Effect of cytokinins on expression of radish CLE genes


CLE (CLAVATA3/ENDOSPERM SURROUNDING REGION) peptides are peptide phytohormones playing the important role in the regulation of various type meristem development and also in the interaction between plants and parasites and symbionts. At the same time, the interaction of CLE peptides with other phytohormones, participating in these processes, is essentially unstudied. Using real-time RT-PCR, we analyzed expression of some genes encoding CLE peptides in radish (Raphanus sativus var. radicula Pers.) under normal conditions and after treatment with the cytokinin 6-benzyladenine. Even after short-term (30 min) action of cytokinin, expression of group A CLE genes was manifold suppressed, whereas cytokinin did not affect expression of group B CLE genes. Radish lines contrasting in the trait “spontaneous tumor formation” demonstrated similar dynamics of CLE gene expression in response to cytokinin treatment but differed in the levels of their expression. A possibility of interaction between CLE peptides and cytokinins in the spontaneous development of tumors in radish lines is discussed. The important part of the work was the selection of optimal reference genes for the analysis of radish gene expression by real-time RT-PCR method. Most stable expression was observed for the genes of ubiquitin (RsUBQ) and glyceraldehydes-3-phosphate dehydrogenase (RsGAPDH).

This is a preview of subscription content, access via your institution.



apical meristem








root apical meristem


reverse transcription with subsequent polymerase chain reaction in real time


shoot apical meristem


Tracheary Element Differentiation Inhibitory Factor


WUSHEL-Related Homeobox


  1. 1.

    Schoof, H., Lenhard, M., Haecker, A., Mayer, K.F., Jürgens, G., and Laux, T., The Stem Cell Population of Arabidopsis Shoot Meristems Is Maintained by a Regulatory Loop between the CLAVATA and WUSCHEL Genes, Cell, 2000, vol. 100, pp. 635–644.

    PubMed  Article  CAS  Google Scholar 

  2. 2.

    Bonello, J.F., Sevilla-Lecoq, S., Berne, A., Risueño, M.C., Dumas, C., and Rogowsky, P.M., ESR Proteins Are Secreted by the Cells of the Embryo Surrounding Region, J. Exp. Bot., 2002, vol. 53, pp. 1559–1568.

    PubMed  Article  CAS  Google Scholar 

  3. 3.

    Stahl, Y. and Simon, R., Plant Primary Meristems: Shared Functions and Regulatory Mechanisms, Curr. Opin. Plant Biol., 2010, vol. 13, pp. 53–58.

    PubMed  Article  CAS  Google Scholar 

  4. 4.

    Mitchum, M.G., Wang, X., and Davis, E.L., Diverse and Conserved Roles of CLE Peptides, Curr. Opin. Plant Biol., 2008, vol. 11, pp. 75–81.

    PubMed  Article  CAS  Google Scholar 

  5. 5.

    Jun, J., Fiume, E., Roeder, A.H., Meng, L., Sharma, V.K., Osmont, K.S., Baker, C., Ha, C.M., Meyerowitz, E.M., Feldman, L.J., and Fletcher, J.C., Comprehensive Analysis of CLE Polypeptide Signaling Gene Expression and Overexpression Activity in Arabidopsis, Plant Physiol., 2010, vol. 154, pp. 1721–1736.

    PubMed  Article  CAS  Google Scholar 

  6. 6.

    Stahl, Y., Wink, R.H., Ingram, G.C., and Simon, R., A Signaling Module Controlling the Stem Cell Niche in Arabidopsis Root Meristems, Curr. Biol., 2009, vol. 19, pp. 909–914.

    PubMed  Article  CAS  Google Scholar 

  7. 7.

    Hirakawa, Y., Kondo, Y., and Fukuda, H., TDIF Peptide Signaling Regulates Vascular Stem Cell Proliferation via the WOX4 Homeobox Gene in Arabidopsis, Plant Cell, 2010, vol. 22, pp. 2618–2629.

    PubMed  Article  CAS  Google Scholar 

  8. 8.

    Ito, Y., Nakanomyo, I., Motose, H., Iwamoto, K., Sawa, S., Dohmae, N., and Fukuda, H., Dodeca-CLE Peptides as Suppressors of Plant Stem Cell Differentiation, Science, 2006, vol. 313, pp. 842–845.

    PubMed  Article  CAS  Google Scholar 

  9. 9.

    Whitford, R., Fernandez, A., de Groodt, R., Ortega, E., and Hilson, P., Plant CLE Peptides from Two Distinct Functional Classes Synergistically Induce Division of Vascular Cells, Proc. Natl. Acad. Sci. USA, 2008, vol. 105, pp. 18625–18630.

    PubMed  Article  CAS  Google Scholar 

  10. 10.

    Yaginuma, H., Hirakawa, Y., Kondo, Y., Ohashi-Ito, K., and Fukuda, H., A Novel Function of TDIF-Related Peptides: Promotion of Axillary Bud Formation, Plant Cell Physiol., 2011, vol. 52, pp. 1354–1364.

    PubMed  Article  CAS  Google Scholar 

  11. 11.

    Mortier, V., den Herder, G., Whitford, R., van de Velde, W., Rombauts, S., d’Haeseleer, K., Holsters, M., and Goormachtig, S., CLE Peptides Control Medicago truncatula Nodulation Locally and Systemically, Plant Physiol., 2010, vol. 153, pp. 222–237.

    PubMed  Article  CAS  Google Scholar 

  12. 12.

