玉米内源茉莉酸调节干旱胁迫下的光合作用和抗旱生理反应
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摘要
[目的]本文旨在探究玉米内源茉莉酸(JA)在抗旱反应中的作用及对内源脱落酸(ABA)合成的影响。[方法]以玉米自交系B73和茉莉酸合成缺少突变体opr7opr8(opr)为试验材料,进行土壤干旱处理和快速干旱处理。土壤干旱处理:生长至V7时期植株断水,13 d后复水为干旱处理;以保持土壤湿润为对照。处理后1、4、7、10、13、16、19 d取样。研究干旱胁迫对B73及opr苗期叶片净光合速率(P_n)、气孔导度(G_s)、胞间CO_2浓度(Ci)、蒸腾速率(T_r),总超氧化物歧化酶(T-SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)活性,丙二醛(MDA)、脯氨酸、可溶性蛋白含量的影响。快速干旱处理:切取V3时期幼苗地上部分,插入空试管为干旱处理;插入装满无菌水的试管为对照。各处理均置于28℃、相对湿度60%、光照强度为10 000 lx的生长箱中,分别于处理后0、1、3、6、9、12 h取样,采用RT-qPCR方法检测ABA合成通路关键基因Zm ZEP1、Zm NCED1和Zm AO1的表达情况。[结果]土壤干旱处理:对照中,opr的气孔密度小于B73,Ci值略低;干旱处理后,opr与B73的P_n、G_s和T_r值均下降,opr更敏感但下降速率慢。干旱7 d后,opr的净光合能力高于B73; T-SOD、POD和CAT活性呈现先上升后下降趋势,opr的酶活性下降时间晚于B73; MDA、可溶性蛋白和游离脯氨酸含量呈上升趋势,但opr的变化幅度小。植株快速干旱处理:opr和B73的Zm NCED1、Zm ZEP1和Zm AO1表达量在对照中无显著差异,干旱后均有上升,但opr中,Zm NCED1、Zm ZEP1及Zm AO1表达量低于B73。[结论]opr在土壤干旱条件下能够维持更高的叶片含水量、光合作用效率和抗氧化酶活性,细胞膜损伤较低,表明玉米内源JA在植物的抗旱反应中起重要作用,内源JA的缺少能够减少水分损失,提高在干旱条件下的生存能力。快速干旱条件下,内源JA的缺少降低了ABA合成途径中的3个关键酶活性,说明抗旱反应中ABA的合成被延缓,引起了ABA调控的植物抗旱反应延迟。
[Objectives]This paper is aimed at revealing the role of endogenous jasmonic acid( JA) in drought-tolerance response and its influence to endogenous abscisic acid( ABA) biosynthesis. [Methods]B73 and JA-deficient line opr7opr8( opr) were used to do soil drought and fast drought treatment. Soil drought treatment: plants were stopped watering for 13 days then re-watered as the drought treatment,and the plants watered routinely were used as the control. The leaves of the plants were sampled at 1,4,7,10,13,16 and 19 days of drought treatment period. Photosynthetic parameters such as net photosynthetic rate( P_n),stomatal conductance( Gs),intercellular carbon dioxide concentration( Ci),transpiration rate( Tr) were measured during the experiment time. The activity of antioxidant enzyme includes total-superoxide dismutase( T-SOD),peroxidase( POD),catalase( CAT),malondialdehyde( MDA)content and osmotic adjustment substances( proline and soluble protein) in leaves of the plants were detected. Fast drought treatment:the seedlings of B73 and opr were cutted off at V3 stage,and placed straightly in tubes with( control) or without water( dehydrate treatment). The detached seedlings were placed in a growth chamber where the condition maintained: temperature at 28 ℃,relative humidity at 60% and illumination at 10 000 lx. The leaves of the plants were sampled at 0,1,3,6,9 h after cut-off. The expression levels of three ABA biosynthesis genes,Zm ZEP1,Zm NCED1 and Zm AO1 were detected by quantitative PCR. [Results]Soil drought treatment: opr had lower stomatal density and Cithan B73 under normal condition. Under drought treatment,P_n,Gsand Trwent downboth in the opr and B73 leaves,but opr slower than B73,and the P_nof opr was higher than B73 at 7 day. The trend of T-SOD,POD and CAT activities rose up first and went down during drought tolerance. MDA,proline and soluble protein contents went up both in opr and B73,but had a minor variation in opr. Fast drought treatment: there were no significantly difference in the relative expression of gene Zm NCED1,Zm ZEP1 and Zm AO1 in control plants,but up-regulated under drought stress. Expression level of gene Zm NCED1,Zm AO1 and Zm ZEP1 in opr was lower than B73. [Conclusions]opr mutant has higher water content,photosynthetic efficiency,antioxidant enzyme activity but lower cellular membrane damage under drought treatment. It is indicated that maize endogenous JA plays an important role in drought response. The deficiency of endogenous JA in the mutant delays the drought response of the mutant plants. Under fast drought treatment,the absence of endogenous JA has delayed the activity of 3 key enzymes in the synthesis pathway of ABA,it indicates that the synthesis of ABA in drought response was delayed.
