脱落酸和蛋白质可逆磷酸化试剂处理对葡萄、苹果果实糖代谢酶活性的效应
|
摘要
脱落酸(Abscisic acid，ABA)是一种具有重要生理功能的植物激素。许多研究表明ABA在果实成熟中起到重要作用，同化物分配也受到ABA的调控。本研究的目的是为了探索ABA的作用机制。首先通过使用不同浓度的ABA对果实圆片进行活体温育，测定各种糖代谢酶的活性变化，从而确定ABA是否对葡萄、苹果果实中糖代谢酶的活性具有调节作用。研究证明20μmol／L的ABA可明显地激活葡萄和苹果果实中可溶性酸性转化酶活性。
为了研究ABA激活效应的发生机制以及在ABA诱导的信号转导途径中是否有蛋白可逆磷酸化反应的参与，进一步探索了以下几方面内容：1、用(±)-ABA和(-)-ABA分别以最适激活浓度(20μmol／L)处理果实圆片。发现(-)-ABA对果实中可溶性酸性转化酶活性无明显的激活作用，从而证实自然活性形式的(+)-ABA是果实成熟发育过程中起重要作用的因素。2、用多种蛋白激酶和蛋白磷酸酶抑制剂处理果实圆片，其作用效果复杂，并未表现明显的一致性。说明在ABA激活糖代谢酶活性的信号传递中可能有多个蛋白激酶或蛋白磷酸酶参与，相互作用，连锁反应。3、用蛋白质合成抑制剂环己亚胺和mRNA转录抑制剂放线菌素D处理果实圆片，对葡萄、苹果果实中酸性转化酶均有影响，但结果不尽相同。在葡萄果实中，酸性转化酶活性被抑制，可能是由于ABA的诱导效应是在mRNA转录和翻译水平上进行的。而在苹果果实中，加入核酸和蛋白质合成抑制剂，反而增强了ABA对苹果果实圆片酸性转化酶活性的激活效应。4、在果实的酶粗提液中进行酸性磷酸酶和碱性磷酸酶处理。发现仅就酸性转化酶来讲，酸性磷酸酶的作用条件与其相符，在葡萄、苹果果实粗提酶液中加入酸性磷酸酶的实验中，该酶对酸性转化酶活性有激活作用。而碱性磷酸酶由于反应缓冲环境与酸性转化酶差异较大，对于酸性转化酶的脱磷酸化反应不适用。
Abscisic acid (ABA) is a vital plant hormone that plays an important role in various plant developmental processes including fruit ripening and carbohydrate partitioning in fruit. This study aims at a better insight into the ABA action mechanism. By incubating the fruit discs in the ABA-contained medium, the results showed that ABA strongly activated the fruit soluble acid invertase. The ABA-induced acid invertase activation appeared significantly 4 hours after 20umol/L ABA incubation in grape berry and 3 hours after 20umol/L ABA in apple fruit. Further experiments were conducted to assess possible role of protein reversible phosphorylation in the ABA-induced acid invertase activation. By adding inhibitors of protein kinase and protein phosphatase, the results indicated that there may exist a complex regulation mechanism involving various protein kinases and protein phosphatases that may interact among them to constitute a complex signaling cascades. The protein synthesis inhibitor cycloheximide and mRNA transcription inhibitor Actidione D were also used to show futher mechanism of ABA action. In grape berry acid invertase activity was suppressed and the inhibitors annulled ABA activation effects. In apple fruit ABA effects were strengthened instead. By using acid phosphatase in the crude extract, dephosporylation helped to enhance the activity of acid invertase.
为了研究ABA激活效应的发生机制以及在ABA诱导的信号转导途径中是否有蛋白可逆磷酸化反应的参与，进一步探索了以下几方面内容：1、用(±)-ABA和(-)-ABA分别以最适激活浓度(20μmol／L)处理果实圆片。发现(-)-ABA对果实中可溶性酸性转化酶活性无明显的激活作用，从而证实自然活性形式的(+)-ABA是果实成熟发育过程中起重要作用的因素。2、用多种蛋白激酶和蛋白磷酸酶抑制剂处理果实圆片，其作用效果复杂，并未表现明显的一致性。说明在ABA激活糖代谢酶活性的信号传递中可能有多个蛋白激酶或蛋白磷酸酶参与，相互作用，连锁反应。3、用蛋白质合成抑制剂环己亚胺和mRNA转录抑制剂放线菌素D处理果实圆片，对葡萄、苹果果实中酸性转化酶均有影响，但结果不尽相同。在葡萄果实中，酸性转化酶活性被抑制，可能是由于ABA的诱导效应是在mRNA转录和翻译水平上进行的。而在苹果果实中，加入核酸和蛋白质合成抑制剂，反而增强了ABA对苹果果实圆片酸性转化酶活性的激活效应。4、在果实的酶粗提液中进行酸性磷酸酶和碱性磷酸酶处理。发现仅就酸性转化酶来讲，酸性磷酸酶的作用条件与其相符，在葡萄、苹果果实粗提酶液中加入酸性磷酸酶的实验中，该酶对酸性转化酶活性有激活作用。而碱性磷酸酶由于反应缓冲环境与酸性转化酶差异较大，对于酸性转化酶的脱磷酸化反应不适用。
Abscisic acid (ABA) is a vital plant hormone that plays an important role in various plant developmental processes including fruit ripening and carbohydrate partitioning in fruit. This study aims at a better insight into the ABA action mechanism. By incubating the fruit discs in the ABA-contained medium, the results showed that ABA strongly activated the fruit soluble acid invertase. The ABA-induced acid invertase activation appeared significantly 4 hours after 20umol/L ABA incubation in grape berry and 3 hours after 20umol/L ABA in apple fruit. Further experiments were conducted to assess possible role of protein reversible phosphorylation in the ABA-induced acid invertase activation. By adding inhibitors of protein kinase and protein phosphatase, the results indicated that there may exist a complex regulation mechanism involving various protein kinases and protein phosphatases that may interact among them to constitute a complex signaling cascades. The protein synthesis inhibitor cycloheximide and mRNA transcription inhibitor Actidione D were also used to show futher mechanism of ABA action. In grape berry acid invertase activity was suppressed and the inhibitors annulled ABA activation effects. In apple fruit ABA effects were strengthened instead. By using acid phosphatase in the crude extract, dephosporylation helped to enhance the activity of acid invertase.