    Mortier, V., de Wever, E., Vuylsteke, M., Holsters, M., and Goormachtig, S., Nodule Numbers Are Governed by Interaction between CLE Peptides and Cytokinin Signaling, Plant J., 2012, doi 10.1111/j.1365-313X.2011.04881.x

    Google Scholar 

  13. 13.

    Dodueva, I.E., Yurlova, E.V., Osipova, M.A., and Lutova, L.A., CLE Peptides Are Universal Regulators of Meristem Development, Russ. J. Plant Physiol., 2012, vol. 59, pp. 14–27.

    Article  CAS  Google Scholar 

  14. 14.

    Narbut, S.I., Voilokov, A.V., Rakhman, M.I., and Maksimenko, O.E., Biometric Analysis of Spontaneous Tumor Formation in Radish Inbreeding Lines, Genetika (Moscow), 1995, vol. 31, pp. 1268–1271.

    Google Scholar 

  15. 15.

    Matveeva, T.V., Frolova, N.V., Smets, R., Dodueva, I.E., Buzovkina, I.S., van Onckelen, H., and Lutova, L.A., Hormonal Control of Tumor Formation in Radish, J. Plant Growth Regul., 2004, vol. 23, pp. 37–43.

    Article  CAS  Google Scholar 

  16. 16.

    Buzovkina, I.S., Kneshke, I., and Lutova, L.A., Simulation of Tumor Formation In Vitro in Radish Lines and Hybrids, Genetika (Moscow), 1993, vol. 29, pp. 1002–1008.

    Google Scholar 

  17. 17.

    Il’ina, E.L., Dodueva, I.E., Ivanova, N.M., and Lutova, L.A., The Effect of Cytokinins on In Vitro Cultured Inbred Lines of Raphanus sativus var. radicula Pers. with Genetically Determined Tumorigenesis, Russ. J. Plant Physiol., 2006, vol. 53, pp. 514–522.

    Article  Google Scholar 

  18. 18.

    Czechowski, T., Stitt, M., Altmann, T., Udvardi, M.K., and Scheible, W.R., Genome-Wide Identification and Testing of Superior Reference Genes for Transcript Normalization in Arabidopsis, Plant Physiol., 2005, vol. 139, pp. 5–17.

    PubMed  Article  CAS  Google Scholar 

  19. 19.

    Reid, K.E., Olsson, N., Schlosser, J., Peng, F., and Lund, S.T., An Optimized Grapevine RNA Isolation Procedure and Statistical Determination of Reference Genes for Real-Time RT-PCR during Berry Development, BMC Plant Biol., 2006, vol. 14, p. 27, doi 10.1186/1471-2229-6-27

    Article  Google Scholar 

  20. 20.

    Vandesompele, J., de Preter, K., Pattyn, F., Poppe, B., van Roy, N., de Paepe, A., and Speleman, F., Accurate Normalization of Real-Time Quantitative RT-PCR Data by Geometric Averaging of Multiple Internal Control Genes, Genome Biol., 2002, vol. 18, R0034.

    Google Scholar 

  21. 21.

    Andersen, C.L., Jensen, J.L., and Orntoft, T.F., Normalization of Real-Time Quantitative Reverse Transcription-PCR Data: A Model-Based Variance Estimation Approach to Identify Genes Suited for Normalization, Applied to Bladder and Colon Cancer Data Sets, Cancer Res., 2004, vol. 64, pp. 5245–5250.

    PubMed  Article  CAS  Google Scholar 

  22. 22.

    Murashige, T. and Skoog, F., A Revised Medium for Rapid Growth and Bioassays with Tobacco Tissue Cultures, Physiol. Plant., 1962, vol. 15, pp. 473–497.

    Article  CAS  Google Scholar 

  23. 23.

    Murray, M.G. and Thompson, W.F., Rapid Isolation of High Molecular Weight DNA, Nucleic Acids Res., 1980, vol. 8, pp. 4321–4325.

    PubMed  Article  CAS  Google Scholar 

  24. 24.

    Inoue, H., Nojima, H., and Okayama, H., High Efficiency Transformation of Escherichia coli with Plasmids, Gene, 1990, vol. 96, pp. 23–28.

    PubMed  Article  CAS  Google Scholar 

  25. 25.

    Pfaffl, M.W., A New Mathematical Model for Relative Quantification in Real-Time RT-PCR, Nucleic Acids Res., 2001, vol. 29, p. e45.

    PubMed  Article  CAS  Google Scholar 

  26. 26.

    Kondo, Y., Hirakawa, Y., Kieber, J.J., and Fukuda, H., CLE Peptides Can Negatively Regulate Protoxylem Vessel Formation via Cytokinin Signaling, Plant Cell Physiol., 2011, vol. 52, pp. 37–48.

    PubMed  Article  CAS  Google Scholar 

Download references

Author information



Corresponding author

Correspondence to I. E. Dodueva.

Additional information

Original Russian Text © I.E. Dodueva, A.S. Kiryushkin, E.V. Yurlova, M.A. Osipova, I.S. Buzovkina, L.A. Lutova, 2013, published in Fiziologiya Rastenii, 2013, Vol. 60, No. 3, pp. 399–407.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Dodueva, I.E., Kiryushkin, A.S., Yurlova, E.V. et al. Effect of cytokinins on expression of radish CLE genes. Russ J Plant Physiol 60, 388–395 (2013). https://doi.org/10.1134/S1021443713020052

Download citation


  • Raphanus sativus
  • CLE peptides
  • cytokinins
  • meristems
  • genetic tumors
  • expression quantification
  • reference genes