热激因子AtHsfA1a突变对热胁迫中拟南芥抗坏血酸过氧化物酶的影响
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摘要
【目的】探讨热激因子AtHsfA1a突变对热胁迫中拟南芥抗坏血酸过氧化物酶的影响。【方法】以T-DNA插入AtHsfA1a基因突变拟南芥及野生型植株为材料,采用分光光度法比较热胁迫下不同基因型植株的抗坏血酸过氧化物酶(APX,EC1.11.1.7)活性的变化,RT-PCR法研究AtHsfA1a对APX表达的影响,染色质免疫沉淀技术和凝胶阻滞电泳分析体内外研究AtHsfA1a与APX基因启动子区的结合情况。【结果】在热胁迫下AtHsfA1a基因突变拟南芥植株中的APX的活性低于野生型,且AtHsfA1a基因突变拟南芥植株中的APX的mRNA的相对量低于野生型;体内研究发现在热胁迫下AtHsfA1a基因突变拟南芥植株中未筛选出APX基因启动子区片段,而野生型筛选出APX基因启动子区片段,体外研究发现AtHsfA1a与APX基因启动子区的结合是直接的。【结论】热激因子AtHsfA1a突变对热胁迫中拟南芥APX的影响是从转录调控水平直接影响APX的表达。
【Objective】Effect of heat shock factor AtHsfA1a mutation on ascorbate peroxidase under heat stress in Arabidopsis thaliana was studied.【Method】T-DNA inserted mutant of Arabidopsis AtHsfA1a and wild-type plants were used as experimental materials to compare activity of Ascorbate peroxidase( APX,EC1. 11. 1. 7) in different genotypes under heat stress by using spectrophotometry. The gene expression levels of the Antioxidant enzyme APX of different Arabidopsis genotypes were analyzed under heat stress by using RT-PCR. The binding of AtHsfA1a to the promoter regions of APX were investigated by using chromatin immunoprecipitation( CHIP) in vivo and electrophoretic mobility shift assay( EMSA) in vitro.【Result】The activity of APX in T-DNA inserted mutant of Arabidopsis AtHsfA1a was less than that in the wild-type plants under heat stress. We also found that the mRNA levels of APX in T-DNA inserted mutant of Arabidopsis were lower than those in wild-type plants. The results showed that the binding of AtHsfA1a to the promoter regions of APX at different degrees existed in vivo and in vitro.【Conclusion】Effect of heat shock factor AtHsfA1a mutation on ascorbate peroxidase under heat stress in Arabidopsis thaliana was directly regulated the expression of APX by the level of transcription.
【Objective】Effect of heat shock factor AtHsfA1a mutation on ascorbate peroxidase under heat stress in Arabidopsis thaliana was studied.【Method】T-DNA inserted mutant of Arabidopsis AtHsfA1a and wild-type plants were used as experimental materials to compare activity of Ascorbate peroxidase( APX,EC1. 11. 1. 7) in different genotypes under heat stress by using spectrophotometry. The gene expression levels of the Antioxidant enzyme APX of different Arabidopsis genotypes were analyzed under heat stress by using RT-PCR. The binding of AtHsfA1a to the promoter regions of APX were investigated by using chromatin immunoprecipitation( CHIP) in vivo and electrophoretic mobility shift assay( EMSA) in vitro.【Result】The activity of APX in T-DNA inserted mutant of Arabidopsis AtHsfA1a was less than that in the wild-type plants under heat stress. We also found that the mRNA levels of APX in T-DNA inserted mutant of Arabidopsis were lower than those in wild-type plants. The results showed that the binding of AtHsfA1a to the promoter regions of APX at different degrees existed in vivo and in vitro.【Conclusion】Effect of heat shock factor AtHsfA1a mutation on ascorbate peroxidase under heat stress in Arabidopsis thaliana was directly regulated the expression of APX by the level of transcription.
