Journal of Plant Research

, Volume 116, Issue 3, pp 241–252 | Cite as

Structure and function of cytokinin oxidase/dehydrogenase genes of maize, rice, Arabidopsis and other species

  • Thomas SchmüllingEmail author
  • Tomáš Werner
  • Michael Riefler
  • Eva Krupková
  • Isabel Bartrina y Manns
JPR Symposium


Cytokinin oxidases/dehydrogenases (CKX) catalyze the irreversible degradation of the cytokinins isopentenyladenine, zeatin, and their ribosides in a single enzymatic step by oxidative side chain cleavage. To date the sequences of 17 fully annotated CKX genes are known, including two prokaryotic genes. The CKX gene families of Arabidopsis thaliana and rice comprise seven and at least ten members, respectively. The main features of CKX genes and proteins are summarized in this review. Individual proteins differ in their catalytic properties, their subcellular localization and their expression domains. The evolutionary development of cytokinin-catabolizing gene families and the individual properties of their members indicate an important role for the fine-tuned control of catabolism to assure proper regulation of cytokinin functions. The use of CKX genes as a tool in studies of cytokinin biology and biotechnological applications is discussed.


Arabidopsis thaliana Cytokinin dehydrogenase Cytokinin oxidase Plant hormones Rice Zea mays 


