Reconstitution of cyanogenesis in barley (Hordeum vulgare L.) and its implications for resistance against the barley powdery mildew fungus |
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Authors: | Kirsten A. Nielsen Maria Hrmova Janni Nyvang Nielsen Karin Forslund Stefan Ebert Carl E. Olsen Geoffrey B. Fincher Birger Lindberg Møller |
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Affiliation: | (1) Plant Biochemistry Laboratory, Department of Plant Biology, Royal Veterinary and Agricultural University, 1871 Frederiksberg C, Copenhagen, Denmark;(2) Center for Molecular Plant Physiology, Royal Veterinary and Agricultural University, 1871 Frederiksberg C, Copenhagen, Denmark;(3) Department of Natural Sciences, Royal Veterinary and Agricultural University, 1871 Frederiksberg C, Copenhagen, Denmark;(4) School of Agriculture and Wine, and the Australian Centre for Plant Functional Genomics, University of Adelaide, Waite Campus, 5064 Glen Osmond, SA, Australia;(5) Present address: Epilepsy Hospital, 4293 Dianalund, Denmark;(6) Present address: Department of Physiological Botany, EBC, Uppsala University, Villavagen 6, 752 36 Uppsala, Sweden |
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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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Keywords: | Cyanogenesis |
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