[Objectives]This paper is aimed at revealing the role of endogenous jasmonic acid( JA) in drought-tolerance response and its influence to endogenous abscisic acid( ABA) biosynthesis. [Methods]B73 and JA-deficient line opr7opr8( opr) were used to do soil drought and fast drought treatment. Soil drought treatment: plants were stopped watering for 13 days then re-watered as the drought treatment,and the plants watered routinely were used as the control. The leaves of the plants were sampled at 1,4,7,10,13,16 and 19 days of drought treatment period. Photosynthetic parameters such as net photosynthetic rate( P_n),stomatal conductance( Gs),intercellular carbon dioxide concentration( Ci),transpiration rate( Tr) were measured during the experiment time. The activity of antioxidant enzyme includes total-superoxide dismutase( T-SOD),peroxidase( POD),catalase( CAT),malondialdehyde( MDA)content and osmotic adjustment substances( proline and soluble protein) in leaves of the plants were detected. Fast drought treatment:the seedlings of B73 and opr were cutted off at V3 stage,and placed straightly in tubes with( control) or without water( dehydrate treatment). The detached seedlings were placed in a growth chamber where the condition maintained: temperature at 28 ℃,relative humidity at 60% and illumination at 10 000 lx. The leaves of the plants were sampled at 0,1,3,6,9 h after cut-off. The expression levels of three ABA biosynthesis genes,Zm ZEP1,Zm NCED1 and Zm AO1 were detected by quantitative PCR. [Results]Soil drought treatment: opr had lower stomatal density and Cithan B73 under normal condition. Under drought treatment,P_n,Gsand Trwent downboth in the opr and B73 leaves,but opr slower than B73,and the P_nof opr was higher than B73 at 7 day. The trend of T-SOD,POD and CAT activities rose up first and went down during drought tolerance. MDA,proline and soluble protein contents went up both in opr and B73,but had a minor variation in opr. Fast drought treatment: there were no significantly difference in the relative expression of gene Zm NCED1,Zm ZEP1 and Zm AO1 in control plants,but up-regulated under drought stress. Expression level of gene Zm NCED1,Zm AO1 and Zm ZEP1 in opr was lower than B73. [Conclusions]opr mutant has higher water content,photosynthetic efficiency,antioxidant enzyme activity but lower cellular membrane damage under drought treatment. It is indicated that maize endogenous JA plays an important role in drought response. The deficiency of endogenous JA in the mutant delays the drought response of the mutant plants. Under fast drought treatment,the absence of endogenous JA has delayed the activity of 3 key enzymes in the synthesis pathway of ABA,it indicates that the synthesis of ABA in drought response was delayed.
引文
[1]孙成韬,焦仁海,番兴明.玉米抗旱育种的研究进展[J].玉米科学,2006,14(6):71-74.Sun C T,Jiao R H,Pan X M. Advances of maize drought tolerance breeding[J]. Journal of Maize Sciences,2006,14(6):71-74(in Chinese with English abstract).
[2] Eberhard S,Finazzi G,Wollman F A. The dynamics of photosynthesis[J]. Annual Review of Genetics,2008,42(1):463-515.
[3] Reddy A R,Chaitanya K V,Vivekanandan M. Drought-induced responses of photosynthesis and antioxidant metabolism in higher plants[J].Journal of Plant Physiology,2004,161(11):1189-1202.