  1. Angra R, Mandahar CL, Gulati A (1990) The possible involvement of cytokinins in the pathogenicity of Helminthosporium maydis. Mycopathologia 109:177–182Google Scholar
  2. Arabidopsis Genome Initiative (2000) Analysis of the genome sequence of the flowering plant Arabidopsis thaliana. Nature 408:796–815PubMedGoogle Scholar
  3. Armstrong DJ (1994) Cytokinin oxidase and the regulation of cytokinin degradation. In: Mok DWS, Mok MC (eds) Cytokinin: chemistry, activity, and function. CRC, Boca Raton, pp 139–154Google Scholar
  4. Armstrong DJ, Firtel RA (1989) Cytokinin oxidase activity in the cellular slime mold Dictyostelium discoideum. Dev Biol 136:491–499Google Scholar
  5. Benková E, Witters E, Van Dongen W, Kolář J, Motyka V, Brzobohatý B, Van Onckelen H, Macháčková I (1999) Cytokinins in tobacco and wheat chloroplasts: occurrence and changes due to light/dark treatment. Plant Physiol 121:245–251CrossRefPubMedGoogle Scholar
  6. Bilyeu KD, Cole JL, Laskey JG, Riekhof WR, Esparza TJ, Kramer MD, Morris RO (2001) Molecular and biochemical characterization of a cytokinin oxidase from maize. Plant Physiol 125:378–386CrossRefPubMedGoogle Scholar
  7. Brownlee BG, Hall RH, Whitty CD (1975) 3-Methyl-2-butenal; an enzymatic degradation product of the cytokinin, N6-(Δ2-isopentenyl)adenine. Can J Biochem 53:37–41PubMedGoogle Scholar
  8. Burch LR, Horgan R (1989) The purification of cytokinin oxidase from Zea mays kernels. Phytochemistry 28:1313–1319CrossRefGoogle Scholar
  9. Burch LR, Horgan R (1992) Cytokinin oxidase and the degradative metabolism of cytokinins. In: Kamínek M, Mok DWS, Zažímalová E (eds) Physiology and biochemistry of cytokinins in plants. SPB Academic, The Hague, pp 29–32Google Scholar
  10. Chatfield JM, Armstrong DJ (1986) Regulation of cytokin oxidase activity in callus tissues of Phaseolus vulgaris L. cv. Great Northern. Plant Physiol 80:493–499Google Scholar
  11. Cheikh N, Jones RJ (1994) Disruption of maize kernel growth and development by heat stress: role of cytokinin/abscisic acid balance. Plant Physiol 106:45–51PubMedGoogle Scholar
  12. Crespi M, Messens E, Caplan AB, Van Montagu M, Desomer J (1992) Fasciation induction by the phytopathogen Rhodococcus fascians depends upon a linear plasmid encoding a cytokinin synthase gene. EMBO J 11:795–804PubMedGoogle Scholar
  13. Crespi M, Vereecke D, Temmerman W, Van Montagu M, Desomer J (1994) The fas operon of Rhodococcus fascians encodes new genes required for efficient fasciation of host plants. J Bacteriol 176:2492–2501PubMedGoogle Scholar
  14. Dietrich JT, Kamínek M, Blevins DG, Reinbott TM, Morris RO (1995) Changes in cytokinins and cytokinin oxidase activity in developing maize kernels and the effects of exogenous cytokinin on kernel development. Plant Physiol Biochem 33:327–336Google Scholar
  15. Doree M, Guern J (1973) Short-time metabolism of some exogenous cytokinins in Acer pseudoplatanus cells. Biochim Biophys Acta 304:611–619CrossRefPubMedGoogle Scholar
  16. Eklöf S, Astot C, Moritz T, Blackwell J, Olsson O, Sandberg G (1996) Cytokinin metabolites and gradients in wild type and transgenic tobacco with moderate cytokinin overproduction. Physiol Plant 98:333–344CrossRefGoogle Scholar
  17. Emanuelsson O, Heijne G von (2001) Prediction of organellar targeting signals. Biochim Biophys Acta 1541:114–119CrossRefPubMedGoogle Scholar
  18. Emanuelsson O, Nielsen H, Brunak S, Heijne G von (2000) Predicting subcellular localization of proteins based on their N-terminal amino acid sequence. J Mol Biol 300:1005–1016CrossRefPubMedGoogle Scholar
  19. Forsyth C, Staden J van (1987) Cytokinin metabolism in tomato plants. II. Metabolites of kinetin and benzyladenine in decapitated roots. Plant Growth Regul 6:277–292Google Scholar
