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J. Biol. Chem., Vol. 279, Issue 6, 3900-3905, February 6, 2004
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**

From the
Centre of Microbial and Plant Genetics, Kasteelpark Arenberg 20, Heverlee B-3001, Belgium, the ¶Institut für Allgemeine Botanik, University of Hamburg, Ohnhorststrasse 18, Hamburg 22609, Germany, and the ||Research Center Borstel, Center for Medicine and Biosciences, Parkallee 22, Borstel 23845, Germany
Growth of the yeast species Candida albicans and Pichia pastoris is inhibited by RsAFP2, a plant defensin isolated from radish seed (Raphanus sativus), at micromolar concentrations. In contrast, gcs-deletion mutants of both yeast species are resistant toward RsAFP2. GCS genes encode UDP-glucose:ceramide glucosyltransferases, which catalyze the final step in the biosynthesis of the membrane lipid glucosylceramide. In an enzyme-linked immunosorbent assay-based binding assay, RsAFP2 was found to interact with glucosylceramides isolated from P. pastoris but not with soybean nor human glucosylceramides. Furthermore, the P. pastoris parental strain is sensitive toward RsAFP2-induced membrane permeabilization, whereas the corresponding gcs-deletion mutant is highly resistant to RsAFP2-mediated membrane permeabilization. A model for the mode of action of RsAFP2 is presented in which all of these findings are linked. Similarly to RsAFP2, heliomicin, a defensin-like peptide from the insect Heliothis virescens, is active on C. albicans and P. pastoris parental strains but displays no activity on the gcs-deletion mutants of both yeast species. Furthermore, heliomicin interacts with glucosylceramides isolated from P. pastoris and soybean but not with human glucosylceramides. These data indicate that structurally homologous anti-fungal peptides present in species from different eukaryotic kingdoms interact with the same target in the fungal plasma membrane, namely glucosylceramides, and as such support the hypothesis that defensins from plants and insects have evolved from a single precursor.
Received for publication, October 10, 2003
* This work was supported in part by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 470, and by the BMBF Project NAPUS 2000 Part 0312252F. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
** Postdoctoral fellow of the Fonds voor Wetenschappelijk Onderzoek-Vlaanderen. Present address: Laboratory of Phytopathology, Wageningen University, The Netherlands.
To whom correspondence should be addressed. Tel.: 32-16-32-9688; Fax: 32-16-32-1966; E-mail: karin.thevissen{at}agr.kuleuven.ac.be.
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