低温下不同H_2O_2抑制剂对四季秋海棠花色素苷合成的影响
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摘要
为了确定不同H_2O_2抑制剂对低温诱导植物花色素苷合成的影响,探讨此过程中不同来源H_2O_2的作用途径和机制,以四季秋海棠为研究试材,采用酶活性分析和qRT-PCR等方法,测定了不同处理下SOD、POD、PAO、GO酶的活性和H_2O_2、MDA的含量,以及花色素苷合成相关酶基因的表达水平。结果表明:低温显著增加了四季秋海棠叶片的SOD、POD、PAO、GO酶活性,促进了H_2O_2的积累和MDA含量的升高,最终引起花色素苷合成相关酶基因(BsPAL、BsCHS、BsF3H、BsANS)表达上调和花色素苷含量的增加;施用光合电子传递链的抑制剂DCMU、SOD的抑制剂DDC、GO酶的抑制剂OX和NADPH氧化酶的抑制剂DPI预处理6 h再给予低温处理,均不同程度地降低了SOD、POD、PAO、GO的活性和H_2O_2、MDA的含量,花色素苷合成相关酶基因表达水平下降,导致花色素苷合成受阻。因此,不同抑制剂预处理,分别降低由光合电子传递链、SOD、GO、NADPH氧化酶介导产生的H_2O_2后,均不同程度地减少了花色素苷含量,表明光合电子传递链、SOD、GO和NADPH氧化酶介导产生的H_2O_2可能参与了低温诱导四季秋海棠叶片花色素苷合成的过程。
To analyze the effects of different H_2O_2 inhibitors on anthocyanin biosynthesis induced by low temperature,to discuss the function principle and function way of different originations of H_2O_2 produced in this process,the activity of SOD,POD,PAO and GO enzymes,the content of H_2O_2 and MDA,and the transcription levels of anthocyanin biosynthesis genes were measured under different treatments in Begonia semperflorens,using the methods of enzyme-activity analysis and qRT-PCR,etc..The results showed that:The activities of SOD,POD,PAO and GO were significantly increased by low temperature,causing H_2O_2 accumulation and an increase in MDA content in Beginia semperflorens.The transcription levels of anthocyanin biosynthesis genes(namely BsPAL,BsCHS,BsF3 H and BsANS) and anthocyanin content were consequently soared in Beginia semperflorens;After the pretreatment with DCMU(an inhibitor of photosynthetic electron transport),DDC(an inhibitor of SOD),OX(an inhibitor of GO) and DPI(an inhibitor ofNADPH oxidase) for 6 hours and then treated with low temperature,in different degrees,the activities of SOD,POD,PAO and GO all decreased,the contents of H_2O_2 and MDA decreased either;blocked partially anthocyanin biosynthesis by decreasing the transcription levels of anthocyanin biosynthesis genes.As a result,pretreatments of all the test inhibitors decreased the contents of H_2O_2 came from photosynthetic electron transport,SOD,GO and NADPH oxidase;and anthocyanin content was also decreased to different level in Beginia semperflorens.We therefore suggested that H_2O_2 produced by photosynthetic electron transport,SOD,GO and NADPH oxidase may be involved in low temperature induced anthocyanin biosynthesis in Beginia semperflorens.
To analyze the effects of different H_2O_2 inhibitors on anthocyanin biosynthesis induced by low temperature,to discuss the function principle and function way of different originations of H_2O_2 produced in this process,the activity of SOD,POD,PAO and GO enzymes,the content of H_2O_2 and MDA,and the transcription levels of anthocyanin biosynthesis genes were measured under different treatments in Begonia semperflorens,using the methods of enzyme-activity analysis and qRT-PCR,etc..The results showed that:The activities of SOD,POD,PAO and GO were significantly increased by low temperature,causing H_2O_2 accumulation and an increase in MDA content in Beginia semperflorens.The transcription levels of anthocyanin biosynthesis genes(namely BsPAL,BsCHS,BsF3 H and BsANS) and anthocyanin content were consequently soared in Beginia semperflorens;After the pretreatment with DCMU(an inhibitor of photosynthetic electron transport),DDC(an inhibitor of SOD),OX(an inhibitor of GO) and DPI(an inhibitor ofNADPH oxidase) for 6 hours and then treated with low temperature,in different degrees,the activities of SOD,POD,PAO and GO all decreased,the contents of H_2O_2 and MDA decreased either;blocked partially anthocyanin biosynthesis by decreasing the transcription levels of anthocyanin biosynthesis genes.As a result,pretreatments of all the test inhibitors decreased the contents of H_2O_2 came from photosynthetic electron transport,SOD,GO and NADPH oxidase;and anthocyanin content was also decreased to different level in Beginia semperflorens.We therefore suggested that H_2O_2 produced by photosynthetic electron transport,SOD,GO and NADPH oxidase may be involved in low temperature induced anthocyanin biosynthesis in Beginia semperflorens.
引文
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[11]Wang J,Guo M,Li Y,et al.High-throughput transcriptome sequencing reveals the role of anthocyanin metabolism in Begonia semperflorens under high light stress[J].Photochemistry&Photobiology,2017,94(1):105-114.