  20. Fox JE, Dyson W, Sood D, McChesney JD (1972) Active forms of the cytokinins. In: Carr DJ (ed) Plant growth regulation. Springer, Berlin Heidelberg New York, pp 449–458Google Scholar
  21. Frébort I, Šebela M, Galuszka P, Werner T, Schmülling T, Peč P (2002) Cytokinin oxidase/cytokinin dehydrogenase assay: optimized procedures and applications. Anal Biochem 306:1–7CrossRefPubMedGoogle Scholar
  22. Galuszka P, Frébort I, Šebela M, Strnad M, Peč P (1999) Cytokinin oxidase: the key enzyme in the biodegradation of cytokinins. In: Strnad M, Peč P, Beck E (eds) Advances in regulation of plant growth and development. Peres, Prague, pp 39–48Google Scholar
  23. Galuszka P, Frébort I, Šebela M, Sauer P, Jacobsen S, Peč P (2001) Cytokinin oxidase or dehydrogenease? Mechanism of cytokinin degradation in cereals. Eur J Biochem 268:450–461CrossRefPubMedGoogle Scholar
  24. Gan S, Amasino RM (1995) Inhibition of leaf senescence by autoregulated production of cytokinin. Science 270:1986–1988PubMedGoogle Scholar
  25. Gerhäuser D, Bopp M (1990) Cytokinin oxidases in mosses. 2. Metabolism of kinetin and benzyladenine in vivo. J Plant Physiol 135:714–718Google Scholar
  26. Gordon ME, Letham DS, Parker CW (1974) The metabolism and translocation of zeatin in intact radish seedlings. Ann Bot 38:809–825Google Scholar
  27. Hare PD, Van Staden J (1994a) Cytokinin oxidase: biochemical features and physiological significance. Physiol Plant 91:128–136CrossRefGoogle Scholar
  28. Hare PD, Van Staden J (1994b) Inhibitory effect of thidiazuron on the activity of cytokinin oxidase isolated from soybean callus. Plant Cell Physiol 35:1121–1125Google Scholar
  29. Houba-Hérin N, Pethe C, d'Alayer J, Laloue M (1999) Cytokinin oxidase from Zea mays: purification, cDNA cloning and expression in moss protoplasts. Plant J 17:615–626PubMedGoogle Scholar
  30. Hwang I, Sheen J (2001) Two-component circuitry in Arabidopsis cytokinin signal transduction. Nature 413:383–389Google Scholar
  31. Jones RJ, Schreiber BMN (1997) Role and function of cytokinin oxidase in plants. Plant Growth Regul 23:123–134Google Scholar
  32. Jones RJ, Schreiber BMN, McNeil K, Brenner ML, Foxon G (1992) Cytokinin levels and oxidase activity during maize kernel development. In: Kamínek M, Mok DWS, Zažímalová E (eds) Physiology and biochemistry of cytokinins in plants. SPB Academic, The Hague, pp 29–32Google Scholar
  33. Kamínek M, Armstrong DJ (1990) Genotypic variation in cytokinin oxidase from Phaseolus callus cultures. Plant Physiol 93:1530–1538Google Scholar
  34. Kamínek M, Trčková M, Motyka V, Gaudinová A (1994) Role of cytokinins in control of wheat grain development and utilization of nutrients. Biol Plant 36:135Google Scholar
  35. Kamínek M, Motyka V, Vaňková R (1997) Regulation of cytokinin content in plant cells. Physiol Plant 101:689–700CrossRefGoogle Scholar
  36. Kaneko T, Nakamura Y, Wolk CP, Kuritz T, Sasamoto S, Watanabe A, Iriguchi M, Ishikawa A, Kawashima K, Kimura T, Kishida Y, Kohara M, Matsumoto M, Matsuno A, Muraki A, Nakazaki N, Shimpo S, Sugimoto M, Takazawa M, Yamada M, Yasuda M, Tabata S (2001) Complete genomic sequence of the filamentous nitrogen-fixing cyanobacterium Anabaena sp. strain PCC 7120 (supplement). DNA Res 8:205–213PubMedGoogle Scholar
  37. Kapoor K, Sharma VK (1981) Effect of growth-promoting chemicals on growth, nitrogen fixation and heterocyst frequency of a blue-green alga. Z Allg Mikrobiol 21:305–311PubMedGoogle Scholar
