Skip to main content
SpringerLink
Log in

Photoaffinity labelling with the cytokinin agonist azido-CPPU of a 34 kDa peptide of the intracellular pathogenesis-related protein family in the moss Physcomitrella patens

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

As in higher plants, the development of the moss Physcomitrella patens is regulated by environmental signals and phytohormones. At the protonema level transition from chloronema to caulonema cells is under auxin control. The formation on second sub-apical caulonema cells of buds that will give rise to the leafy gametophore requires cytokinins. Using [3H]azidoCPPU (1-(2-azido-6-chloropyrid-4-yl)-3-(4-[3H])phenylurea), a photoactivatable cytokinin agonist, we have specifically photolabelled a soluble 34 kDa protein of P. patens. Urea derivatives were very efficient competitors of photolabelling while purine-type cytokinins were poor competitors. The protein UBP34 was purified by affinity chromatography and the sequences of six internal peptides obtained. A cDNA encoding UBP34 was cloned by screening a P. patens protonema cDNA library with a probe amplified by PCR using degenerate primers designed from the peptide sequences. The UBP34 amino acid sequence shows an average sequence identity of 42% with both intracellular PR proteins and the BetV1-related family of plant allergens. Recombinant UBP34 expressed in Escherichia coli was confirmed to bind azidoCPPU.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

REFERENCES

  • Ashton, N.W. and Cove, D.J. 1977. The isolation and preliminary characterisation of auxotrophic and analogue resistant mutants of the moss Physcomitrella patens. Mol. Gen. Genet. 154: 87–95.

    Google Scholar 

  • Ashton, N.W., Grimsley, N.H. and Cove, D.J. 1979a. Analysis of gametophytic development in the moss, Physcomitrella patens, using auxin and cytokinin-resistant mutants. Planta 144: 427–435.

    Google Scholar 

  • Ashton, N.W., Cove, D.J. and Featherstone, D.R. 1979b. The isolation and physiological analysis of mutants of the moss Physcomitrella patens which over-produce gametophores. Planta 144: 437–442.

    Google Scholar 

  • Bleecker, A.B. 1999. Ethylene perception and signaling: an evolutionary perspective. Trends Plant Sci. 4: 269–274.

    Article  PubMed  Google Scholar 

  • Brault, M., Maldiney, R. and Miginiac, E. 1997. Cytokinin-binding proteins. Physiol. Plant. 100: 520–527.

    Google Scholar 

  • Carpin, S., Laffer, S., Schoentgen, F., Valenta, R., Chénieux, J.C., Rideau, M. and Hamdi, S. 1998. Molecular characterization of a cytokinin-inducible periwinkle protein showing sequence homology with pathogenesis-related proteins and the Bet v 1 allergen family. Plant Mol. Biol. 36: 791–798.

    Google Scholar 

  • Corpet, F. 1988. Multiple sequence alignment with hierarchical clustering. Nucl. Acids Res. 16: 10881–10890.

    PubMed  Google Scholar 

  • Corse, J., Pacovsky, R.S., Brandon, D.L. and McKeon, T.A. 1992. Identification of cytokinin receptors by means of structure-activity response. In: M. Kamineck, D.W.S. Mok and E. Zazimalova (Eds.) Physiology and Biochemistry of Cytokinins in Plants, SPB Academic Publishing, The Hague, Netherlands, pp. 211–214.

    Google Scholar 

  • Cove, D.J. 1992. Regulation of development in the moss, Physcomitrella patens. In: S. Brody, D.J. Cove, S. Ottolenghi and V.E.A. Russo (Eds.) Developmental Biology: A Molecular Genetic Approach, Springer-Verlag, Heidelberg, Germany, pp. 179–188.

    Google Scholar 

  • Christianson, M.L. and Hornbuckle, J.S. 1999. Phenylurea cytokinins assayed for induction of shoot buds in the moss Funaria hygrometrica. Am. J. Bot. 86: 1645–1648.

    PubMed  Google Scholar 

  • Deikman, J. and Ulrich, M. 1995. A novel cytokinin-resistant mutant of Arabidopsis with abbreviated shoot development. Planta 195: 440–449.

