异源过表达水稻OsSAPP3基因促进拟南芥叶片衰老
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摘要
蛋白磷酸酶催化的蛋白质可逆磷酸化反应是叶片衰老的关键环节。该研究筛选并克隆了1个新的参与水稻(Oryza sativa)叶片衰老调控的PP2C基因OsSAPP3。研究表明,OsSAPP3的启动子在Pro_(OsSAPP3)-GUS转基因拟南芥(Arabidopsis thaliana)的莲座叶中有活性,并且活性以依赖叶龄方式增加。利用CaMV 35S启动子驱动组成型异源过表达OsSAPP3导致转基因拟南芥无法正常生长。用可诱导型启动子GVG系统驱动OsSAPP3异源过表达导致转基因拟南芥出现莲座叶变小、数量增加、叶片早衰及抽薹开花提前等早衰表型。外源诱导OsSAPP3基因异源过表达后,利用实时荧光定量PCR检测到SAG12、WRKY6和NAC2等衰老标志基因显著上调表达。研究结果表明, OsSAPP3是参与水稻叶片衰老的正向调控因子。
Protein phosphatase-catalyzed reversible phosphorylation of proteins is a key aspect of leaf senescence. We screened and cloned a new PP2C gene, Os SAPP3, involved in leaf senescence regulation in rice. The promoter of OsSAPP3 was active in rosette leaves of Pro OsSAPP3-GUS transgenic Arabidopsis and its activity increased with leaf age. With heterologous overexpression of OsSAPP3 driven by a CaMV 35S promoter, transgenic Arabidopsis could not grow normally. Therefore, OsSAPP3 heterologous overexpression was induced by the inducible promoter GVG system. Exogenous inducible heterologous overexpression of OsSAPP3 led to a precocious leaf senescence phenotype in transgenic Arabidopsis including smaller rosette leaves and increased number, premature leaf senescence, and advanced bolting and flowering. Additionally, quantitative RT-PCR showed that exogenous inducible heterologous overexpression of OsSAPP3 upregulated the expression of critical senescence-related transcription factor genes, including SAG12, WRKY6, and NAC2. In summary, OsSAPP3 is a positive factor involved in regulating leaf senescence in rice.
Protein phosphatase-catalyzed reversible phosphorylation of proteins is a key aspect of leaf senescence. We screened and cloned a new PP2C gene, Os SAPP3, involved in leaf senescence regulation in rice. The promoter of OsSAPP3 was active in rosette leaves of Pro OsSAPP3-GUS transgenic Arabidopsis and its activity increased with leaf age. With heterologous overexpression of OsSAPP3 driven by a CaMV 35S promoter, transgenic Arabidopsis could not grow normally. Therefore, OsSAPP3 heterologous overexpression was induced by the inducible promoter GVG system. Exogenous inducible heterologous overexpression of OsSAPP3 led to a precocious leaf senescence phenotype in transgenic Arabidopsis including smaller rosette leaves and increased number, premature leaf senescence, and advanced bolting and flowering. Additionally, quantitative RT-PCR showed that exogenous inducible heterologous overexpression of OsSAPP3 upregulated the expression of critical senescence-related transcription factor genes, including SAG12, WRKY6, and NAC2. In summary, OsSAPP3 is a positive factor involved in regulating leaf senescence in rice.
引文
李合生(2000).植物生理生化实验原理和技术.北京:高等教育出版社.pp.134-137.
牟少亮,李秀娟,官德义,赖燕,何水林(2011).水稻蛋白磷酸酶ABI2基因启动子的克隆和功能分析.热带作物学报32,2293-2297.
孙高阳,吴向远,叶琳琳,孟淑君,阎鹏帅,汤继华,郭战勇(2017).玉米衰老相关基因在2个杂交种及其亲本中的表达分析.河南农业大学学报2,140-148.
翁华,冉亮,魏群(2003).植物蛋白磷酸酶及其在植物抗逆中的作用.植物学报20,609-615.