  38. Laloue M, Fox JE (1985) Characterization of an imine intermediate in the degradation of isopentenylated cytokinins by a cytokinin oxidase from wheat. In: Boop M, Knoop B, Rademacher W (eds) Abstracts of the 12th international conference on plant growth substances, Heidelberg, p 23Google Scholar
  39. Laloue M, Fox JE (1989) Cytokinin oxidase from wheat: partial purification and general properties. Plant Physiol 90:899–906Google Scholar
  40. Laloue M, Pethe C (1982) Dynamics of cytokinin metabolism in tobacco cells. In: Wareing PF (ed) Plant growth substances 1982. Academic Press, London, pp 186–195Google Scholar
  41. Leister D (2003) Chloroplast research in the genomic age. Trends Genet 19:47–56CrossRefPubMedGoogle Scholar
  42. Lejeune P, Prinsen E, Van Onckelen HA, Bernier G (1998). Hormonal control of ear abortion in a stress-sensitive maize (Zea mays) inbred. Aust J Plant Physiol 25:481–488Google Scholar
  43. Letham DS, Wilson MM, Parker CW, Jenkins ID, Macleod JK, Summons RE (1975) Regulators of cell division in plant tissue. XXIII. The identity of an unusual metabolite of 6-benzylaminopurine. Biochim Biophys Acta 399:61–70CrossRefPubMedGoogle Scholar
  44. Letham DS, Tao GQ, Parker CW (1982) An overview of cytokinin metabolism. In: Wareing PF (ed) Plant growth substances. Academic Press, London, pp 143–152Google Scholar
  45. Libreros-Minotta CA, Tipton PA (1995) A colorimetric assay for cytokinin oxidase. Anal Biochem 231:339–341CrossRefPubMedGoogle Scholar
  46. Lohrmann J, Sweere U, Zabaleta E, Bäuerle I, Keitel C, Kozma-Bognar L, Brennicke A, Kudla J, Schäfer E, Harter K (2001) The response regulator ARR2: a pollen-specific transcription factor involved in the expression of nuclear-encoded mitochondrial complex I genes. Mol Gen Genomics 265:2–13CrossRefGoogle Scholar
  47. Lynch J (1995) Root architecture and plant productivity. Plant Physiol 109:7–13PubMedGoogle Scholar
  48. Martin W, Rujan T, Richly E, Hansen A, Cornelsen S, Lins T, Leister D, Stoebe B, Hasegawa M, Penny D (2002) Evolutionary analysis of Arabidopsis, cyanobacterial, and chloroplast genomes reveals plastid phylogeny and thousands of cyanobacterial genes in the nucleus. Proc Natl Acad Sci USA 99:12246–12251CrossRefPubMedGoogle Scholar
  49. McGaw BA, Horgan R (1983) Cytokinin catabolism and cytokinin oxidase. Phytochemistry 22:1103–1105CrossRefGoogle Scholar
  50. McNeil KJ, Brenner ML, Jones RJ (1991) Effect of benzylamino-purine and thidiazuron on maize kernel development. Proc Plant Growth Regul Soc Am 17:16–17Google Scholar
  51. Miernyk JA (1979) Abscisic acid inhibition of kinetin nucleotide formation in germinating lettuce seeds. Physiol Plant 45:63–66Google Scholar
  52. Miernyk JA, Blaydes (1977) Short term metabolism of radioactive kinetin during lettuce seed germination. Physiol Plant 39:4–8Google Scholar
  53. Mills LJ, Van Staden J (1978) Extraction of cytokinins from maize, smut tumors of maize and Ustilago maydis cultures. Physiol Plant Pathol 13:73–80Google Scholar
  54. Mok MC (1994) Cytokinins and plant development: an overview. In Mok, DWS, Mok MC (eds) Cytokinins: Chemistry, activity, and function. CRC, Boca Raton, pp 155–166Google Scholar
  55. Mok DW, Mok MC (2001) Cytokinin metabolism and action. Annu Rev Plant Physiol Plant Mol Biol 52:89–118Google Scholar
  56. Morris RO (1995) Genes specifying auxin and cytokinin biosynthesis in prokaryotes. In: Davies PJ (ed) Plant hormones: physiology, biochemistry and molecular biology. Kluwer Academic, Dordrecht, pp 318–339Google Scholar
  57. Morris RO, Bilyeu KD, Laskey JG, Cheikh N (1999) Isolation of a gene encoding a glycosylated cytokinin oxidase from maize. Biochem Biophys Res Comm 255:328–333PubMedGoogle Scholar
  58. Motyka V, Kamínek M (1994) Cytokinin oxidase from auxin and cytokinin-dependent callus cultures of tobacco Nicotiana tabacum L.). J Plant Growth Regul 13:1–9Google Scholar