引文
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[7]Liu H,Liao H,Charng Y.The role of class A1 heat shock factors(HSFA1s)in response to heat and other stresses in Arabidopsis[J].Plant Cell Environ,2011,34:738-751.
[8]Imlay J A.Pathway of oxidative damage[J].Annu Rev Microbiol,2003,57:395-418.
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[10]Foyer C H,Noctor G.Oxidant and antioxidant signaling in plants:a reevaluation of the concept of oxidative stress in a physiological context[J].Plant Cell Environ,2005,28:1056-1071.
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[12]Davletova S,Rizhsky L,Liang H,et al.Cytosolic ascorbate peroxidase 1 is a central component of the reactive oxygen gene network of Arabidopsis[J].Plant Cell,2005,17:268-281.
[13]周静,曾学英,贺维,等.低温胁迫下蓝花楹的耐寒生理机制分析[J].西南农业学报,2016,29:74-80.
[14]Storozhenko S.The heat-shock element is a functional component of the Arabidopsis APX.I gene promoter[J].Plant Physiol,2005,118:1005-1014.
[15]Panchuk I I,Volkov R A,Schoffl F.Heat stress and heat shock transcription factor-dependent expression and activity of ascorbate peroxidase in Arabidopsis[J].Plant Physiol,2002,29:838-853.
[2]Akerfelt M,Morimoto R I,Sistonen L.Heat shock factors:integrators of cell stress,development and lifespan[J].Molecular Cell Biology,2010,11:545-555.
[3]von Koskull-Doring P,Scharf K,Nover L.The diversity of plant heat stress transcription factors[J].Trends Plant Sci,2007,12:452-457.
[4]Nover L.Arabidopsis and the heat stress transcription factor world:how many heat stress transcription factors do we need?[J].Cell Stress Chaperones,2001(6):177-189.
[5]Guo LH,Chen S N,Liu KH,et al.Isolation of heat shock factor Hsf A1a-binding sites in vivo revealed variations of heat shock elements in Arabidopsis thaliana[J].Plant Cell Physiol,2008,49:1306-1315.
[6]Liu Y,Zhang C,Chen J,et al.Arabidopsis heat shock factor Hsf A1a directly senses heat stress,p H changes,and hydrogen peroxide via the engagement of redox state[J].Plant Physiology and Biochemistry,2013,64:92-98.
[7]Liu H,Liao H,Charng Y.The role of class A1 heat shock factors(HSFA1s)in response to heat and other stresses in Arabidopsis[J].Plant Cell Environ,2011,34:738-751.
[8]Imlay J A.Pathway of oxidative damage[J].Annu Rev Microbiol,2003,57:395-418.
[9]Mittler H.Reactive oxygen gene network of plants[J].Trends in Plant Science,2004(9):490-498.
[10]Foyer C H,Noctor G.Oxidant and antioxidant signaling in plants:a reevaluation of the concept of oxidative stress in a physiological context[J].Plant Cell Environ,2005,28:1056-1071.
[11]Busch W,Wunderlich M,Sch9ffl F.Identification of novel heat shock factor-dependent genes and biochemical pathways in Arabidopsis thaliana[J].The Plant Journal,2005,41:1-14.
[12]Davletova S,Rizhsky L,Liang H,et al.Cytosolic ascorbate peroxidase 1 is a central component of the reactive oxygen gene network of Arabidopsis[J].Plant Cell,2005,17:268-281.
[13]周静,曾学英,贺维,等.低温胁迫下蓝花楹的耐寒生理机制分析[J].西南农业学报,2016,29:74-80.
[14]Storozhenko S.The heat-shock element is a functional component of the Arabidopsis APX.I gene promoter[J].Plant Physiol,2005,118:1005-1014.
[15]Panchuk I I,Volkov R A,Schoffl F.Heat stress and heat shock transcription factor-dependent expression and activity of ascorbate peroxidase in Arabidopsis[J].Plant Physiol,2002,29:838-853.