Abstract
Somatic embryogenesis of carrot (Daucus carota L.) is inhibited by the glycosylation inhibitor tunicamycin. This inhibition is reversible by the addition of correctly glycosylated glycoproteins which have been secreted into the culture medium. To identify the proteins responsible for complementation, glycoproteins present in the medium of embryo cultures were purified and tested for their activity in the tunicamycin inhibition/ complementation assay. A 38-kDa glycoprotein was purified that could restore embryogenesis to more than 50% of that in untreated controls. This 38-kDa glycoprotein was identified as a heme-containing peroxidase on the basis of its A405/A280 ratio (Reinheit Zahl or RZ) and enzyme activity. The 38-kDa peroxidase consisted of four different cationic isoenzymes of which only one or possibly two appeared active in the complementation assay. The cationic peroxidase isoenzymes from the carrot medium could be effectively replaced by cationic horseradish peroxidases which depended on their catalytic properties for their ability to restore tunicamycin-inhibited somatic embryogenesis.
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Abbreviations
- IEF:
-
isoelectric focussing
- HRP c:
-
cationic horse-radish peroxidase
- pI:
-
isoelectric point
- SDS-PAGE:
-
sodium dodecyl sulfate-polyacrylamide gel electrophoresis
- TFMS:
-
trifluoromethanesulfonic acid
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This work was supported by grant of the Stichting Innovatiefonds Plantenveredeling (J.C.) and partially by grant of the European Commision (S.C. de V.). We thank Dr. T. Hendriks, Department of Molecular Biology, Agricultural University Wageningen, Wageningen, The Netherlands for helpful discussions and Dr. A.J. Fleming, Botanisches Institut, Universität Bern, Switzerland, Dr. L. Faye, Physiologie Cellulaire, Université de Rouen, Mont-Saint-Aignan, France and Dr. S.C. Fry, Department of Botany, University of Edinburgh, UK for many stimulating discussions and for critical reading of the manuscript.
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Cordewener, J., Booij, H., van der Zandt, H. et al. Tunicamycin-inhibited carrot somatic embryogenesis can be restored by secreted cationic peroxidase isoenzymes. Planta 184, 478–486 (1991). https://doi.org/10.1007/BF00197895
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DOI: https://doi.org/10.1007/BF00197895