  59. Motyka V, Faiss M, Strnad M, Kamínek M, Schmülling T (1996) Changes in cytokinin content and cytokinin oxidase activity in response to derepression of ipt gene transcription in transgenic tobacco calli and plants. Plant Physiol 112:1035–1043PubMedGoogle Scholar
  60. Motyka V, Vaňková R, Čapková V, Petrášek J, Kamínek M, Schmülling T (2003) Cytokinin-induced upregulation of cytokinin oxidase activity in tobacco includes changes in enzyme glycosylation and secretion. Physiol Plant 117:11–21Google Scholar
  61. Mushegian AR, Koonin EV (1995) A putative FAD-binding domain in a distinct group of oxidases including a protein involved in plant development. Protein Sci 4:1243–1244PubMedGoogle Scholar
  62. Nakai K, Horton P (1999) PSORT: a program for detecting sorting signals in proteins and predicting their subcellular localization. Trends Biochem Sci 24:34–36PubMedGoogle Scholar
  63. Pačes V, Werstiuk E, Hall RH (1971) Conversion of N6-(Δ2-isopentenyl)adenosine to adenosine by enzyme activity in tobacco tissue. Plant Physiol 48:775–778Google Scholar
  64. Page RD (1996) TreeView: an application to display phylogenetic trees on personal computers. Comput Appl Biosci 12:357–358PubMedGoogle Scholar
  65. Palni LMS, Burch L, Horgan R (1988) The effect of auxin concentration on cytokinin stability and metabolism. Planta 174:231–234Google Scholar
  66. Parker CW, Letham DS (1973) Regulation of cell division in plant tissues. XVI. Metabolism of zeatin by radish cotyledons and hypocotyls. Planta 114:199–218Google Scholar
  67. Peeters N, Small I (2001) Dual targeting to mitochondria and chloroplasts. Biochim Biophys Acta 1541:54–63CrossRefPubMedGoogle Scholar
  68. Redig P, Schmülling T, Van Onckelen H (1996a) Analysis of cytokinin metabolism in ipt transgenic tobacco by liquid chromatography-tandem mass spectrometry. Plant Physiol: 112:141–148Google Scholar
  69. Redig P, Shaul O, Inze D, Van Montagu M, Van Onckelen H (1996b) Levels of endogenous cytokinins, indole-3-acetic acid and abscisic acid during the cell cycle of synchronized tobacco BY-2 cells. FEBS Lett 391:175–180PubMedGoogle Scholar
  70. Redig P, Motyka V, Van Onckelen H, Kamínek M (1997) Regulation of cytokinin oxidase activity in tobacco callus expressing the T-DNA ipt gene. Physiol Plant 99:89–96CrossRefGoogle Scholar
  71. Sasaki T, Matsumoto T, Yamamoto K, Sakata K, Baba T, Katayose Y, Wu J, Niimura Y, Cheng Z, Nagamura Y, Antonio BA, Kanamori H, Hosokawa S, Masukawa M, Arikawa K, Chiden Y, Hayashi M, Okamoto M, Ando T, Aoki H, Arita K, Hamada M, Harada C, Hijishita S, Honda M, Ichikawa Y, Idonuma A, Iijima M, Ikeda M, Ikeno M, Itoh S, Itoh T, Itoh Y, Iwabuchi A, Kamiya K, Karasawa W, Katagiri S, Kikuta A, Kobayashi N, Kono I, Machita K, Maehara T, Mizuno H, Mizubayashi T, Mukai Y, Nagasaki H, Nakashima, M, Nakama Y, Nakamichi Y, Nakamura M, Namiki N, Negishi M, Ohta I, Ono N, Saji S, Sakai K, Shibata M, Shimokawa T, Shomura A, Song J, Takazaki Y, Terasawa K, Tsuji K, Waki K, Yamagata H, Yamane H, Yoshiki S, Yoshihara R, Yukawa K, Zhong H, Iwama H, Endo T, Ito H, Hahn JH, Kim HI, Eun MY, Yano M, Jiang J, Gojobori T (2002) The genome sequence and structure of rice chromosome 1. Nature 420:312–316CrossRefPubMedGoogle Scholar
  72. Schmülling T (2001) CREam of cytokinin signalling: receptor identified. Trends Plant Sci 6:281–284CrossRefPubMedGoogle Scholar
  73. Schmülling T, Schäfer S, Romanov G (1997) Cytokinins as regulators of gene expression. Physiol Plant 100:505–519CrossRefGoogle Scholar