[4]张仁和,郑友军,马国胜,等.干旱胁迫对玉米苗期叶片光合作用和保护酶的影响[J].生态学报,2011,31(5):1303-1311.Zhang R H,Zheng Y J,Ma G S,et al. Effects of drought stress on photosynthetic traits and protective enzyme activity in maize seedling[J]. Acta Ecologica Sinica,2011,31(5):1303-1311(in Chinese with English abstract).
[5] Smirnoff N. The role of active oxygen in the response of plants to water deficit and desiccation[J]. New Phytologist,1993,125(1):27-58.
[6]高辉,张红芳,袁思安,等.植物内源激素对干旱胁迫的响应研究[J].绿色科技,2013(11):5-7.Gao H,Zhang H F,Yuan S A,et al. Response of plant endogenous hormone to drought threat stress[J]. Journal of Green Science and Technology,2013(11):5-7(in Chinese).
[7]沈慧,林琳,李群,等. ABA与SA信号通路交互作用的研究[C]//中国植物逆境生理生态与分子生物学学术研讨会.乌鲁木齐,2006.Shen H,Lin L,Li Q,et al. Study on the interaction between ABA and SA signaling pathway[C]//Symposium on physiological ecology and molecular biology of Chinese plants. Urumqi,2006(in Chinese).
[8]蔡昆争,董桃杏,徐涛.茉莉酸类物质(JAs)的生理特性及其在逆境胁迫中的抗性作用[J].生态环境学报,2006,15(2):397-404.Cai K Z,Dong T X,Xu T. The physiological roles and resistance control in stress environment of jasmonates[J]. Ecology and Environment,2006,15(2); 397-404(in Chinese with English abstract).
[9] Zhu J K. Salt and drought stress signal transduction in plants[J]. Annu Rev Plant Biol,2002,53:247-273.
[10] Finkelstein R. Abscisic acid synthesis and response[J]. The Arabidopsis Book,2013,11:e0166.
[11] Roychoudhury A,Paul S,Basu S. Cross-talk between abscisic acid-dependent and abscisic acid-independent pathways during abiotic stress[J].Plant Cell Rep,2013,32(7):985-1006.
[12]桂连友,刘树生,陈宗懋.外源茉莉酸和茉莉酸甲酯诱导植物抗虫作用及其机理[J].昆虫学报,2004,47(4):507-514.Gui L Y,Liu S S,Chen Z X. Plant resistance to insects induced by application of exogenous jasmonic acid and methyl jasmonate[J]. Acta Entomologica Sinica,2004,47(4):507-514(in Chinese with English abstract).
[13]王瑜,吴丽芳,余增亮.茉莉酸及其甲酯在植物诱导抗病性中的作用[J].生物学杂志,2000,17(1):11-12.Wang Y,Wu L F,Yu Z L. The role of jasmonic acid and methy jasmonate in plant induced disease resistance[J]. Journal of Biology,2000,17(1):11-12(in Chinese with English abstract).
[14]董桃杏,蔡昆争,张景欣,等.茉莉酸甲酯(MeJ A)对水稻幼苗的抗旱生理效应[J].生态环境,2007,16(4):1261-1265.Dong T X,Cai K Z,Zhang J X,et al. The physiological roles of methyl jasmonate(MeJ A)in drought resistance of rice seedlings[J]. Ecology and Environment,2007,16(4):1261-1265(in Chinese with English abstract).
[15] Dhakarey R,Raorane M L,Treumann A. Physiological and proteomic analysis of the rice mutant cpm2 suggests a negative regulatory role of jasmonic acid in drought tolerance[J]. Frontiers in Plant Science,2017,8:1903.
[16] Schaller F,Schaller A,Stintzi A. Biosynthesis and metabolism of jasmonates[J]. Journal of Plant Growth Regulation,2005,23(3):179-199.
[17] Ziegler J,Stenzel I,Hause B,et al. Molecular cloning of allene oxide cyclase:the enzyme establishing the stereochemistry of octadecanoids and jasmonates[J]. Journal of Biological Chemistry,2000,275(25):19132-19138.
[18] Schaller F,Biesgen C,Müssig C,et al. 12-oxophytodienoate reductase 3(OPR3)is the isoenzyme involved in jasmonate biosynthesis[J]. Planta,2000,210(6):979-984.
[19] Savchenko T,Kolla V A,Wang C Q. Functional convergence of oxylipin and abscisic acid pathways controls stomatal closure in response to drought[J]. Plant Physiology,2014,164(3):1151-1160.