肖冬,崔燕娇,王宁宁(2014).叶片衰老过程中的蛋白激酶和蛋白磷酸酶.植物生理学报50,1267-1273.
徐凡(2012).GmSARK和AtSARK基因调控叶片衰老分子机制的研究.博士论文.天津:南开大学.pp.1-12.
徐凡,李鹏丽,安宝燕,苑玲玲,孟涛,岳慧琴,王宁宁(2010).大豆诱导型启动子驱动类受体蛋白激酶Gm SARK转基因植物分析.植物学报45,149-156.
徐娜,徐江民,蒋玲欢,饶玉春(2017).水稻叶片早衰成因及分子机理研究进展.植物学报52,102-112.
杨同文,李成伟(2014).植物叶片衰老的表观遗传调控.植物学报49,729-737.
郑建敏,张涛,郑家奎(2009).水稻叶片衰老相关基因的研究进展.基因组学与应用生物学28,1010-1019.
Aoyama T,Chua NH(1997).A glucocorticoid-mediated transcriptional induction system in transgenic plants.Plant J 11,605-612.
Bhatnagar N,Min MK,Choi EH,Kim N,Moon SJ,Yoon I,Kwon T,Junq KH,Kim BG(2017).The protein phosphatase 2C clade a protein OsPP2C51 positively regulates seed germination by directly inactivating Osb ZIP10.Plant Mol Biol 93,389-401.
Boyes DC,Zayed AM,Ascenzi R,McCaskill AJ,Hoffman NE,Davis KR,G?rlach J(2001).Growth stage-based phenotypic analysis of Arabidopsis:a model for high throughput functional genomics in plants.Plant Cell 13,1499-1510.
Buchanan-Wollaston V,Page T,Harrison E,Breeze E,Lim PO,Nam HG,Lin JF,Wu SH,Swidzinski J,Ishizaki K,Leaver CJ(2005).Comparative transcriptome analysis reveals significant differences in gene expression and signaling pathways between developmental and dark/starvation-induced senescence in Arabidopsis.Plant J 42,567-585.
Castro PH,Lilay GH,Mu?oz-Mérida A,Schjoerring JK,Azevedo H,Assun??o AGL(2017).Phylogenetic analysis of F-bZIP transcription factors indicates conservation of the zinc deficiency response across land plants.Sci Rep 7,3806.
Chen C,Yu Y,Ding XD,Liu BD,Duanmu H,Zhu D,Sun XL,Cao L,Zaib-Un-Nisa,Li Q,Zhu YM(2018).Genome-wide analysis and expression profiling of PP2Cclade D under saline and alkali stresses in wild soybean and Arabidopsis.Protoplasma 255,643-654.
Jiang YJ,Liang G,Yang SZ,Yu DQ(2014).Arabidopsis WRKY57 functions as a node of convergence for jasmonic acid-and auxin-mediated signaling in jasmonic acidinduced leaf senescence.Plant Cell 26,230-245.
Lim PO,Hong GN(2007).Aging and senescence of the leaf organ.J Plant Biol 50,291-300.
Lim PO,Woo HR,Hong GN(2003).Molecular genetics of leaf senescence in Arabidopsis.Trends Plant Sci 8,272-278.
Liu D,Gong QQ,Ma YY,Li PL,Li JP,Yang SH,Yuan LL,Yu YQ,Pan DD,Xu F,Wang NN(2010).cp Sec A,a thylakoid protein translocase subunit,is essential for photosynthetic development in Arabidopsis.J Exp Bot 61,1655-1669.
Liu L,Xu W,Hu XS,Liu HJ,Lin YJ(2016).W-box and G-box elements play important roles in early senescence of rice flag leaf.Sci Rep 6,20881.
Menges M,Hennig L,Gruissem W,Murray JAH(2002).Cell cycle-regulated gene expression in Arabidopsis.J Biol Chem 277,41987-42002.