  74. Schwacke R, Schneider A, Graaff E van der, Fischer K, Catoni E, Desimone M, Frommer WB, Flügge UI, Kunze R (2003) ARAMEMNON, a novel database for Arabidopsis integral membrane proteins. Plant Physiol 131:16–26CrossRefPubMedGoogle Scholar
  75. Shudo K (1994) Chemistry of phenylurea cytokinins. In: Mok DWS, Mok MC (eds) Cytokinins: chemistry, activity, and function. CRC, Boca Raton, pp 35–42Google Scholar
  76. Singh S, Palni LMS, Letham DS (1992) Cytokinin biochemistry in relation to leaf senescence. V. Endogeneous cytokinin levels and metabolism of zeatin riboside in leaf discs from green and senescent tobacco (Nicotiana rustica) leaves. J Plant Physiol 139:279–283Google Scholar
  77. Stock AM, Robinson VL, Goudreau PN (2000) Two-component signal transduction. Annu Rev Biochem 69:183–215Google Scholar
  78. Takei K, Sakakibara H, Sugiyama T (2001) Identification of genes encoding adenylate isopentenyltransferase, a cytokinin biosynthesis enzyme, in Arabidopsis thaliana. J Biol Chem 276:26405–26410CrossRefPubMedGoogle Scholar
  79. Terrine C, Laloue M (1980) Kinetics of N6-(Δ2-isopentenyl)adenosine degradation in tobacco cells: evidence of regulatory mechanism under the control of cytokinins. Plant Physiol 65:1090–1095Google Scholar
  80. Thomas JC, Katterman RF (1986) Cytokinin activity induced by thidiazuron. Plant Physiol 81:681–683Google Scholar
  81. Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680PubMedGoogle Scholar
  82. Turner JE, Mok MC, Mok DWS (1985) Zeatin metabolism in fruits of Phaseolus: comparison between embryos, seed coat and pod tissues. Plant Physiol 79:321–322Google Scholar
  83. Van Kast CA, Laten H (1987) Cytokinin utilization by adenine requiring mutants of the yeast Saccharomyces cerevisiae. Plant Physiol 83:726–727Google Scholar
  84. Vision TJ, Brown DG, Tanksley SD (2000) The origins of genomic duplications in Arabidopsis. Science 290:2114–2117PubMedGoogle Scholar
  85. Wang J, Letham DS (1995) Cytokinin oxidase: purification by affinity chromatography and activation by caffeic acid. Plant Sci 112:161–166CrossRefGoogle Scholar
  86. Werner T, Motyka V, Strnad M, Schmülling T (2001) Regulation of plant growth by cytokinin. Proc Natl Acad Sci USA 98:10487–10492CrossRefPubMedGoogle Scholar
  87. Whitty CD, Hall RH (1974) A cytokinin oxidase in Zea mays. Can J Biochem 52:787–799Google Scholar
  88. Wilkins MR, Lindskog I, Gasteiger E, Bairoch A, Sanchez JC, Hochstrasser DF, Appel RD (1997) Detailed peptide characterisation using PEPTIDEMASS: a World-Wide Web accessible tool. Electrophoresis 18:403–408PubMedGoogle Scholar
  89. Yamada H, Suzuki T, Terada K, Takei K, Ishikawa K, Miwa K, Yamashino T, Mizuno T (2001) The Arabidopsis AHK4 histidine kinase is a cytokinin-binding receptor that transduces cytokinin signals across the membrane. Plant Cell Physiol 42:1017–1023Google Scholar
  90. Yang SH, Yu H, Goh CJ (2002) Isolation and characterization of the orchid cytokinin oxidase DSCKX1 promoter. J Exp Bot 53:1899–1907CrossRefPubMedGoogle Scholar
  91. Yang SH, Yu H, Goh CJ (2003) Functional characterisation of a cytokinin oxidase gene DSCKX1 in Dendrobium orchid. Plant Mol Biol 51:237–248CrossRefPubMedGoogle Scholar
  92. Zhang R, Zhang X, Wang J, Letham DS, Mckinney SA, Higgins TJV (1995) The effect of auxin on cytokinin levels and metabolism in transgenic tobacco tissues expressing an ipt gene. Planta 196:84–94Google Scholar

Copyright information

© The Botanical Society of Japan and Springer-Verlag  2003

Authors and Affiliations

  • Thomas Schmülling
    • 1
    Email author
  • Tomáš Werner
    • 1
  • Michael Riefler
    • 1
  • Eva Krupková
    • 1
  • Isabel Bartrina y Manns
    • 1
  1. 1.Institute of Biology, Applied GeneticsFU BerlinBerlinGermany

Personalised recommendations