玉米PHYB1基因的克隆、改造及其在拟南芥中的功能分析
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摘要
光是一个重要的环境因子,影响植物生长发育的诸多方面。植物通过多种光受体感受环境中的光信号,其中以红光与远红光受体——光敏色素的研究最为透彻。研究表明,将拟南芥phyB的104位点与361位点的酪氨酸(Y)残基改造为苯丙氨酸(F)残基可增强其活性。为了研究玉米光敏色素B1(PHYB1)的功能,构建了玉米光敏色素B1基因的3种重组质粒:pZmPHYB1∷ZmPHYB1~(WT)、pZmPHYB1∷ZmPHYB1~(Y98F)(对应拟南芥Y104F突变)和pZmPHYB1∷ZmPHYB1~(Y359F)(对应拟南芥Y361F突变),并将其转入拟南芥phyB-9突变体中,然后对转基因株系进行表型分析。结果表明,ZmPHYB1可抑制phyB-9突变体下胚轴及叶柄的伸长;ZmPHYB1可与拟南芥AtPIF5互作并诱导下游避荫反应响应基因和生长素合成基因的表达;Y98F和Y359F氨基酸的替换可增强ZmPHYB1的活性。研究结果表明ZmPHYB1在拟南芥中具有介导避荫反应的作用,同时也将为玉米耐密株型改良提供参考。
As an important environmental factor, light affects many aspects of plant growth and development. Plants use several classes of photoreceptors to perceive the light signals in the ambient environment. Among the photoreceptors, phytochromes, which sense red and far-red light signals, are best studied so far. Previous studies have shown that changing the tyrosine(Y) at position 104 and 361 of Arabidopsis phyB to phenylalanine(F) can enhance its activity. To investigate the function of maize phytochrome B1, we constructed three binary vectors: Wild type pZmPHYB1∷ZmPHYB1~(WT), pZmPHYB1∷ZmPHYB1~(Y98F)(mimicking Y104 F of Arabidopsis phyB) and pZmPHYB1∷ZmPHYB1~(Y359F )(mimicking Y361 F of Arabidopsis phyB), and transformed them into the Arabidopsis phyB-9 loss-of-function mutant. Phenotypic analyses of the transgenic lines showed that the maize PHYB1 gene can largely complement the Arabidopsis phyB-9 mutant phenotypes, including hypocotyl elongation and petiole elongation. Furthermore, ZmPHYB1 can physically interact with the Arabidopsis PIF5 protein and induce the expression of downstream shade avoidance responsive and auxin biosynthesis related genes. The Y98 F and Y359 F amino acid substitutions can enhance the activity of ZmPHYB1. In summary, our results indicated that ZmPHYB1 plays an important role in mediating shade avoidance response in Arabidopsis, and provided useful clues for genetic improvement maize plant architecture for adapting to high-density planting.
As an important environmental factor, light affects many aspects of plant growth and development. Plants use several classes of photoreceptors to perceive the light signals in the ambient environment. Among the photoreceptors, phytochromes, which sense red and far-red light signals, are best studied so far. Previous studies have shown that changing the tyrosine(Y) at position 104 and 361 of Arabidopsis phyB to phenylalanine(F) can enhance its activity. To investigate the function of maize phytochrome B1, we constructed three binary vectors: Wild type pZmPHYB1∷ZmPHYB1~(WT), pZmPHYB1∷ZmPHYB1~(Y98F)(mimicking Y104 F of Arabidopsis phyB) and pZmPHYB1∷ZmPHYB1~(Y359F )(mimicking Y361 F of Arabidopsis phyB), and transformed them into the Arabidopsis phyB-9 loss-of-function mutant. Phenotypic analyses of the transgenic lines showed that the maize PHYB1 gene can largely complement the Arabidopsis phyB-9 mutant phenotypes, including hypocotyl elongation and petiole elongation. Furthermore, ZmPHYB1 can physically interact with the Arabidopsis PIF5 protein and induce the expression of downstream shade avoidance responsive and auxin biosynthesis related genes. The Y98 F and Y359 F amino acid substitutions can enhance the activity of ZmPHYB1. In summary, our results indicated that ZmPHYB1 plays an important role in mediating shade avoidance response in Arabidopsis, and provided useful clues for genetic improvement maize plant architecture for adapting to high-density planting.
引文
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[2] Vierstra R D,Zhang J.Phytochrome signaling:Solving the gordian knot with microbial relatives [J].Trends Plant Sci.,2011,16(8):417-426.
[3] Li J G,Li G,Wang H Y,et al..Phytochrome signaling mechanisms [J].Arabidopsis Book,2002,3(3):e0148.
[4] Andrea C,Giovanna S,Massimiliano S,et al..Dynamics of the shade-avoidance response in Arabidopsis [J].Plant Physiol.,2013,163(1):331-353.
[5] Pablo L,Quail P H.PIFs:Pivotal components in a cellular signaling hub [J].Trends Plant Sci.,2011,16(1):19-28.
[6] Lorrain S,Allen T,Duek P G,et al..Phytochrome-mediated inhibition of shade avoidance involves degradation of growth-promoting bHLH transcription factors [J].Plant J.,2010,53(2):312-323.
[7] Stamm P,Kumar P P.The phytohormone signal network regulating elongation growth during shade avoidance [J].J.Exp.Bot.,2010,61(11):2889.