Quirino BF,Noh YS,Himelblau E,Amasino RM(2000).Molecular aspects of leaf senescence.Trends Plant Sci 5,278-282.
Robatzek S,Somssich IE(2002).Targets of AtWRKY6regulation during plant senescence and pathogen defense.Genes Dev 16,1139-1149.
Sakuraba Y,Schelbert S,Park SY,Han SH,Lee BD,Andrès CB,Kessler F,H?rtensteiner S,Paek NC(2012).STAY-GREEN and chlorophyll catabolic enzymes interact at light-harvesting complex ii for chlorophyll detoxification during leaf senescence in Arabidopsis.Plant Cell 24,507-518.
Schweighofer A,Kazanaviciute V,Scheikl E,Teige M,Doczi R,Hirt H,Schwanninger M,Kant M,Schuurink R,Mauch F,Buchala A,Cardinale F,Meskiene I(2007).The PP2C-type phosphatase AP2C1,which negatively regulates MPK4 and MPK6,modulates innate immunity,jasmonic acid,and ethylene levels in Arabidopsis.Plant Cell 19,2213-2224.
Shen XJ,Guo X,Zhao D,Zhang Q,Jiang YZ,Wang YT,Peng X,Wei Y,Zhai ZF,Zhao W,Li TH(2017).Cloning and expression profiling of the PacSnRK2 and PacPP2Cgene families during fruit development,ABA treatment,and dehydration stress in sweet cherry.Plant Physiol Biochem 119,275-285.
Wang CL,Lu GQ,Hao YQ,Guo HM,Guo Y,Zhao J,Cheng HM(2017).ABP9,a maize bZIP transcription factor,enhances tolerance to salt and drought in transgenic cotton.Planta 246,453-469.
Wang FB,Liu JC,Chen MX,Zhou LJ,Li ZW,Zhao Q,Pan G,Zaidi SHR,Cheng FM(2016).Involvement of abscisic acid in PSII photodamage and D1 protein turnover for light-induced premature senescence of rice flag leaves.PLo S One 11,e0161203.
Xiao D,Cui YJ,Xu F,Xu XX,Gao GX,Wang YX,Guo ZX,Wang D,Wang NN(2015).Senescence-suppressed protein phosphatase directly interacts with the cytoplasmic domain of senescence-associated receptor-like kinase and negatively regulates leaf senescence in Arabidopsis.Plant Physiol 169,1275-1291.
Xu F,Meng T,Li PL,Yu YQ,Cui YJ,Wang YX,Gong QQ,Wang NN(2011).A soybean dual-specificity kinase,Gm SARK,and its Arabidopsis homolog,AtSARK,regulate leaf senescence through synergistic actions of auxin and ethylene.Plant Physiol 157,2131-2153.
Xue TT,Wang D,Zhang SZ,Ehlting J,Ni F,Jakab S,Zheng CC,Zhong Y(2008).Genome-wide and expression analysis of protein phosphatase 2C in rice and Arabidopsis.BMC Genomics 9,550.
Zhang KW,Gan SS(2012).An abscisic acid-AtNAP transcription factor-SAG113 protein phosphatase 2C regulatory chain for controlling dehydration in senescing Arabidopsis leaves.Plant Physiol 158,961-969.
Zhao Y,Chan ZL,Gao JH,Xing L,Cao MJ,Yu CM,Hu YL,You J,Shi HT,Zhu YF,Gong YH,Mu ZX,Wang HQ,Deng X,Wang PC,Bressan RA,Zhu JK(2016).ABAreceptor PYL9 promotes drought resistance and leaf senescence.Proc Natl Acad Sci USA 113,1949-1954.
牟少亮,李秀娟,官德义,赖燕,何水林(2011).水稻蛋白磷酸酶ABI2基因启动子的克隆和功能分析.热带作物学报32,2293-2297.