引文
陈发河，蔡慧农，冯作山，张维一，廖康．葡萄浆果发育过程中激素水平变化．植物生理与分子生物学学报．2002，28：391—395．
陈尚武，张大鹏．ABA和Fluridone对苹果果实成熟的影响．植物生理学报．2000，26：123—129
陈尚武，张大鹏．苹果果实发育晚期ABA的来源与代谢．园艺学报．1998，25：313-318
陈长征，李伯良，夏其昌等．蛋白激酶的结构与功能．见敖世洲主编蛋白质可逆磷酸化对细胞活动的调节．上海科学出版社．1994，P170-200
刘慧英，朱祝军．转化酶在高等植物蔗糖代谢中的作用研究进展．植物学通报．2002，19(6)：666-674
刘璞，陈珈，植物激素脱落酸的信号转导．植物生理学通讯．2000，36(2)：165-170
吕英民，张大鹏．果实发育过程中糖的积累．植物生理学讯．，2000，36(3)：258～265
吕英明，张大鹏，严海燕．苹果果实韧皮部及其周围薄壁组织的超微结构观察和功能分析．植物学报．2000，42：32-42
吕英明，张大鹏，严海燕．苹果果实糖卸载机制的研究．园艺学报．1999，26：141-146
王永章，张大鹏．果糖和葡萄糖诱导了苹果果实酸性转化酶翻译后的抑制性调节．中国科学．(Ser C) 2002，45：309-321
张大鹏，许雪峰，张子莲，贾文锁．葡萄果实始熟机理的研究—缓慢生长期外施激素和环剥的效应．园艺学报．1997，24(1)：1-7
张大鹏，张子莲，陈伽．葡萄果实发育过程中脱落酸结合蛋白动力学特性的变化．植物学报．1996，38(12)：932-935
周庆红，李成琼，匡全．生物学杂志Vol，20 No．3 Jun，2003 1-4
张承才，彭玲．蛋白激酶在植物生长及发育中作用[M]．见：许智宏，刘春明主编．植物发育的分子生理．北京：科学出版社，1998，172-192．
孙大业，马力耕主编．细胞信号转导(第二版)[M]．北京：科学出版社，1998，350-355
郭艳林．植物体内蛋白质磷酸化及其在信号传递中的作用[J]．植物理学通讯，1991，27(5)：385-390．
梁小娥，张大鹏，贾文锁．苹果和葡萄果实蛋白激酶特性分析．植物生理学报，2000，26(3)：257-262
梁小娥，张大鹏，贾文锁．苹果和葡萄果实发育过程中蛋白激酶特性的比较研究．植物学报 2000，42(11)：1131-1136
周庆红，李成琼，匡全．植物蛋白激酶研究进展．生物学杂志，2003，20(3)：1-4
Addicott FT, Carns HR. History and introduction. In : Addicott FT (ed) . Abscisic Acid. New York : Praeger Sci , 1983. 1～21
Akio O,Hidokazu I, Takanori S, Shigeo N, Tuyoshi I, Masashi H. Suppression of acid invertase activity by antisense RNA modifies the sugar composition of tomato fruit. Plant Cell Physiol. 1995,36 (2) :369～376
Ackerson R.C. Invertase activity and abscisic in developing soybean reproductive structures. Crop Science. 1985,25:615-620
Anderson M N, Asch F, Wu Y, Jensen C R, Naested h, Mogensen V O&Koch R E. Soluble invertase expression is an early target of drought stress during the critical, abortion-sensing phase of young ovary development in maize. Plant Physiology. 2002,130:591-604
Anderson R S, Ewing E E&Hedges S. Inhibition of potato tuber invertase by an endogenous inhibitor. Plant Physiology. 1980,66:451-456
Armstrong F, Leung J, Grabov A, Brearley J, Giraudat J, Blatt MR. Sensitivity to abscisic acid of guard cell K~+ channels is suppressed by ABI121, a mutant Arabidopsis gene encoding a putative protein phosphate. Proc Natl Acad Sci USA. 1995,92:9520-9524
Assmann SM. Signal transduction in guard cells. Annu Rev Cell Biol.1993,9:345-375
Aurelio G C, Steven D, Verhey, et al. An abscisic acid -induced protein kinase PKABAL mediates abscisic acid - suppressed gene exression in barey aleurone layers. Plant Biol.1999,96:1767-1772
Beruter J&Studer F M E. Comparison of Sorbitol transport in excised tissue discs and cortex tissue of intact apple fruit. Journal of Plant Physiology. 1995,146:95-102
Beruter J. Sugar accumulation and changes in the activities of related enzyme during development of apple fruit. Journal of Plant Physiology. 1985,121:331-334
Beruter J, Studer F M E&Ruedi P. Sorbitol and sucrose partitioning in the growing apple fruit. Journal of Plant Physiology. 1997,151:269-276
Beruter J. Effect of abscisic acid on sorbitol uptake in growing apple fruit. Journal of Experimental Botany. 1983,34:737-743
Beruter J&Studer F. M. E. The effect of girdling on carbohydrate portioning in the growing
apple fruit. Journal of Plant Physiology. 1997,51:277-285
Beruter J..Effect of abscisic on sorbitol uptake in growing apple fruits. Journal of Experimental Botany. 1983,34:737-743
Bowler C ,Neuhaus G, Yamagata H ,Cgua N-H. Cyclic GMP and calcium mediated phytochrome phototransduction [J] . Cell. 1994, 77: 73～81.