[20] Zhang J,Simmons C,Yalpani N,et al. Genomic analysis of the 12-oxo-phytodienoic acid reductase gene family of Zea mays[J]. Plant Molecular Biology,2005,59(2):323-343.
[21] Yan Y C,Christensen S,Isakeit T,et al. Disruption of OPR7 and OPR8 reveals the versatile functions of jasmonic acid in maize development and defense[J]. Plant Cell,2012,24(4):1420-1436.
[22] Sedmak J J,Grossberg S E. A rapid,sensitive,and versatile assay for protein using coomassie brilliant blue G250[J]. Analytical Biochemistry,1977,79(1/2):544-552.
[23]徐俊增,彭世彰,魏征,等.节水灌溉水稻叶片胞间CO2浓度及气孔与非气孔限制[J].农业工程学报,2010,26(7):76-80.Xu J Z,Peng S Z,Wei Z,et al. Intercellular CO2concentration and stomatal or non-stomatal limitation of rice under water saving irrigation.[J].Transactions of the Chinese Society of Agricultural Engineering,2010,26(7):76-80(in Chinese with English abstract).
[24]刘海艳,杨丽洁,丁艳锋,等. NO对水稻孕穗期干旱胁迫下叶片光合及产量的影响[J].南京农业大学学报,2017,40(2):195-202.DOI:10.7685/jnau.201605012.Liu H Y,Yang L J,Ding Y F,et al. Effects of nitric oxide on photosynthesis and yield of rice under drought stress at booting stage[J]. Journal of Nanjing Agricultural University,2017,40(2):195-202(in Chinese with English abstract).
[25]马旭凤.水分亏缺对玉米生理指标、形态特性及解剖结构的影响[D].杨凌:西北农林科技大学,2010.Ma X F. Effect of water deficit on maize physiology,morphology and anatomical structure[D]. Yangling:Northwest A&F University,2010(in Chinese with English abstract).
[2] Eberhard S,Finazzi G,Wollman F A. The dynamics of photosynthesis[J]. Annual Review of Genetics,2008,42(1):463-515.
[3] Reddy A R,Chaitanya K V,Vivekanandan M. Drought-induced responses of photosynthesis and antioxidant metabolism in higher plants[J].Journal of Plant Physiology,2004,161(11):1189-1202.
[4]张仁和,郑友军,马国胜,等.干旱胁迫对玉米苗期叶片光合作用和保护酶的影响[J].生态学报,2011,31(5):1303-1311.Zhang R H,Zheng Y J,Ma G S,et al. Effects of drought stress on photosynthetic traits and protective enzyme activity in maize seedling[J]. Acta Ecologica Sinica,2011,31(5):1303-1311(in Chinese with English abstract).
[5] Smirnoff N. The role of active oxygen in the response of plants to water deficit and desiccation[J]. New Phytologist,1993,125(1):27-58.
[6]高辉,张红芳,袁思安,等.植物内源激素对干旱胁迫的响应研究[J].绿色科技,2013(11):5-7.Gao H,Zhang H F,Yuan S A,et al. Response of plant endogenous hormone to drought threat stress[J]. Journal of Green Science and Technology,2013(11):5-7(in Chinese).
[7]沈慧,林琳,李群,等. ABA与SA信号通路交互作用的研究[C]//中国植物逆境生理生态与分子生物学学术研讨会.乌鲁木齐,2006.Shen H,Lin L,Li Q,et al. Study on the interaction between ABA and SA signaling pathway[C]//Symposium on physiological ecology and molecular biology of Chinese plants. Urumqi,2006(in Chinese).
[8]蔡昆争,董桃杏,徐涛.茉莉酸类物质(JAs)的生理特性及其在逆境胁迫中的抗性作用[J].生态环境学报,2006,15(2):397-404.Cai K Z,Dong T X,Xu T. The physiological roles and resistance control in stress environment of jasmonates[J]. Ecology and Environment,2006,15(2); 397-404(in Chinese with English abstract).
[9] Zhu J K. Salt and drought stress signal transduction in plants[J]. Annu Rev Plant Biol,2002,53:247-273.
[10] Finkelstein R. Abscisic acid synthesis and response[J]. The Arabidopsis Book,2013,11:e0166.
[11] Roychoudhury A,Paul S,Basu S. Cross-talk between abscisic acid-dependent and abscisic acid-independent pathways during abiotic stress[J].Plant Cell Rep,2013,32(7):985-1006.
