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Reconstitution of cyanogenesis in barley (Hordeum vulgare L.) and its implications for resistance against the barley powdery mildew fungus

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Abstract

Barley (Hordeum vulgare L.) produces a leucine-derived cyanogenic β-d-glucoside, epiheterodendrin that accumulates specifically in leaf epidermis. Barley leaves are not cyanogenic, i.e. they do not possess the ability to release hydrogen cyanide, because they lack a cyanide releasing β-d-glucosidase. Cyanogenesis was reconstituted in barley leaf epidermal cells through single cell expression of a cDNA encoding dhurrinase-2, a cyanogenic β-d-glucosidase from sorghum. This resulted in a 35–60% reduction in colonization rate by an obligate parasite Blumeria graminis f. sp. hordei, the causal agent of barley powdery mildew. A database search for barley homologues of dhurrinase-2 identified a (1,4)-β-d-glucan exohydrolase isozyme βII that is located in the starchy endosperm of barley grain. The purified barley (1,4)-β-d-glucan exohydrolase isozyme βII was found to hydrolyze the cyanogenic β-d-glucosides, epiheterodendrin and dhurrin. Molecular modelling of its active site based on the crystal structure of linamarase from white clover, demonstrated that the disposition of the catalytic active amino acid residues was structurally conserved. Epiheterodendrin stimulated appressoria and appressorial hook formation of B. graminis in vitro, suggesting that loss of cyanogenesis in barley leaves has enabled the fungus to utilize the presence of epiheterodendrin to facilitate host recognition and to establish infection.

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Abbreviations

Glc:

Glucose

GFP:

Green fluorescent

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Acknowledgements

We thank Mogens Houmøller for generously providing the B. graminis f. sp. Hordei isolate. Professor Asim Esen, Virginia Polytechnic Institute and State University Blacksburg, VA, USA is thanked for helpful discussions. This work was supported by grants from the Danish National Research Foundation to Center for Molecular Plant Physiology (PlaCe), from the Swedish Agricultural Research Council to KF, by a EU Marie Curie training grant to SE, and from the Australian Research Council and the Grains Research and Development Corporation to GBF.

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  1. Kirsten A. Nielsen

    Present address: Epilepsy Hospital, 4293, Dianalund, Denmark

  2. Karin Forslund

    Present address: Department of Physiological Botany, EBC, Uppsala University, Villavagen 6, 752 36, Uppsala, Sweden

Authors and Affiliations

  1. Plant Biochemistry Laboratory, Department of Plant Biology, Royal Veterinary and Agricultural University, 1871, Frederiksberg C, Copenhagen, Denmark

    Kirsten A. Nielsen, Janni Nyvang Nielsen, Karin Forslund, Stefan Ebert & Birger Lindberg Møller

  2. Center for Molecular Plant Physiology, Royal Veterinary and Agricultural University, 1871, Frederiksberg C, Copenhagen, Denmark

    Kirsten A. Nielsen & Birger Lindberg Møller

  3. Department of Natural Sciences, Royal Veterinary and Agricultural University, 1871, Frederiksberg C, Copenhagen, Denmark

    Carl E. Olsen

  4. School of Agriculture and Wine, and the Australian Centre for Plant Functional Genomics, University of Adelaide, Waite Campus, 5064, Glen Osmond, SA, Australia

    Maria Hrmova & Geoffrey B. Fincher

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  1. Kirsten A. Nielsen
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Correspondence to Birger Lindberg Møller.

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Nielsen, K.A., Hrmova, M., Nielsen, J.N. et al. Reconstitution of cyanogenesis in barley (Hordeum vulgare L.) and its implications for resistance against the barley powdery mildew fungus. Planta 223, 1010–1023 (2006). https://doi.org/10.1007/s00425-005-0158-z

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