Bracho G.E. &Whitaker J.R. Purification and partial characterization of potato invertase and its endogenous proteinaceous inhibitor. Plant Physiology. 1990,92:386-394
Burch LR, Davies HV, Cuthbert EM & Machray G.C. Purification of soluble invertase from potato. Phytochemistry. 1992,131:1901-1904
Bussis D, Heineke D, Sonnewaid U, Willmitzer L, Raschke K & Heldt H W. Solute accumulation and decreased photosynthesis in leaves of potato plants expressing yeast derived invertase either in the apoplast, vacuole or cytosol. Planta. 1997,202:126-136
Burbridge. Characterization of the ABA deficienttomato mutant notabilis and its relationship with maize VP14[J]. Planta. 1999 ,17(4):427-431
Chen J Q &Black C C. Biochemical and immunological properties of alkaline invertase isolated from sprouting soybean hypocotyls. Archives in Biochemistry & Biophysics. 1992,295:61-69
Coombe B. G. The grape berry as a sink. ActaHorticulturae. 1989,239:149-158
Davies C.,Boss P.K. & Robinson S.P. Treatment of grape berries, a nonclimacteric fruit with a synthetic auxin, retards ripening and alters the expression of developmental regulated genes. Plant Physiology. 1997, 115:1155-1161
Davis C, Robinson S P. Sugar accumulation in grape berries. Plant Physiol .1996,111:275-283
Estruch J J, Beltran J P. Changes in invertase activities precede ovary growth induced by gibberellic acid in pisum sativum. Physiol Plant.1991,81: 319～326
Ehness R , Ecker M , Roitsch T. Glucose and stress independently regulate source and sink metabolism and defense mechanism via signal transduction pathways involving protein phosphorylation. Plant Cell. 1997,9: 1825～1841
Elliot K.J.,Butler W.O.,Dickinson C.D.,Konno Y.,Vedvick T.S.,Fitzmaurice L,Mirkov T.E.
Isolation and characterization of fruit vacuolar invertase genes from two tomato species and temporal differences in mRNA levels during fruit ripening. Plant Molecular. Biology. 1993,21:515-524
Famiani F.,Walker R.P.,Tecsi L.,Chen Z.H.,Proietti P. &Leegood R.C. An immunohistochemical study of the comparmentation of metabolism during the development of grape berries. Journal of Experimental Botany. 2000,51:675-683
Foyer C.H. The basis for source-sink interaction in leaves. Plant Physiology &Biochemistry. 1987,25:649-659
Ferreira PCG, The Arabidopsis functional homolog of the P34cdc2 protein kinase[J].Plant Cell.1991,3:531-540.
Frommer W.B.&Sonnewald U. Molecular analysis of carbon partitioning in solanaceous species. Journal of Experimental Botany. 1995,46:587-607
Geotz M & Roitsch T. The different pH optima and subtrate specifities of extracellular an vacuolar invertases from plants are determined by a single amino acid substitution. Plant Journal. 1999,20:707-711
Geota M.&Roitsch T. Identification of amino acid essential for enzymetic activity of plant invertase. Journal of Plant Physiology. 2000,157:581-585
Godt D.E.&Roitsch T. Regulation and tissue-specific distribution of mRNAs for three extracellular invertase isoenzymes of tomato suggests:an important function in establishing and maintaining sink metabolism. Plant Physiology. 1997,80:464-471
Goupil P.,Loncle D.,Druart N.,Bellettre A.&Rambour S. Influence of ABA on nitrate reductase activity and carbohydrate metabolism in chicory roots. Journal of Experimental Botany. 1998, 49:1855-1862
Greenland A.J. & Lewis D.H. The acid invertase activity of the developing third leaf of oat Ⅰ:Changes in activity of invertase and concentrations of ethanol-soluble carbohydrates. New Phytologist. 1981,88:265-277
Godt DE, Riegel A, Roitsch T. Regulation of sucrose synthase expression in chenopodium rubrum: characterization of sugar induced expression in photoautotrophic suspension
cultures and sink tissue specific expression in plants. J Plant Physiol.1995,146:231～236
Grabov A , leung J , Giraudat J , Blatt MR. Alteration of anion channel kinetics in wild2type and abi121 transgenic Nicotiana benthamiana guard cells by abscisic acid. Plant J. 1997,12:203～213
Ghosheh N S, Rayle D h, Carpita N C & Kaufman P B, Cell wall and enzyme changes during gravivespos of the leaf-sheet pulvinus of oat. Plant Physiology. 1990,157:581-585
Harper, J F. ,A calcium- dependent protein kinase with a regulatory domain similar to calmodulin[J]. Science 1993,252: 951～954.
Hawker J.S. Changes in the activities of enzymes concerned with sugar metabolism during the development of grape berries. Phytochemistry. 1969,8:9-17
Harmon AC, Yoo BC ,Mccaffery C. Pseudo substrate inhibition of CDPK, a protein kinases with a calmodulin - like domain [J] .Biochem. 1994 ,33: 7278～7287.
Hidaka et al. Molecular pharmacology of protein kinase. Annu rev Pharmacol Toxical, 1992.32:377-397
Hideaki Yamaguchi, Yoshinori Kanayama and Shohei Yamaki. Purification and properties of NAD-dependent sorbitol dehydrogenase from apple fruit. Plant Cell Physiol. 1994, 25(6):887-892
Hanks S K, et al. Protein kinases catalytic domain sequence database: identification of conserved features of primary structure and classification of family members[J]. Methods Enzymol, 1991, 200: 38-62.