孙高阳,吴向远,叶琳琳,孟淑君,阎鹏帅,汤继华,郭战勇(2017).玉米衰老相关基因在2个杂交种及其亲本中的表达分析.河南农业大学学报2,140-148.
翁华,冉亮,魏群(2003).植物蛋白磷酸酶及其在植物抗逆中的作用.植物学报20,609-615.
肖冬,崔燕娇,王宁宁(2014).叶片衰老过程中的蛋白激酶和蛋白磷酸酶.植物生理学报50,1267-1273.
徐凡(2012).GmSARK和AtSARK基因调控叶片衰老分子机制的研究.博士论文.天津:南开大学.pp.1-12.
徐凡,李鹏丽,安宝燕,苑玲玲,孟涛,岳慧琴,王宁宁(2010).大豆诱导型启动子驱动类受体蛋白激酶Gm SARK转基因植物分析.植物学报45,149-156.
徐娜,徐江民,蒋玲欢,饶玉春(2017).水稻叶片早衰成因及分子机理研究进展.植物学报52,102-112.
杨同文,李成伟(2014).植物叶片衰老的表观遗传调控.植物学报49,729-737.
郑建敏,张涛,郑家奎(2009).水稻叶片衰老相关基因的研究进展.基因组学与应用生物学28,1010-1019.
Aoyama T,Chua NH(1997).A glucocorticoid-mediated transcriptional induction system in transgenic plants.Plant J 11,605-612.
Bhatnagar N,Min MK,Choi EH,Kim N,Moon SJ,Yoon I,Kwon T,Junq KH,Kim BG(2017).The protein phosphatase 2C clade a protein OsPP2C51 positively regulates seed germination by directly inactivating Osb ZIP10.Plant Mol Biol 93,389-401.
Boyes DC,Zayed AM,Ascenzi R,McCaskill AJ,Hoffman NE,Davis KR,G?rlach J(2001).Growth stage-based phenotypic analysis of Arabidopsis:a model for high throughput functional genomics in plants.Plant Cell 13,1499-1510.
Buchanan-Wollaston V,Page T,Harrison E,Breeze E,Lim PO,Nam HG,Lin JF,Wu SH,Swidzinski J,Ishizaki K,Leaver CJ(2005).Comparative transcriptome analysis reveals significant differences in gene expression and signaling pathways between developmental and dark/starvation-induced senescence in Arabidopsis.Plant J 42,567-585.
Castro PH,Lilay GH,Mu?oz-Mérida A,Schjoerring JK,Azevedo H,Assun??o AGL(2017).Phylogenetic analysis of F-bZIP transcription factors indicates conservation of the zinc deficiency response across land plants.Sci Rep 7,3806.
Chen C,Yu Y,Ding XD,Liu BD,Duanmu H,Zhu D,Sun XL,Cao L,Zaib-Un-Nisa,Li Q,Zhu YM(2018).Genome-wide analysis and expression profiling of PP2Cclade D under saline and alkali stresses in wild soybean and Arabidopsis.Protoplasma 255,643-654.
Jiang YJ,Liang G,Yang SZ,Yu DQ(2014).Arabidopsis WRKY57 functions as a node of convergence for jasmonic acid-and auxin-mediated signaling in jasmonic acidinduced leaf senescence.Plant Cell 26,230-245.
Lim PO,Hong GN(2007).Aging and senescence of the leaf organ.J Plant Biol 50,291-300.
Lim PO,Woo HR,Hong GN(2003).Molecular genetics of leaf senescence in Arabidopsis.Trends Plant Sci 8,272-278.
Liu D,Gong QQ,Ma YY,Li PL,Li JP,Yang SH,Yuan LL,Yu YQ,Pan DD,Xu F,Wang NN(2010).cp Sec A,a thylakoid protein translocase subunit,is essential for photosynthetic development in Arabidopsis.J Exp Bot 61,1655-1669.