[12]Zhang J,Chen C,Zhang D,et al.Reactive oxygen species produced via plasma membrane NADPH oxidase regulate anthocyanin synthesis in apple peel[J].Planta,2014,240(5):1023-1035.
[13]SimonovicováM,HuttováJ,Mistrík I,et al.Root growth inhibition by aluminum is probably caused by cell death due to peroxidase-mediated hydrogen peroxide production[J].Protoplasma,2004,224(1-2):91-98.
[14]She X P,Huang A X,Yun R.Hydrogen peroxide generated by copper amine oxidase involved in adventitious root formation in mung bean hypocotyl cuttings[J].Australian Journal of Botany,2010,58(8):656-662.
[15]穆茜,张丹丹,李千惠,等.5个海棠品种果实品质分析及评价[J].南方农业学报,2018,49(5):971-978.
[16]陈琳琳,吴瑞姣,罗思谦,等.4种PCR体系对湖北海棠DNA扩增效果的对比[J].山西农业科学,2013,41(8):771-773.
[17]马廷蕊.NO和H2O2对马铃薯花青素合成调控及信号转导[D].兰州:甘肃农业大学,2013.
[18]Foyer C H,Bloom A J,Queval G,et al.Photorespiratory metabolism:genes,mutants,energetics,and redox signaling[J].Annual Review of Plant Biology,2009,60(1):455-484.
[19]冯汉青,白晶月,管冬冬,等.胞外H2O2及NADPH氧化酶参与了铜胁迫对植物细胞死亡的诱导[J].植物研究,2015,35(5):710-715.
[2]Sivankalyani V,Feygenberg O,Diskin S,et al.Increased anthocyanin and flavonoids in mango fruit peel are associated with cold and pathogen resistance[J].Postharvest Biology&Technology,2016,111:132-139.
[3]Das P K,Dong H S,Choi S B,et al.Sugar-hormone crosstalk in anthocyanin biosynthesis[J].Molecules&Cells,2012,34(6):501-507.
[4]Hughes N M.Winter leaf reddening in‘evergreen’species.[J].New Phytologist,2011,190(3):573-581.
[5]Mita S,Murano N,Akaike M,et al.Mutants of Arabidopsis thaliana with pleiotropic effects on the expression of the gene for beta-amylase and on the accumulation of anthocyanin that are inducible by sugars[J].Plant Journal,2010,11(4):841-851.
[6]Brennan T,Frenkel C.Involvement of hydrogen peroxide in the regulation of senescence in pear[J].Plant Physiology,1977,59(3):411-416.
[7]Cakmak I,Horst W J.Effect of aluminium on lipid peroxidation,superoxide dismutase,catalase,and peroxidase activities in root tips of soybean(Glycine max)[J].Physiologia Plantarum,2010,83(3):463-468.
[8]汪天,郭世荣,刘俊,等.多胺氧化酶检测方法的改进及其在低氧水培黄瓜根系中的应用[J].植物生理学报,2004,40(3):358-360.
[9]Booker F,Reid C,Brunschonharti S,et al.Photosynthesis and photorespiration in soybean[Glycine max(L.)Merr.]chronically exposed to elevated carbon dioxide and ozone[J].Journal of Experimental Botany,1997,48(315):1843-1852.
[10]郭美丽,王佳婉,王珂,等.外源ABA对低温不同光周期下四季秋海棠叶片花色素苷的诱导机理[J].西北植物学报,2016,36(10):1999-2007.
[11]Wang J,Guo M,Li Y,et al.High-throughput transcriptome sequencing reveals the role of anthocyanin metabolism in Begonia semperflorens under high light stress[J].Photochemistry&Photobiology,2017,94(1):105-114.
[12]Zhang J,Chen C,Zhang D,et al.Reactive oxygen species produced via plasma membrane NADPH oxidase regulate anthocyanin synthesis in apple peel[J].Planta,2014,240(5):1023-1035.
[13]SimonovicováM,HuttováJ,Mistrík I,et al.Root growth inhibition by aluminum is probably caused by cell death due to peroxidase-mediated hydrogen peroxide production[J].Protoplasma,2004,224(1-2):91-98.
[14]She X P,Huang A X,Yun R.Hydrogen peroxide generated by copper amine oxidase involved in adventitious root formation in mung bean hypocotyl cuttings[J].Australian Journal of Botany,2010,58(8):656-662.
[15]穆茜,张丹丹,李千惠,等.5个海棠品种果实品质分析及评价[J].南方农业学报,2018,49(5):971-978.
[16]陈琳琳,吴瑞姣,罗思谦,等.4种PCR体系对湖北海棠DNA扩增效果的对比[J].山西农业科学,2013,41(8):771-773.
[17]马廷蕊.NO和H2O2对马铃薯花青素合成调控及信号转导[D].兰州:甘肃农业大学,2013.
[18]Foyer C H,Bloom A J,Queval G,et al.Photorespiratory metabolism:genes,mutants,energetics,and redox signaling[J].Annual Review of Plant Biology,2009,60(1):455-484.
[19]冯汉青,白晶月,管冬冬,等.胞外H2O2及NADPH氧化酶参与了铜胁迫对植物细胞死亡的诱导[J].植物研究,2015,35(5):710-715.