Advertisement

Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Cytokinin oxidase PpCKX1 plays regulatory roles in development and enhances dehydration and salt tolerance in Physcomitrella patens

Abstract

Key message

PpCKX1 localizes to vacuoles and is dominantly expressed in the stem cells. PpCKX1 regulates developmental changes with increased growth of the rhizoid and enhances dehydration and salt tolerance.

Abstract

Cytokinins (CKs) are plant hormones that regulate plant development as well as many physiological processes, such as cell division, leaf senescence, control of shoot/root ratio, and reproductive competence. Cytokinin oxidases/dehydrogenases (CKXs) control CK concentrations by degradation, and thereby influence plant growth and development. In the moss Physcomitrella patens, an evolutionarily early divergent plant, we identified six putative CKXs that, by phylogenetic analysis, form a monophyletic clade. We also observed that ProPpCKX1:GUS is expressed specifically in the stem cells and surrounding cells and that CKX1 localizes to vacuoles, as indicated by Pro35S:PpCKX1-smGFP. Under normal growth conditions, overexpression of PpCKX1 caused many phenotypic changes at different developmental stages, and we suspected that increased growth of the rhizoid could affect those changes. In addition, we present evidence that the PpCKX1-overexpressor plants show enhanced dehydration and salt stress tolerance. Taken together, we suggest that PpCKX1 plays regulatory roles in development and adaptation to abiotic stresses in this evolutionarily early land plant species.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Abbreviations

FM4-64:

N-(3-Triethylammoniumpropyl)-4-(6-(4-(diethylamino) phenyl) hexatrienyl pyridinium dibromide

iP:

N6-Isopentenyladenine

PpCKX:

Physcomitrella patens Cytokinin oxidase

qRT-PCR:

Quantitative real time-PCR

WT:

Wild-type

References

  1. Aki SS, Mikami T, Naramoto S, Nishihama R, Ishizaki K, Kojima M, Takebayashi Y, Sakakibara H, Kyozuka J, Kohchi T, Umeda M (2019) Cytokinin Signaling Is Essential for Organ Formation in Marchantia polymorpha. Plant and Cell Physiol 60(8):1842–1854

  2. Ashikari M et al (2005) Cytokinin oxidase regulates rice grain production. Science 309:741–745

  3. Ashton NW, Cove DJ (1977) The isolation and preliminary characterisation of auxotrophic and analogue resistant mutants of the moss, Physcomitreila patens. Mol Gen Genet 154:87–95

  4. Avalbaev AM, Somov KA, Yuldashev RA, Shakirova FM (2012) Cytokinin oxidase is key enzyme of cytokinin degradation. Biochemistry (Moscow) 77:1354–1361

  5. Cho SH, Quatrano RS, Shin JS (2007) Transgenesis of Physcomitrella patens. Transgenic Plant Journal 1:99–103

  6. Coudert Y, Palubicki W, Ljung K, Novak O, Leyser O, Harrison CJ (2015) Three ancient hormonal cues co-ordinate shoot branching in a moss. eLife 4:e06808

  7. Dym O, Eisenberg D (2001) Sequence-structure analysis of FAD-containing proteins. Protein Sci 10:1712–1728

  8. Farrow SC, Emery RJN (2012) Concurrent profiling of indole-3-acetic acid, abscisic acid, and cytokinins and structurally related purines by high-performance-liquid-chromatography tandem electrospray mass spectrometry. Plant Methods 8(1):42

  9. Frank W, Ratnadewi D, Reski R (2005) Physcomitrella patens is highly tolerant against drought, salt and osmotic stress. Planta 220:384–394

  10. Frebort I, Kowalska M, Hluska T, Frebortova J, Galuszka P (2011) Evolution of cytokinin biosynthesis and degradation. J Exp Bot 62:2431–2452

  11. Frebortova J, Greplova M, Seidl MF, Heyl A, Frebort I (2015) Biochemical characterization of putative adenylate dimethylallyltransferase and cytokinin dehydrogenase from Nostoc sp. PCC 7120. PLoS ONE 10:0138468

  12. Gao SP et al (2014) CYTOKININ OXIDASE/DEHYDROGENASE4 integrates cytokinin and auxin signaling to control rice crown root formation. Plant Physiol 165:1035–1046

  13. Gu R, Fu J, Guo S, Duan F, Wang Z, Mi G, Yuan L (2010) Comparative expression and phylogenetic analysis of maize cytokinin dehydrogenase/oxidase (CKX) gene family. J Plant Growth Regul 29:428–440

  14. Ha S, Vankova R, Yamaguchi-Shinozaki K, Shinozaki K, Tran LS (2012) Cytokinins: metabolism and function in plant adaptation to environmental stresses. Trends Plant Sci 17:172–179

  15. Joshi MV, Loria R (2007) Streptomyces turgidiscabies possesses a functional cytokinin biosynthetic pathway and produces leafy galls. Mol Plant Microbe Interact 20:751–758

  16. Kamisugi Y et al (2006) The mechanism of gene targeting in Physcomitrella patens: homologous recombination, concatenation and multiple integration. Nucleic Acids Res 34:6205–6214

  17. Kofuji R, Hasebe M (2014) Eight types of stem cells in the life cycle of the moss Physcomitrella patens. Curr Opin Plant Biol 17:13–21

  18. Komatsu K, Nishikawa Y, Ohtsuka T, Taji T, Quatrano RS, Tanaka S, Sakata Y (2009) Functional analyses of the ABI1-related protein phosphatase type 2C reveal evolutionarily conserved regulation of abscisic acid signaling between Arabidopsis and the moss Physcomitrella patens. Plant Mol Biol 70:327–340

