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Papers In Press, published online ahead of print April 1, 2005
J. Lipid Res., doi:10.1194/jlr.M400457-JLR200

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Journal of Lipid Research, Vol. 46, 759-768, April 2005
Copyright © 2005 by American Society for Biochemistry and Molecular Biology

Neutral glycolipids of the filamentous fungus Neurospora crassa: altered expression in plant defensin-resistant mutants

Chaeho Park^*, Beau Bennion, Isabelle E. J. A. François, Kathelijne K. A. Ferket, Bruno P. A. Cammue, Karin Thevissen and Steven B. Levery^1,

^* Complex Carbohydrate Research Center and Department of Biochemistry and Molecular Biology, University of Georgia, Athens, GA 30602-7229
Department of Chemistry, University of New Hampshire, Durham, NH 03824-3598
Center of Microbial and Plant Genetics, Katholieke Universiteit Leuven, B-3001 Heverlee-Leuven, Belgium

Published, JLR Papers in Press, January 16, 2005. DOI 10.1194/jlr.M400457-JLR200

The online version of this article (available at http://www.jlr.org) contains an additional figure and table.

¹ To whom correspondence should be addressed. e-mail: slevery{at}cisunix.unh.edu

To defend themselves against fungal pathogens, plants produce numerous antifungal proteins and peptides, including defensins, some of which have been proposed to interact with fungal cell surface glycosphingolipid components. Although not known as a phytopathogen, the filamentous fungus Neurospora crassa possesses numerous genes similar to those required for plant pathogenesis identified in fungal pathogens (Galagan, J. E., et al. 2003. Nature 422: 859–868), and it has been used as a model for studying plant-phytopathogen interactions targeting fungal membrane components (Thevissen, K., et al. 2003. Peptides. 24: 1705–1712). For this study, neutral glycolipid components were extracted from wild-type and plant defensin-resistant mutant strains of N. crassa. The structures of purified components were elucidated by NMR spectroscopy and mass spectrometry. Neutral glycosphingolipids of both wild-type and mutant strains were characterized as ß-glucopyranosylceramides, but those of the mutants were found with structurally altered ceramides. Although the wild type expressed a preponderance of N-2'-hydroxy-(E)-³-octadecenoate as the fatty-N-acyl component attached to the long-chain base (4E,8E)-9-methyl-4,8-sphingadienine, the mutant ceramides were found with mainly N-2'-hydroxyhexadecanoate instead. In addition, the mutant strains expressed highly increased levels of a sterol glucoside identified as ergosterol-ß-glucoside.

The potential implications of these findings with respect to defensin resistance in the N. crassa mutants are discussed.

Abbreviations: CID, collision-induced dissociation; ESI, electrospray ionization; Fa, fatty acyl; GlcCer, ß-glucopyranosylceramide; GlcSte, steryl-ß-glucopyranoside; HPTLC, high-performance thin-layer chromatography; TOF, time-of-flight; UGT, UDP-Glc:sterol ß-D-glucosyltransferase

Supplementary key words ergosterol • sphingolipid • sterol • glucoside • ceramide • cerebroside • electrospray ionization • nuclear magnetic resonance spectroscopy • mass spectrometry • collision-induced dissociation • tandem mass spectrometry

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