    Article  PubMed  Google Scholar 

  • Dias, M., Mornet, R. and Laloue, M. 1995. Synthesis, azidotetrazole equilibrium studies and biological activity of 1-(2-azido-6-chloro-pyrid-4-yl)-3-phenylurea, a photoaffinity labeling reagent for cytokinin-binding proteins. Bioorg. Med. Chem. 3: 361–366.

    PubMed  Google Scholar 

  • Faure, J.D. and Howell, S.H. 1999. Cytokinin perception and signal transduction. In: P.J. Hooykaas, M.A. Hall and K.R. Libbenga (Eds.) Biochemistry and Molecular Biology of Plant Hormones, Elsevier Science, Amsterdam, pp. 461–474.

    Google Scholar 

  • Fujimoto, Y., Nagata, R., Fukasawa, H., Yano, K., Azuma, M., Iida, A., Sugimoto, S., Shudo, K. and Hashimoto, Y. 1998. Purification and cDNA cloning of cytokinin-specific binding protein from mung bean (Vigna radiata). Eur. J. Biochem. 258: 794–802.

    PubMed  Google Scholar 

  • Girke, T., Schmidt, H., Zähninger, U., Reski, R. and Heinz, E. 1998. Identification of a novel)-acyl-group desaturase by targeted gene disruption in Physcomitrella patens. Plant J. 15: 39–48.

    PubMed  Google Scholar 

  • Girod, P.A., Fu, H., Zrßd, J.-P. and Vierstra, R.D. 1999. Multiubiquitin chain binding subunit MCB21 (RPN10) of the 26S proteasome is essential for developmental progression in Physcomitrella patens. Plant Cell 11: 1457–1471.

    PubMed  Google Scholar 

  • Gonneau, M., Mornet, R. and Laloue, M. 1998. A Nicotiana plumbaginifolia protein labeled with an azido cytokinin agonist is a glutathione S-transferase. Physiol. Plant. 103: 114–124.

    Google Scholar 

  • Henrie, R.N., Green, C.M., Yeager, W.H. and Ball, T.F. 1998. Activity optimization of pyridinyl N-oxide urea cytokinin mimics. J. Agric. Food Chem. 36: 626–633.

    Google Scholar 

  • Hofmann, A.H., Codon, A.C., Ivascu, C., Russo, V.E.A., Knight, C., Cove, D., Schaefer, D.G., Chakhparonian, M. and Zrßd, J.P. 1999. A specific member of the Cab multigene family can be efficiently targeted and disrupted in the moss Physcomitrella patens. Mol. Gen. Genet. 261: 92–99.

    Google Scholar 

  • Hoffmann-Sommergruber, K., Vanek-Krebitz, M., Radauer, C., Wen J., Ferreira, F., Scheiner, O. and Breitender, H. 1997. Genomic characterization of members of the Bet v 1 family: genes coding for allergens and pathogenesis-related proteins share intron positions. Gene 197: 91–100.

    PubMed  Google Scholar 

  • Kakimoto, T. 1998. Cytokinin signaling. Curr. Opin. Plant Biol. 1: 399–403

    PubMed  Google Scholar 

  • Kakimoto, T. 1996. CKI1, a histidine kinase homolog implicated in cytokinin signal transduction. Science 274: 982–985.

    Article  PubMed  Google Scholar 

  • Kobayashi, K., Fukuda, M., Igarashi, D. and Sunaoshi, M. 2000. Cytokinin-binding proteins from tobacco callus share homology with osmotin-like protein and an endochitinase. Plant Cell Physiol. 41: 148–157.

    PubMed  Google Scholar 

  • Laemmli, U.K. Cleavage of structural proteins during the assembly of the head of the bacteriophage T4. Nature 227: 680–685.

  • Machuka, J., Bashiardes, S., Ruben, E., Spooner, K., Cuming, A., Knight, C. and Cove, D. 1999. Sequence analysis of expressed sequence tags from an ABA-treated cDNA library identifies stress response genes in the moss Physcomitrella patens.Plant Cell Physiol. 40: 378–387.

    PubMed  Google Scholar 

  • Nagata, R., Kawachi, E., Hasimoto, Y. and Shudo, K. 1993. Cytokinin-specific binding protein in etiolated mung bean seedlings. Biochem. Biophys. Res. Commun. 191: 543–549.