Huber J L A, Huber S C. Site - specific serine phosphorylation of spinach leaf sucrose-phosphate synthase. Biochem J. 1992,28:877-882
Huber S C, Huber J L, Pao-Chi Liao, et al. Phosphorylation of serine-15 of maize leaf sucrose synthase. Plant Physiol. 1992,99:1275-1278
Hurley J R, Dean A M, Sohl J L, Koshland D E J, et al. Regulation of an enzyme by phosphorylation at the active site. Science. 1990,249:1012-1016
Isla M. I., Salerno G.,Pontis H.,Vattuone MA.& Sampietro A.R. Purification and properties of the soluble acid invertase fron Oryza Sativa. Phytochemistry. 1995,38:321-325
Islam M.S.,Matsui T.,&Yoshid Y. Carbohydrate content and the activity of sucrose synthase, sucrose phosphate and acid invertase in different tomato cultivars during fruit development. Scientia Horticulturae. 1996,65:125-136
Iwatsubo. NakagawaH. OguraN. HirabayashiY. &SatoT. Acid invertase of melon fruit: immunochemical detection of acid invertase. Plant Cell&Physiology. 1992,33(8):1127-1133
Jang J C& Sheen J. Sugar sensing in higher plants. The Plant Cell.1994,6:1665-1679
Kato T , Kubota S . Properties of invertase in sugar storage tissues of citrus fruit and changes in their activities during maturation. Physiol Plant. 1978.42:67～72
Kim J.Y.,Aline M, Sylvain G, Mahe A, Guy S, Brangeon J, Roche O, Chourey P S&Prioul J L. Characterization of two members of the maize gene family, Incw3 and Incw4, encoding cell-wall invertase. Gene. 2000,245:89-102
Kingstom S A H &Pollock C J. Soluble acid invertase activity in leaves is independent of species differences in leaf carbohydrates, diurnal sugar profiles and path of phloem loading. New Phytologist, 1999. 139:293-292
Klann E M, Cheltelat R T&Bennett B. Antisense acid invertase(TIV1)gene alters soluble sugar composition and size in transgenic tomato fruit. Plant Physiology, 1996.112:1321-1330
Knight J S & Gray J C. Expression of genes encoding the tobacco chloropolast phosphate translocator is not light-regulated and is repressed by sucrose. Molecular & General Genetic, 1994.242:586-594
Kobashi K, Gemma H&Iwahori S. Sugar accumulation in peach fruit as affected by abscisic acid(ABA)treatment in relation to some sugar metabolizing enzymes. Journal. of the Japanese Society for Horticultural Science, 1999.68:465-470
Leung J, Micehelle B D. Arabidopsis ABA response gene ABI1:feature of a calcium-modulated protein phosphatase. Science, 1994,264:1448-1451
Li J , Wang XQ , Watson MB , Assmann SM. Regulation of abscisic acid induced stomatal closure and anion channels by guard cell AAPK kinase. Science. 2000,287 : 300～303
Lowell C A , Tomlinson P T. Sucrose-metabolizing enzymes in transport tissues and adjiacent sink structures in developing citrus fruit . Plant Physiol. 1989,90:1394～
1402
Leigh R A, Reea T, Fuller WA. The localization of acid invertase activity and sucrose in the vacuoles of storage roots of beet root (Beta vulgaris) . Biochem J, 1979.178:539～547
Miron D, Schaffer A. Sucrose phosphate synthase, sucrose synthase and invertase activities in developing fruit of Lycopersicon esculintum Mill. And sucrose accumulation in Lycopersicon hirsutum humb and bonp. Plant Physiology. 1991, 95:623-627
Nishihama R ,Baanon H, Shibata W, et al. Plant homologues of components of MAPK(mitogen - activated protein kinase) signal pathways in yeast and animal cell [J] . Plant Cell Physiol.1995 ,36(5) :7479～7486.
Patrick J W. Phloem unloading: sieve element unloading and post-sieve element transport. Annual of Review Plant Physiology and Plant Molecular Biology. 1997,48:191-222
Palejwala V A, Parikh H R, Modi V V, The role of abscisic acid in the ripening of grapes. Physiol Plant. 1985,65:498-502
Pei ZM , Murata Y, Benning G, Thomine S , Klusener B ,Allen GJ , Grill E , Schroeder JL. Calcium channels activated by hydrogen peroxide mediate abscisic acid signalling in guard cells. Nature. 2000,406 : 731～734
Riesmeier JW, Hirner B , Frommer WB. Expression of the sucrose transporter from potato correlates with the sink2to2source transition in leaves. Plant Cell, 1993 ,5:1591～1598
Renner R , Schuler K, Sonnewald U. Soluble acid invertase determines the hexose-to-sucrose ratio in cold- stored potato tubers. Planta. 1996,198 :246～252
Roberts DM, Calcium-modulated proteins: Targets of intracellular calcium signals in higher plants. Annue Rev plant Physiol, Plant Mol Bio.1992,43:375
Schroeder J I, Kwak JM, Allen GJ. Guard cell abscisic acid signalling and engineering drought hardiness in plants. Nature. 2001, 410 : 327～330
Stone JM, Walker JC , Plant protein kinase families and signal transduction[J]. Plant Physiol. 1995,105: 451～457
Sylvie L , Eckhard B , Hanjo H et al. The dual function of sugar carriers : transport
and sugar sensing. Plant Cell.1999,11:707～726
Smeekens S. Sugar2Induced signal transduction in plants. Annu Rev Plant Physiol Plant Mol Biol. 2000,51:49～82
Smith RD , Walker JC. Plant protein phosphatases. A nnu Rev Plant Physiol Plant Mol Biol.1996,47:101～125
Sturm A. Moleculor characterization and functional analysis of sucrose-cleaving enzymes in carrot. Journal of Experimental Botany. 1996,47:1187-1192
Strum A. Invertase:primary structures, functions and roles in plant development and sucrose portioning. Plant Physiology. 1999, 121:1-7
Takeda S, Mano S, Ohto M, et al. Inhibitors of protein phosphatase 1 and 2A block the sugar inducible gene expression in plants. Plant Physiol, 1994,106:567～574
Tymowska L Z&Kreis M. The plant invertase: physiology, biochemistry and molecular biology. Advance in Botany Research. 1998,28:71-117
Trewavas A J, Malho R. Signal perception and transduction:the origin of the phenotype. Plant Cell.1997,9:1181-1195
Unger C, Hardegger M, Lienhard S &Strum A. cDNA cloning of carrot soluble acid β -fructofuranosidase from Daucus carota. European Journal of Biochemistry. 204:915-921
Vendrell M. &Buesa, 1989, Relationship between ABA content and ripening of apples. Acta Horticulturae. 1994,258:389-396
Verhey S D, Lomax T L. Signal transduction in vascular plants. J Plant Growth Regul. 1993, 12: 167-169
Walker R P&Pollock C J. The Purification and characterization of soluble acid invertase from coleptile of wheat. Journal of Experimental Botany. 1993,44:1029-1037
Weber H, A role for sugar transporters during seed development: molecular characterization of a hexose and a sucrise carrier in fava bean seeds. The Plant Cell.1997,9:895-908
Wei L, et al. Protein kinase superfamily-comparisons of sequence data with three-dimensional structures[J].Curr. Opin. Struct. Biol.1994, (4): 450-455.