[8] Nito K,Wong C L,Yates J,et al..Tyrosine phosphorylation regulates the activity of phytochrome photoreceptors [J].Cell Rep.,2013,3(6):1970-1979.
[9] Zhang J R ,Stankey R J,Vierstra R D.Structure-guided engineering of plant phytochrome b with altered photochemistry and light signaling [J].Plant Physiol.,2013,161(3):1445-1457.
[10] Halliday K J,Koornneef M,Whitelam G C.Phytochrome B and at least one other phytochrome mediate the accelerated flowering response of Arabidopsis thaliana L.to low red/far-red ratio [J].Plant Physiol.,1994,104(4):1311-1315.
[11] Kutschera U,Briggs W R.Seedling development in buckwheat and the discovery of the photomorphogenic shade-avoidance response [J].Plant Biol.,2013,15(6):931-940.
[12] Smith H,Whitelam G C.The shade avoidance syndrome:Multiple responses mediated by multiple phytochromes [J].Plant Cell Envir.,2010,20(6):840-844.
[13] Boardman N K.Comparative photosynthesis of sun and shade plants [J].Ann.Rev.Plant Physiol.,1977,28(1):355-377.
[14] Middleton L.Shade-tolerant flowering plants:Adaptations and horti-cultural implications[J].Acta Hortic.,doi:10.17660/ActaHortic.2001.552.9.
[15] Smith H.Light quality,photoperception,and plant strategy[J].Ann.Rev.Plant Physiol.,1982,33(1):481-518.
[16] Bennetzen J L.Patterns in grass genome evolution [J].Curr.Opin.Plant Biol.,2007,10(2):176-181.
[17] Childs K L.The sorghum photoperiod sensitivity gene,Ma3,encodes a phytochrome B [J].Plant Physiol.,1997,113(2):611-619.
[18] Kebrom T H,Brutnell T P,Finlayson S A.Suppression of sorghum axillary bud outgrowth by shade,phyB and defoliation signalling pathways [J].Plant Cell Envir.,2010,33(1):48-58.
[19] Kebrom T H,Brutnell T P.The molecular analysis of the shade avoidance syndrome in the grasses has begun [J].J.Exp.Bot.,2007,58(12):3079-3089.
[20] Wang H,Wu G,Zhao B,et al..Regulatory modules controlling early shade avoidance response in maize seedlings [J].BMC Genom.,2016,17(1):269.
[21] 杨慧莲,韩旭东,郑风田.全球主产国(地区)玉米生产、贸易、消费及库存状况对比——基于1996/1997-2016/2017产季数据测算[J].世界农业,2017(6):28-35.
[22] Duvick D N.Genetic progress in yield of United States maize (Zea mays L.)[J].Maydica,2005,50(3):193-202.
[23] Liu Y,Xie Y,Wang H,et al..Light and ethylene coordinately regulate the phosphate starvation response through transcriptional regulation of PHOSPHATE STARVATION RESPONSE1 [J].Plant Cell,2017,29(9):268-2017.
[24] Murray M G,Thompson W F.Rapid isolation of high molecular weight plant DNA [J].Nucl.Acids Res.,1980,8(19):4321-4325.
[25] Clough S J,Bent A F.Floral dip:A simplified method for agrobacterium-mediated transformation of Arabidopsis thaliana [J].Plant J.,2010,16(6):735-743.
[26] Walley J W,Sartor R C,Shen Z.Integration of omic networks in a developmental atlas of maize [J].Science,2016,353(6301):814-818.
[27] Fankhauser C,Casal J J.Phenotypic characterization of a photomorphogenic mutant [J].Plant J.,2004,39(5):747-760.
[28] López-Juez E,Nagatani A,Tomizawa K,et al..The cucumber long hypocotyl mutant lacks a light-stable PHYB-like phytochrome [J].Plant Cell,1992,4(3):241-251.
[29] Somers D E,Sharrock R A,Tepperman J M,et al..The hy3 long hypocotyl mutant of arabidopsis is deficient in phytochrome B [J].Plant Cell,1991,3(12):1263-1274.
[30] Kebrom T H,Burson B L,Finlayson S A.Phytochrome B represses teosinte branched1 expression and induces sorghum axillary bud outgrowth in response to light signals [J].Plant Physiol.,2006,140(3):1109-1117.
[31] Sawers R J,Sheehan M J,Brutnell T P.Cereal phytochromes:targets of selection,targets for manipulation?[J].Trends Plant Sci.,2005,10(3):138-143.
[32] 王元东,段民孝,邢锦丰,等.玉米理想株型育种的研究进展与展望[J].玉米科学,2008,16(3):47-50.
[33] 张世煌.论玉米高密度育种和生产上的密植[J].北京农业,2009(29):1-2.
[34] Dubois P G,Olsefski G T,Sherry F G,et al..Physiological and genetic characterization of end-of-day far-red light response in maize seedlings [J].Plant Physiol.,2010,154(1):173-186.
[35] Chen C Y,Hou Y H,Rui S,et al..Effects of planting density on yield performance and density-tolerance analysis for maize hybrids [J].Acta Agron.Sin.,2010,36(7):1153-1160.
[36] 陈德龙.不同抗倒能力玉米品种茎秆及根系等相关特征研究[D].长春:吉林农业大学,硕士学位论文,2015.