    PubMed  Google Scholar 

  • Nogué, F., Jullien, M., Mornet, R. and Laloue, M. 1995. The response of a cytokinin resistant mutant is highly specific and permits a new cytokinin bioassay. Plant Growth Regul. 17: 87–94.

    Google Scholar 

  • Nogué, F., Mornet, R. and Laloue, M. 1996. Specific photoaffinity labeling of a thylakoid membrane protein with an azido-cytokinin agonist. Plant Growth Regul. 18: 51–58.

    Google Scholar 

  • Osmark, P., Boyle, B. and Brisson, N. 1998. Sequential and structural homology between intracellular pathogenesis proteins and a group of latex proteins. Plant Mol. Biol. 38: 1243–1246.

    Google Scholar 

  • Pozueta-Romero, J., Klein, M., Houlné, G., Schantz, M-L., Meyer, B. and Schantz R. 1995. Characterization of a family of genes encoding a fruit-specific wound-stimulated protein of bell pepper (Capsicum annuum): identification of a new family of transposable elements. Plant Mol. Biol. 28: 1011–1025.

    Google Scholar 

  • Reski, R. 1998. Development, genetics and molecular biology of mosses. Bot. Acta 111: 1–15.

    Google Scholar 

  • Saraste, M., Sibbad, P.R. and Wittinghofer, A. 1990. The P-loop: a common motif in ATP-and GTP-binding proteins. Trends Biochem. Sci. 19: 430–434.

    Google Scholar 

  • Schaefer, D. and Zrßd, J.-P. 1997. Efficient gene targeting in the moss Physcomitrella patens. Plant J. 11: 1195–1206.

    PubMed  Google Scholar 

  • Schmülling, T., Schäfer, S. and Romanov G. 1997. Cytokinins as regulators of gene expression. Physiol. Plant. 100: 505–519.

    Google Scholar 

  • Skinner, M.K. and Griswold, M.D. 1983. Fluorographic detection of radioactivity in polyacryamide gels with 2,5-diphenyloxazole in acetic acid and its comparison with existing procedures. Biochem. J. 209: 281–284.

    PubMed  Google Scholar 

  • Skoog, F., Schmitz, R.Y., Bock, R.M. and Hecht, S.M. 1973. Cytokinin antagonists: synthesis and physiological effects of 7-substituted 3-methyl pyrazolo(4,3-d)pyrimidines. Phytochemistry 12: 25–37.

    Google Scholar 

  • Strepp, R., Scholz, S., Kruse, S., Speth, V. and Reski, R. 1998. Plant nuclear gene knockout reveals a role in plastid division for the homolog of the bacterial cell division protein FtsZ, an ancestral tubulin. Proc. Natl. Acad. Sci. USA 95: 4368–4373.

    Google Scholar 

  • Takahashi, S., Shudo, K., Okamoto, T., Yamada, K. and Isogai, Y. 1978. Cytokinin activity of N-phenyl-N-(pyridyl)urea derivatives. Phytochemistry 17: 1201–1207.

    Google Scholar 

  • Vanek-Krebitz, M., Hoffmann-Sommergruber, K., Lainer da Camara Machado, H., Susani, M., Ebner, C., Kraft, D. and Scheiner, O. 1995. Cloning and sequencing of Mal d1, the major allergen from apple (Malus domestica) and its immunological relation ship to Bet V1, the major birch pollen allergen. Biochem. Biophys. Res. Commun. 2114: 538–551.

    Google Scholar 

  • Wang, T.L., Cove, D.J., Beutelmann, P. and Hartmann, E. 1980. Isopentenyladenine from mutants of the moss, Physcomitrella patens. Phytochemistry 19: 1103–1105.

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Laboratoire de Biologie Cellulaire, Institut National de la Recherche Agronomique, INRA-Versailles-Grignon, 78026, Versailles-Cedex, France

    Martine Gonneau, Sylvère Pagant, Florent Brun & Michel Laloue

Authors
  1. Martine Gonneau
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sylvère Pagant
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Florent Brun
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Michel Laloue
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Gonneau, M., Pagant, S., Brun, F. et al. Photoaffinity labelling with the cytokinin agonist azido-CPPU of a 34 kDa peptide of the intracellular pathogenesis-related protein family in the moss Physcomitrella patens. Plant Mol Biol 46, 539–548 (2001). https://doi.org/10.1023/A:1010693213437

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1010693213437

Search

Navigation