Weschke W. Panitz R, Gubatz S, Wang Q, Radchuk R, Weber H & Wobus U. The role of invertases
and hexose transporters in controlling sugar ratios in maternal and filial tissues of barley caryopses during early development. Plant Journal. 2003,33:395-411
Winter H & Huber S C. Regulation of sucrose and regulation of activity of key enzymes. Critical Reviews In Plant Science. 2000,19(1):31-67
Woodson W R & Wang H. Invertases of carnation petals: partial purification ,characterization and changes in activity during petal growth. Physiologia Plantarum. 1987,71:224-228
Wu L L, Song I, Kim D &Kaufman P B. Molecular basis of the increase in invertase activity elicited by gravistimulation invertase oat shoot pulvini. Journal of Plant Physiology. 1993,142:179-183
Xiao W Y, Sheen J & Jang J C. The role of hexokinase in plant sugar transduction and growth and development. Plant Molecular Biology. 2000,44(4): 451-461
Yamaki S. Sorbitol oxidase converting sorbitol to glucose in apple leaf. Plant & Cell Physiol. 1980,21:591-599
Yamaki S.& Ishikawa K. Role of four sorbitol related enzymes and invertase in seasonal alteration of sugar metabolism in apple tissue. Journal of American Society Horticultural Science, 1986,111:134-137
Yelle S Chtelate R T, Dorasis M, Devema J W & Bennett A B. Sink metabolism in tomato fruit Ⅳ:genetic and bilchemical analysis of sucrose accumulation. Plant Physiology. 1991,95:1026-1035
Yin J H, Gao F F & Hu G B. The regulation of litchi maturation and coloration abscisic acid and ethylene. Acta Horticulture. 2001,558:293-296
Zhang D P, Lu Y M, Wang Y Z, Duan C Q & Yan H Y. Acid invertase is predominantly localized to cell walls of both partically symplasmically isolated sieve element/companion cell complex and parenchyma cells in developing apple fruits. Plant Cell & Environment. 2001,24:691-702
Zrenner R, Salanoubat M & Willmitzer L. Evidence of the crucial role of sucrose synthase for sink strength using transgenic potato plants. Plant Journal. 1995,7:97-107
Zrenner R, Schuler K & Sonnewald V. Soluble acid invertase determines the hexose to
sucrose ratio in cold-stored potata tubers. Planta. 1996,198:246-25
陈尚武，张大鹏．ABA和Fluridone对苹果果实成熟的影响．植物生理学报．2000，26：123—129
陈尚武，张大鹏．苹果果实发育晚期ABA的来源与代谢．园艺学报．1998，25：313-318
陈长征，李伯良，夏其昌等．蛋白激酶的结构与功能．见敖世洲主编蛋白质可逆磷酸化对细胞活动的调节．上海科学出版社．1994，P170-200
刘慧英，朱祝军．转化酶在高等植物蔗糖代谢中的作用研究进展．植物学通报．2002，19(6)：666-674
刘璞，陈珈，植物激素脱落酸的信号转导．植物生理学通讯．2000，36(2)：165-170
吕英民，张大鹏．果实发育过程中糖的积累．植物生理学讯．，2000，36(3)：258～265
吕英明，张大鹏，严海燕．苹果果实韧皮部及其周围薄壁组织的超微结构观察和功能分析．植物学报．2000，42：32-42
吕英明，张大鹏，严海燕．苹果果实糖卸载机制的研究．园艺学报．1999，26：141-146
王永章，张大鹏．果糖和葡萄糖诱导了苹果果实酸性转化酶翻译后的抑制性调节．中国科学．(Ser C) 2002，45：309-321
张大鹏，许雪峰，张子莲，贾文锁．葡萄果实始熟机理的研究—缓慢生长期外施激素和环剥的效应．园艺学报．1997，24(1)：1-7
张大鹏，张子莲，陈伽．葡萄果实发育过程中脱落酸结合蛋白动力学特性的变化．植物学报．1996，38(12)：932-935
周庆红，李成琼，匡全．生物学杂志Vol，20 No．3 Jun，2003 1-4
张承才，彭玲．蛋白激酶在植物生长及发育中作用[M]．见：许智宏，刘春明主编．植物发育的分子生理．北京：科学出版社，1998，172-192．
孙大业，马力耕主编．细胞信号转导(第二版)[M]．北京：科学出版社，1998，350-355
郭艳林．植物体内蛋白质磷酸化及其在信号传递中的作用[J]．植物理学通讯，1991，27(5)：385-390．
梁小娥，张大鹏，贾文锁．苹果和葡萄果实蛋白激酶特性分析．植物生理学报，2000，26(3)：257-262
梁小娥，张大鹏，贾文锁．苹果和葡萄果实发育过程中蛋白激酶特性的比较研究．植物学报 2000，42(11)：1131-1136
周庆红，李成琼，匡全．植物蛋白激酶研究进展．生物学杂志，2003，20(3)：1-4
Addicott FT, Carns HR. History and introduction. In : Addicott FT (ed) . Abscisic Acid. New York : Praeger Sci , 1983. 1～21
Akio O,Hidokazu I, Takanori S, Shigeo N, Tuyoshi I, Masashi H. Suppression of acid invertase activity by antisense RNA modifies the sugar composition of tomato fruit. Plant Cell Physiol. 1995,36 (2) :369～376
Ackerson R.C. Invertase activity and abscisic in developing soybean reproductive structures. Crop Science. 1985,25:615-620
Anderson M N, Asch F, Wu Y, Jensen C R, Naested h, Mogensen V O&Koch R E. Soluble invertase expression is an early target of drought stress during the critical, abortion-sensing phase of young ovary development in maize. Plant Physiology. 2002,130:591-604
Anderson R S, Ewing E E&Hedges S. Inhibition of potato tuber invertase by an endogenous inhibitor. Plant Physiology. 1980,66:451-456