Liu L,Xu W,Hu XS,Liu HJ,Lin YJ(2016).W-box and G-box elements play important roles in early senescence of rice flag leaf.Sci Rep 6,20881.
Menges M,Hennig L,Gruissem W,Murray JAH(2002).Cell cycle-regulated gene expression in Arabidopsis.J Biol Chem 277,41987-42002.
Quirino BF,Noh YS,Himelblau E,Amasino RM(2000).Molecular aspects of leaf senescence.Trends Plant Sci 5,278-282.
Robatzek S,Somssich IE(2002).Targets of AtWRKY6regulation during plant senescence and pathogen defense.Genes Dev 16,1139-1149.
Sakuraba Y,Schelbert S,Park SY,Han SH,Lee BD,Andrès CB,Kessler F,H?rtensteiner S,Paek NC(2012).STAY-GREEN and chlorophyll catabolic enzymes interact at light-harvesting complex ii for chlorophyll detoxification during leaf senescence in Arabidopsis.Plant Cell 24,507-518.
Schweighofer A,Kazanaviciute V,Scheikl E,Teige M,Doczi R,Hirt H,Schwanninger M,Kant M,Schuurink R,Mauch F,Buchala A,Cardinale F,Meskiene I(2007).The PP2C-type phosphatase AP2C1,which negatively regulates MPK4 and MPK6,modulates innate immunity,jasmonic acid,and ethylene levels in Arabidopsis.Plant Cell 19,2213-2224.
Shen XJ,Guo X,Zhao D,Zhang Q,Jiang YZ,Wang YT,Peng X,Wei Y,Zhai ZF,Zhao W,Li TH(2017).Cloning and expression profiling of the PacSnRK2 and PacPP2Cgene families during fruit development,ABA treatment,and dehydration stress in sweet cherry.Plant Physiol Biochem 119,275-285.
Wang CL,Lu GQ,Hao YQ,Guo HM,Guo Y,Zhao J,Cheng HM(2017).ABP9,a maize bZIP transcription factor,enhances tolerance to salt and drought in transgenic cotton.Planta 246,453-469.
Wang FB,Liu JC,Chen MX,Zhou LJ,Li ZW,Zhao Q,Pan G,Zaidi SHR,Cheng FM(2016).Involvement of abscisic acid in PSII photodamage and D1 protein turnover for light-induced premature senescence of rice flag leaves.PLo S One 11,e0161203.
Xiao D,Cui YJ,Xu F,Xu XX,Gao GX,Wang YX,Guo ZX,Wang D,Wang NN(2015).Senescence-suppressed protein phosphatase directly interacts with the cytoplasmic domain of senescence-associated receptor-like kinase and negatively regulates leaf senescence in Arabidopsis.Plant Physiol 169,1275-1291.
Xu F,Meng T,Li PL,Yu YQ,Cui YJ,Wang YX,Gong QQ,Wang NN(2011).A soybean dual-specificity kinase,Gm SARK,and its Arabidopsis homolog,AtSARK,regulate leaf senescence through synergistic actions of auxin and ethylene.Plant Physiol 157,2131-2153.
Xue TT,Wang D,Zhang SZ,Ehlting J,Ni F,Jakab S,Zheng CC,Zhong Y(2008).Genome-wide and expression analysis of protein phosphatase 2C in rice and Arabidopsis.BMC Genomics 9,550.
Zhang KW,Gan SS(2012).An abscisic acid-AtNAP transcription factor-SAG113 protein phosphatase 2C regulatory chain for controlling dehydration in senescing Arabidopsis leaves.Plant Physiol 158,961-969.
Zhao Y,Chan ZL,Gao JH,Xing L,Cao MJ,Yu CM,Hu YL,You J,Shi HT,Zhu YF,Gong YH,Mu ZX,Wang HQ,Deng X,Wang PC,Bressan RA,Zhu JK(2016).ABAreceptor PYL9 promotes drought resistance and leaf senescence.Proc Natl Acad Sci USA 113,1949-1954.
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