-6229
Roepke J.,Salim V.,Wu M.,Thamm A.M.,Murata J.,Ploss K.,Boland W.,and De Luca V.,2010,Vinca drug components accumulate exclusively in leaf exudates of Madagascar periwinkle,Proc.Natl.Acad.Sci.USA,107(34):15287-15292
Smith J.I.,Smart N.J.,Kurz W.G.W.,and Misawa M.,1987,Stimulation of indole alkaloid production in cell-suspensions cultures of Catharanthus roseus by abscisic acid,Planta Med.,53:470-474
Van der Fits L.,and Memelink J.,2000,ORCA3,a jasmonate-responsive transcriptional regulator of plant primary and secondary metabolism,Science,289(5477):295-297
Wasilewska A.,Vlad F.,Sirichandra C.,Redko Y.,Jammes F.,Valon C.,Frei dit Frey N.,and Leung J.,2008,An update on abscisic acid signaling in plants and more…,Molecular Plant,1(2):198-217
Xiong L.M.,and Zhu J.K.,2003,Regulation of abscisic acid biosynthesis,Plant Physiol.,133(1):29-36
Yu F.,and De Luca V.,2013,ATP-binding cassette transporter controls leaf surface secretion of anticancer drug components in Catharanthus roseus,Proc.Natl.Acad.Sci.USA,110(39):15830-15835
Zhao J.,Zhu W.H.,and Hu Q.,2001,Effects of light and plant growth regulators on the biosynthesis of vindoline and other indole alkaloids in Catharanthus roseus callus cultures,Plant Growth Regul.,33(1):43-49
Zhou M.L.,Zhu X.M.,Shao J.R.,Wu Y.M.,and Tang Y.X.,2010,Transcriptional response of the catharanthine biosynthesis pathway to methyl jasmonate/nitric oxide elicitation in Catharanthus roseus hairy root culture,Appl.Microbiol.Biotechnol.,88(3):737-750
El-Sayed M.,and Verpoorte R.,2004,Growth,metabolic profiling and enzymes activities of Catharanthus roseus seedlings treated with plant growth regulators,Plant Growth Regul.,44(1):53-58
Niu Y.L.,Jiang X.M.,and Xu X.Y.,2016,Cloning and VIGSvector construction of GRAS transcription factors family gene SIGLD1 from tomato,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),35(8):2155-2160(牛义岭,姜秀明,许向阳,2016,番茄GRAS转录因子家族基因SIGLD1的克隆及VIGS载体构建,基因组学与应用生物学,35(8):2155-2160)
Pan Q.F.,Chen Y.,Wang Q.,Yuan F.,Xing S.H.,Tian Y.S.,Zhao J.Y.,Sun X.F.,and Tang K.X.,2010,Effect of plant growth regulators on the biosynthesis of vinblastine,vindoline and catharanthine in Catharanthus roseus,Plant Growth Regul.,60(2):133-141
Pan Q.F.,Mustafa N.R.,Tang K.X.,Choi Y.H.,and Verpoorte R.,2016,Monoterpenoid indole alkaloids biosynthesis and its regulation in Catharanthus roseus:a literature review from genes to metabolites,Phytochem.Rev.,15(2):221-250
Papon N.,Bremer J.,Vansiri A.,Andreu F.,Rideau M.,and Creche J.,2005,Cytokinin and ethylene control indole alkaloid production at the level of the MEP/terpenoid pathways in Catharanthus roseus suspension cells,Planta Med.,71(6):572-574
Pasquali G.,Goddijn O.J.M.,De Waal A.,Verpoorte R.,Schilperoort R.A.,Hoge J.H.C.,and Memelink J.,1992,Coordinated regulation of two indole alkaloid biosynthetic genes from Catharanthus roseus by auxin and elicitors,Plant Mol.Biol.,18(6):1121-1131
Pre M.,Atallah M.,Champion A.,De Vos M.,Pieterse C.M.J.,and Memelink J.,2008,The AP2/ERF domain transcription factor ORA59 integrates jasmonic acid and ethylene signals in plant defense,Plant Physiol.,147(3):1347-1357
Qu Y.,Easson M.L.A.E.,Froese J.,Simionescu R.,Hudlicky T.,and De Luca V.,2015,Completion of the seven-step pathway from tabersonine to the anticancer drug precursor vindoline and its assembly in yeast,Proc.Natl.Acad.Sci.USA,112(19):6224-6229
Roepke J.,Salim V.,Wu M.,Thamm A.M.,Murata J.,Ploss K.,Boland W.,and De Luca V.,2010,Vinca drug components accumulate exclusively in leaf exudates of Madagascar periwinkle,Proc.Natl.Acad.Sci.USA,107(34):15287-15292
Smith J.I.,Smart N.J.,Kurz W.G.W.,and Misawa M.,1987,Stimulation of indole alkaloid production in cell-suspensions cultures of Catharanthus roseus by abscisic acid,Planta Med.,53:470-474
Van der Fits L.,and Memelink J.,2000,ORCA3,a jasmonate-responsive transcriptional regulator of plant primary and secondary metabolism,Science,289(5477):295-297
Wasilewska A.,Vlad F.,Sirichandra C.,Redko Y.,Jammes F.,Valon C.,Frei dit Frey N.,and Leung J.,2008,An update on abscisic acid signaling in plants and more…,Molecular Plant,1(2):198-217
Xiong L.M.,and Zhu J.K.,2003,Regulation of abscisic acid biosynthesis,Plant Physiol.,133(1):29-36
Yu F.,and De Luca V.,2013,ATP-binding cassette transporter controls leaf surface secretion of anticancer drug components in Catharanthus roseus,Proc.Natl.Acad.Sci.USA,110(39):15830-15835
Zhao J.,Zhu W.H.,and Hu Q.,2001,Effects of light and plant growth regulators on the biosynthesis of vindoline and other indole alkaloids in Catharanthus roseus callus cultures,Plant Growth Regul.,33(1):43-49
Zhou M.L.,Zhu X.M.,Shao J.R.,Wu Y.M.,and Tang Y.X.,2010,Transcriptional response of the catharanthine biosynthesis pathway to methyl jasmonate/nitric oxide elicitation in Catharanthus roseus hairy root culture,Appl.Microbiol.Biotechnol.,88(3):737-750