Armstrong F, Leung J, Grabov A, Brearley J, Giraudat J, Blatt MR. Sensitivity to abscisic acid of guard cell K~+ channels is suppressed by ABI121, a mutant Arabidopsis gene encoding a putative protein phosphate. Proc Natl Acad Sci USA. 1995,92:9520-9524
Assmann SM. Signal transduction in guard cells. Annu Rev Cell Biol.1993,9:345-375
Aurelio G C, Steven D, Verhey, et al. An abscisic acid -induced protein kinase PKABAL mediates abscisic acid - suppressed gene exression in barey aleurone layers. Plant Biol.1999,96:1767-1772
Beruter J&Studer F M E. Comparison of Sorbitol transport in excised tissue discs and cortex tissue of intact apple fruit. Journal of Plant Physiology. 1995,146:95-102
Beruter J. Sugar accumulation and changes in the activities of related enzyme during development of apple fruit. Journal of Plant Physiology. 1985,121:331-334
Beruter J, Studer F M E&Ruedi P. Sorbitol and sucrose partitioning in the growing apple fruit. Journal of Plant Physiology. 1997,151:269-276
Beruter J. Effect of abscisic acid on sorbitol uptake in growing apple fruit. Journal of Experimental Botany. 1983,34:737-743
Beruter J&Studer F. M. E. The effect of girdling on carbohydrate portioning in the growing
apple fruit. Journal of Plant Physiology. 1997,51:277-285
Beruter J..Effect of abscisic on sorbitol uptake in growing apple fruits. Journal of Experimental Botany. 1983,34:737-743
Bowler C ,Neuhaus G, Yamagata H ,Cgua N-H. Cyclic GMP and calcium mediated phytochrome phototransduction [J] . Cell. 1994, 77: 73～81.
Bracho G.E. &Whitaker J.R. Purification and partial characterization of potato invertase and its endogenous proteinaceous inhibitor. Plant Physiology. 1990,92:386-394
Burch LR, Davies HV, Cuthbert EM & Machray G.C. Purification of soluble invertase from potato. Phytochemistry. 1992,131:1901-1904
Bussis D, Heineke D, Sonnewaid U, Willmitzer L, Raschke K & Heldt H W. Solute accumulation and decreased photosynthesis in leaves of potato plants expressing yeast derived invertase either in the apoplast, vacuole or cytosol. Planta. 1997,202:126-136
Burbridge. Characterization of the ABA deficienttomato mutant notabilis and its relationship with maize VP14[J]. Planta. 1999 ,17(4):427-431
Chen J Q &Black C C. Biochemical and immunological properties of alkaline invertase isolated from sprouting soybean hypocotyls. Archives in Biochemistry & Biophysics. 1992,295:61-69
Coombe B. G. The grape berry as a sink. ActaHorticulturae. 1989,239:149-158
Davies C.,Boss P.K. & Robinson S.P. Treatment of grape berries, a nonclimacteric fruit with a synthetic auxin, retards ripening and alters the expression of developmental regulated genes. Plant Physiology. 1997, 115:1155-1161
Davis C, Robinson S P. Sugar accumulation in grape berries. Plant Physiol .1996,111:275-283
Estruch J J, Beltran J P. Changes in invertase activities precede ovary growth induced by gibberellic acid in pisum sativum. Physiol Plant.1991,81: 319～326
Ehness R , Ecker M , Roitsch T. Glucose and stress independently regulate source and sink metabolism and defense mechanism via signal transduction pathways involving protein phosphorylation. Plant Cell. 1997,9: 1825～1841
Elliot K.J.,Butler W.O.,Dickinson C.D.,Konno Y.,Vedvick T.S.,Fitzmaurice L,Mirkov T.E.
Isolation and characterization of fruit vacuolar invertase genes from two tomato species and temporal differences in mRNA levels during fruit ripening. Plant Molecular. Biology. 1993,21:515-524
Famiani F.,Walker R.P.,Tecsi L.,Chen Z.H.,Proietti P. &Leegood R.C. An immunohistochemical study of the comparmentation of metabolism during the development of grape berries. Journal of Experimental Botany. 2000,51:675-683
Foyer C.H. The basis for source-sink interaction in leaves. Plant Physiology &Biochemistry. 1987,25:649-659
Ferreira PCG, The Arabidopsis functional homolog of the P34cdc2 protein kinase[J].Plant Cell.1991,3:531-540.