  19. Komatsu K et al (2013) Group A PP2Cs evolved in land plants as key regulators of intrinsic desiccation tolerance. Nat Commun 4:2219

  20. Kowalska M et al (2010) Vacuolar and cytosolic cytokinin dehydrogenases of Arabidopsis thaliana: heterologous expression, purification and properties. Phytochemistry 71:1970–1978

  21. Lindner A-C et al (2014) Isopentenyltransferase-1 (IPT1) knockout in Physcomitrella together with phylogenetic analyses of IPTs provide insights into evolution of plant cytokinin biosynthesis. J Exp Bot 65:2533–2543

  22. Liu Z et al (2013) Identification, expression, and comparative genomic analysis of the IPT and CKX gene families in Chinese cabbage (Brassica rapa ssp. pekinensis). BMC Genomics 14:594

  23. Malito E, Coda A, Bilyeu KD, Fraaije MW, Mattevi A (2004) Structures of michaelis and product complexes of plant cytokinin dehydrogenase: implications for flavoenzyme catalysis. J Mol Biol 341:1237–1249

  24. Menand B, Yi K, Jouannic S, Hoffmann L, Ryan E, Linstead P, Schaefer DG, Dolan L (2007) An ancient mechanism controls the development of cells with a rooting function in land plants. Science 316:1477–1480

  25. Mok DW, Mok MC (2001) Cytokinin metabolism and action. Annu Rev Plant Physiol Plant Mol Biol 52:89–118

  26. Nishiyama R et al (2011) Analysis of cytokinin mutants and regulation of cytokinin metabolic genes reveals important regulatory roles of cytokinins in drought, salt and abscisic acid responses, and abscisic acid biosynthesis. Plant Cell 23:2169–2183

  27. Oda Y et al (2009) Microtubules regulate dynamic organization of vacuoles in Physcomitrella patens. Plant Cell Physiol 50:855–868

  28. Peshev D, Vergauwen R, Moglia A, Hideg E, Van den Ende W (2013) Towards understanding vacuolar antioxidant mechanisms: a role for fructans? J Exp Bot 64:1025–1038

  29. Pils B, Heyl A (2009) Unraveling the evolution of cytokinin signaling. Plant Physiol 151:782–791

  30. Schaefer D, Zryd JP, Knight CD, Cove DJ (1991) Stable transformation of the moss Physcomitrella patens. Mol Gen Gene 226:418–424

  31. Schmulling T, Werner T, Riefler M, Krupkova E, Bartrina y Manns I (2003) Structure and function of cytokinin oxidase/dehydrogenase genes of maize, rice, Arabidopsis and other species. J Plant Res 116:241–252

  32. Tam TH, Catarino B, Dolan L (2015) Conserved regulatory mechanism controls the development of cells with rooting functions in land plants. Proc Natl Acad Sci USA 112:E3959–E3968

  33. von Schwartzenberg K (2006) Moss biology and phytohormones–cytokinins in Physcomitrella. Plant Biol (Stuttg) 8:382–388

  34. von Schwartzenberg K, Nunez MF, Blaschke H, Dobrev PI, Novak O, Motyka V, Strnad M (2007) Cytokinins in the bryophyte Physcomitrella patens: analyses of activity, distribution, and cytokinin oxidase/dehydrogenase overexpression reveal the role of extracellular cytokinins. Plant Physiol 145:786–800

  35. Vyroubalova S et al (2009) Characterization of new maize genes putatively involved in cytokinin metabolism and their expression during osmotic stress in relation to cytokinin levels. Plant Physiol 151:433–447

  36. Werner T, Schmulling T (2009) Cytokinin action in plant development. Curr Opin Plant Biol 12:527–538

  37. Werner T, Motyka V, Strnad M, Schmulling T (2001) Regulation of plant growth by cytokinin. Proc Natl Acad Sci USA 98:10487–10492

  38. Werner T, Motyka V, Laucou V, Smets R, Van Onckelen H, Schmulling T (2003) Cytokinin-deficient transgenic Arabidopsis plants show multiple developmental alterations indicating opposite functions of cytokinins in the regulation of shoot and root meristem activity. Plant Cell 15:2532–2550

  39. Werner T, Nehnevajova E, Köllmer I, Novák O, Strnad M, Krämer U, Schmülling T (2010) Root-specific reduction of cytokinin causes enhanced root growth, drought tolerance, and leaf mineral enrichment in Arabidopsis and tobacco. Plant Cell 22:3905–3920

  40. Wolters H, Jurgens G (2009) Survival of the flexible: hormonal growth control and adaptation in plant development. Nat Rev Genet 10:305–317

  41. Zwack PJ, Rashotte AM (2015) Interactions between cytokinin signalling and abiotic stress responses. J Exp Bot 66:4863–4871

Download references

Acknowledgements

We thank the Korea Basic Science Institute (Seoul) for helping with cytokinin analysis with HPLC-MS/MS. This research was supported by two grants (PJ011065 and PJ01367001) from the Next-Generation BioGreen 21 Program funded by the Rural Development Administration, Republic of Korea. This work was also partially supported by Korea University.

Author information

Correspondence to Jeong Sheop Shin.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Communicated by Youn-Il Park.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary file1 (DOCX 3218 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Hyoung, S., Cho, S.H., Chung, J.H. et al. Cytokinin oxidase PpCKX1 plays regulatory roles in development and enhances dehydration and salt tolerance in Physcomitrella patens. Plant Cell Rep 39, 419–430 (2020). https://doi.org/10.1007/s00299-019-02500-3

Download citation

Keywords

  • Abiotic stress tolerance
  • Cytokinin oxidase
  • Development
  • Evolution
  • Hormone
  • Physcomitrella patens