[12]桂连友,刘树生,陈宗懋.外源茉莉酸和茉莉酸甲酯诱导植物抗虫作用及其机理[J].昆虫学报,2004,47(4):507-514.Gui L Y,Liu S S,Chen Z X. Plant resistance to insects induced by application of exogenous jasmonic acid and methyl jasmonate[J]. Acta Entomologica Sinica,2004,47(4):507-514(in Chinese with English abstract).
[13]王瑜,吴丽芳,余增亮.茉莉酸及其甲酯在植物诱导抗病性中的作用[J].生物学杂志,2000,17(1):11-12.Wang Y,Wu L F,Yu Z L. The role of jasmonic acid and methy jasmonate in plant induced disease resistance[J]. Journal of Biology,2000,17(1):11-12(in Chinese with English abstract).
[14]董桃杏,蔡昆争,张景欣,等.茉莉酸甲酯(MeJ A)对水稻幼苗的抗旱生理效应[J].生态环境,2007,16(4):1261-1265.Dong T X,Cai K Z,Zhang J X,et al. The physiological roles of methyl jasmonate(MeJ A)in drought resistance of rice seedlings[J]. Ecology and Environment,2007,16(4):1261-1265(in Chinese with English abstract).
[15] Dhakarey R,Raorane M L,Treumann A. Physiological and proteomic analysis of the rice mutant cpm2 suggests a negative regulatory role of jasmonic acid in drought tolerance[J]. Frontiers in Plant Science,2017,8:1903.
[16] Schaller F,Schaller A,Stintzi A. Biosynthesis and metabolism of jasmonates[J]. Journal of Plant Growth Regulation,2005,23(3):179-199.
[17] Ziegler J,Stenzel I,Hause B,et al. Molecular cloning of allene oxide cyclase:the enzyme establishing the stereochemistry of octadecanoids and jasmonates[J]. Journal of Biological Chemistry,2000,275(25):19132-19138.
[18] Schaller F,Biesgen C,Müssig C,et al. 12-oxophytodienoate reductase 3(OPR3)is the isoenzyme involved in jasmonate biosynthesis[J]. Planta,2000,210(6):979-984.
[19] Savchenko T,Kolla V A,Wang C Q. Functional convergence of oxylipin and abscisic acid pathways controls stomatal closure in response to drought[J]. Plant Physiology,2014,164(3):1151-1160.
[20] Zhang J,Simmons C,Yalpani N,et al. Genomic analysis of the 12-oxo-phytodienoic acid reductase gene family of Zea mays[J]. Plant Molecular Biology,2005,59(2):323-343.
[21] Yan Y C,Christensen S,Isakeit T,et al. Disruption of OPR7 and OPR8 reveals the versatile functions of jasmonic acid in maize development and defense[J]. Plant Cell,2012,24(4):1420-1436.
[22] Sedmak J J,Grossberg S E. A rapid,sensitive,and versatile assay for protein using coomassie brilliant blue G250[J]. Analytical Biochemistry,1977,79(1/2):544-552.
[23]徐俊增,彭世彰,魏征,等.节水灌溉水稻叶片胞间CO2浓度及气孔与非气孔限制[J].农业工程学报,2010,26(7):76-80.Xu J Z,Peng S Z,Wei Z,et al. Intercellular CO2concentration and stomatal or non-stomatal limitation of rice under water saving irrigation.[J].Transactions of the Chinese Society of Agricultural Engineering,2010,26(7):76-80(in Chinese with English abstract).
[24]刘海艳,杨丽洁,丁艳锋,等. NO对水稻孕穗期干旱胁迫下叶片光合及产量的影响[J].南京农业大学学报,2017,40(2):195-202.DOI:10.7685/jnau.201605012.Liu H Y,Yang L J,Ding Y F,et al. Effects of nitric oxide on photosynthesis and yield of rice under drought stress at booting stage[J]. Journal of Nanjing Agricultural University,2017,40(2):195-202(in Chinese with English abstract).
[25]马旭凤.水分亏缺对玉米生理指标、形态特性及解剖结构的影响[D].杨凌:西北农林科技大学,2010.Ma X F. Effect of water deficit on maize physiology,morphology and anatomical structure[D]. Yangling:Northwest A&F University,2010(in Chinese with English abstract).