Frommer W.B.&Sonnewald U. Molecular analysis of carbon partitioning in solanaceous species. Journal of Experimental Botany. 1995,46:587-607
Geotz M & Roitsch T. The different pH optima and subtrate specifities of extracellular an vacuolar invertases from plants are determined by a single amino acid substitution. Plant Journal. 1999,20:707-711
Geota M.&Roitsch T. Identification of amino acid essential for enzymetic activity of plant invertase. Journal of Plant Physiology. 2000,157:581-585
Godt D.E.&Roitsch T. Regulation and tissue-specific distribution of mRNAs for three extracellular invertase isoenzymes of tomato suggests:an important function in establishing and maintaining sink metabolism. Plant Physiology. 1997,80:464-471
Goupil P.,Loncle D.,Druart N.,Bellettre A.&Rambour S. Influence of ABA on nitrate reductase activity and carbohydrate metabolism in chicory roots. Journal of Experimental Botany. 1998, 49:1855-1862
Greenland A.J. & Lewis D.H. The acid invertase activity of the developing third leaf of oat Ⅰ:Changes in activity of invertase and concentrations of ethanol-soluble carbohydrates. New Phytologist. 1981,88:265-277
Godt DE, Riegel A, Roitsch T. Regulation of sucrose synthase expression in chenopodium rubrum: characterization of sugar induced expression in photoautotrophic suspension
cultures and sink tissue specific expression in plants. J Plant Physiol.1995,146:231～236
Grabov A , leung J , Giraudat J , Blatt MR. Alteration of anion channel kinetics in wild2type and abi121 transgenic Nicotiana benthamiana guard cells by abscisic acid. Plant J. 1997,12:203～213
Ghosheh N S, Rayle D h, Carpita N C & Kaufman P B, Cell wall and enzyme changes during gravivespos of the leaf-sheet pulvinus of oat. Plant Physiology. 1990,157:581-585
Harper, J F. ,A calcium- dependent protein kinase with a regulatory domain similar to calmodulin[J]. Science 1993,252: 951～954.
Hawker J.S. Changes in the activities of enzymes concerned with sugar metabolism during the development of grape berries. Phytochemistry. 1969,8:9-17
Harmon AC, Yoo BC ,Mccaffery C. Pseudo substrate inhibition of CDPK, a protein kinases with a calmodulin - like domain [J] .Biochem. 1994 ,33: 7278～7287.
Hidaka et al. Molecular pharmacology of protein kinase. Annu rev Pharmacol Toxical, 1992.32:377-397
Hideaki Yamaguchi, Yoshinori Kanayama and Shohei Yamaki. Purification and properties of NAD-dependent sorbitol dehydrogenase from apple fruit. Plant Cell Physiol. 1994, 25(6):887-892
Hanks S K, et al. Protein kinases catalytic domain sequence database: identification of conserved features of primary structure and classification of family members[J]. Methods Enzymol, 1991, 200: 38-62.
Huber J L A, Huber S C. Site - specific serine phosphorylation of spinach leaf sucrose-phosphate synthase. Biochem J. 1992,28:877-882
Huber S C, Huber J L, Pao-Chi Liao, et al. Phosphorylation of serine-15 of maize leaf sucrose synthase. Plant Physiol. 1992,99:1275-1278
Hurley J R, Dean A M, Sohl J L, Koshland D E J, et al. Regulation of an enzyme by phosphorylation at the active site. Science. 1990,249:1012-1016
Isla M. I., Salerno G.,Pontis H.,Vattuone MA.& Sampietro A.R. Purification and properties of the soluble acid invertase fron Oryza Sativa. Phytochemistry. 1995,38:321-325
Islam M.S.,Matsui T.,&Yoshid Y. Carbohydrate content and the activity of sucrose synthase, sucrose phosphate and acid invertase in different tomato cultivars during fruit development. Scientia Horticulturae. 1996,65:125-136
Iwatsubo. NakagawaH. OguraN. HirabayashiY. &SatoT. Acid invertase of melon fruit: immunochemical detection of acid invertase. Plant Cell&Physiology. 1992,33(8):1127-1133
Jang J C& Sheen J. Sugar sensing in higher plants. The Plant Cell.1994,6:1665-1679
Kato T , Kubota S . Properties of invertase in sugar storage tissues of citrus fruit and changes in their activities during maturation. Physiol Plant. 1978.42:67～72
Kim J.Y.,Aline M, Sylvain G, Mahe A, Guy S, Brangeon J, Roche O, Chourey P S&Prioul J L. Characterization of two members of the maize gene family, Incw3 and Incw4, encoding cell-wall invertase. Gene. 2000,245:89-102
Kingstom S A H &Pollock C J. Soluble acid invertase activity in leaves is independent of species differences in leaf carbohydrates, diurnal sugar profiles and path of phloem loading. New Phytologist, 1999. 139:293-292
Klann E M, Cheltelat R T&Bennett B. Antisense acid invertase(TIV1)gene alters soluble sugar composition and size in transgenic tomato fruit. Plant Physiology, 1996.112:1321-1330
Knight J S & Gray J C. Expression of genes encoding the tobacco chloropolast phosphate translocator is not light-regulated and is repressed by sucrose. Molecular & General Genetic, 1994.242:586-594
Kobashi K, Gemma H&Iwahori S. Sugar accumulation in peach fruit as affected by abscisic acid(ABA)treatment in relation to some sugar metabolizing enzymes. Journal. of the Japanese Society for Horticultural Science, 1999.68:465-470
Leung J, Micehelle B D. Arabidopsis ABA response gene ABI1:feature of a calcium-modulated protein phosphatase. Science, 1994,264:1448-1451
Li J , Wang XQ , Watson MB , Assmann SM. Regulation of abscisic acid induced stomatal closure and anion channels by guard cell AAPK kinase. Science. 2000,287 : 300～303
Lowell C A , Tomlinson P T. Sucrose-metabolizing enzymes in transport tissues and adjiacent sink structures in developing citrus fruit . Plant Physiol. 1989,90:1394～
1402
Leigh R A, Reea T, Fuller WA. The localization of acid invertase activity and sucrose in the vacuoles of storage roots of beet root (Beta vulgaris) . Biochem J, 1979.178:539～547
Miron D, Schaffer A. Sucrose phosphate synthase, sucrose synthase and invertase activities in developing fruit of Lycopersicon esculintum Mill. And sucrose accumulation in Lycopersicon hirsutum humb and bonp. Plant Physiology. 1991, 95:623-627
Nishihama R ,Baanon H, Shibata W, et al. Plant homologues of components of MAPK(mitogen - activated protein kinase) signal pathways in yeast and animal cell [J] . Plant Cell Physiol.1995 ,36(5) :7479～7486.
Patrick J W. Phloem unloading: sieve element unloading and post-sieve element transport. Annual of Review Plant Physiology and Plant Molecular Biology. 1997,48:191-222
Palejwala V A, Parikh H R, Modi V V, The role of abscisic acid in the ripening of grapes. Physiol Plant. 1985,65:498-502
Pei ZM , Murata Y, Benning G, Thomine S , Klusener B ,Allen GJ , Grill E , Schroeder JL. Calcium channels activated by hydrogen peroxide mediate abscisic acid signalling in guard cells. Nature. 2000,406 : 731～734
Riesmeier JW, Hirner B , Frommer WB. Expression of the sucrose transporter from potato correlates with the sink2to2source transition in leaves. Plant Cell, 1993 ,5:1591～1598
Renner R , Schuler K, Sonnewald U. Soluble acid invertase determines the hexose-to-sucrose ratio in cold- stored potato tubers. Planta. 1996,198 :246～252
Roberts DM, Calcium-modulated proteins: Targets of intracellular calcium signals in higher plants. Annue Rev plant Physiol, Plant Mol Bio.1992,43:375
Schroeder J I, Kwak JM, Allen GJ. Guard cell abscisic acid signalling and engineering drought hardiness in plants. Nature. 2001, 410 : 327～330
Stone JM, Walker JC , Plant protein kinase families and signal transduction[J]. Plant Physiol. 1995,105: 451～457
Sylvie L , Eckhard B , Hanjo H et al. The dual function of sugar carriers : transport
and sugar sensing. Plant Cell.1999,11:707～726
Smeekens S. Sugar2Induced signal transduction in plants. Annu Rev Plant Physiol Plant Mol Biol. 2000,51:49～82
Smith RD , Walker JC. Plant protein phosphatases. A nnu Rev Plant Physiol Plant Mol Biol.1996,47:101～125
Sturm A. Moleculor characterization and functional analysis of sucrose-cleaving enzymes in carrot. Journal of Experimental Botany. 1996,47:1187-1192
Strum A. Invertase:primary structures, functions and roles in plant development and sucrose portioning. Plant Physiology. 1999, 121:1-7
Takeda S, Mano S, Ohto M, et al. Inhibitors of protein phosphatase 1 and 2A block the sugar inducible gene expression in plants. Plant Physiol, 1994,106:567～574
Tymowska L Z&Kreis M. The plant invertase: physiology, biochemistry and molecular biology. Advance in Botany Research. 1998,28:71-117
Trewavas A J, Malho R. Signal perception and transduction:the origin of the phenotype. Plant Cell.1997,9:1181-1195
Unger C, Hardegger M, Lienhard S &Strum A. cDNA cloning of carrot soluble acid β -fructofuranosidase from Daucus carota. European Journal of Biochemistry. 204:915-921
Vendrell M. &Buesa, 1989, Relationship between ABA content and ripening of apples. Acta Horticulturae. 1994,258:389-396
Verhey S D, Lomax T L. Signal transduction in vascular plants. J Plant Growth Regul. 1993, 12: 167-169
Walker R P&Pollock C J. The Purification and characterization of soluble acid invertase from coleptile of wheat. Journal of Experimental Botany. 1993,44:1029-1037
Weber H, A role for sugar transporters during seed development: molecular characterization of a hexose and a sucrise carrier in fava bean seeds. The Plant Cell.1997,9:895-908
Wei L, et al. Protein kinase superfamily-comparisons of sequence data with three-dimensional structures[J].Curr. Opin. Struct. Biol.1994, (4): 450-455.
Weschke W. Panitz R, Gubatz S, Wang Q, Radchuk R, Weber H & Wobus U. The role of invertases
and hexose transporters in controlling sugar ratios in maternal and filial tissues of barley caryopses during early development. Plant Journal. 2003,33:395-411
Winter H & Huber S C. Regulation of sucrose and regulation of activity of key enzymes. Critical Reviews In Plant Science. 2000,19(1):31-67
Woodson W R & Wang H. Invertases of carnation petals: partial purification ,characterization and changes in activity during petal growth. Physiologia Plantarum. 1987,71:224-228
Wu L L, Song I, Kim D &Kaufman P B. Molecular basis of the increase in invertase activity elicited by gravistimulation invertase oat shoot pulvini. Journal of Plant Physiology. 1993,142:179-183
Xiao W Y, Sheen J & Jang J C. The role of hexokinase in plant sugar transduction and growth and development. Plant Molecular Biology. 2000,44(4): 451-461
Yamaki S. Sorbitol oxidase converting sorbitol to glucose in apple leaf. Plant & Cell Physiol. 1980,21:591-599
Yamaki S.& Ishikawa K. Role of four sorbitol related enzymes and invertase in seasonal alteration of sugar metabolism in apple tissue. Journal of American Society Horticultural Science, 1986,111:134-137
Yelle S Chtelate R T, Dorasis M, Devema J W & Bennett A B. Sink metabolism in tomato fruit Ⅳ:genetic and bilchemical analysis of sucrose accumulation. Plant Physiology. 1991,95:1026-1035
Yin J H, Gao F F & Hu G B. The regulation of litchi maturation and coloration abscisic acid and ethylene. Acta Horticulture. 2001,558:293-296
Zhang D P, Lu Y M, Wang Y Z, Duan C Q & Yan H Y. Acid invertase is predominantly localized to cell walls of both partically symplasmically isolated sieve element/companion cell complex and parenchyma cells in developing apple fruits. Plant Cell & Environment. 2001,24:691-702
Zrenner R, Salanoubat M & Willmitzer L. Evidence of the crucial role of sucrose synthase for sink strength using transgenic potato plants. Plant Journal. 1995,7:97-107
Zrenner R, Schuler K & Sonnewald V. Soluble acid invertase determines the hexose to
sucrose ratio in cold-stored potata tubers. Planta. 1996,198:246-25