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Originally published In Press as doi:10.1194/jlr.M700414-JLR200 on December 21, 2007

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Journal of Lipid Research, Vol. 49, 597-606, March 2008
Copyright © 2008 by American Society for Biochemistry and Molecular Biology

Identification and characterization by electrospray mass spectrometry of endogenous Drosophila sphingadienes

Henrik Fyrst*, Xinyi Zhang{dagger}, Deron R. Herr§, Hoe Sup Byun**, Robert Bittman**, Van H. Phan§, Greg L. Harris§ and Julie D. Saba1,*

* Children's Hospital Oakland Research Institute, Oakland, CA 94609
{dagger} Bruker Daltonics, Inc., Beijing 100081, China
§ Department of Biology and Molecular Biology Institute, San Diego State University, San Diego, CA 92007
** Department of Chemistry and Biochemistry, Queens College of City University of New York, Flushing, NY 11367-1597

Published, JLR Papers in Press, December 21, 2007.

1 To whom correspondence should be addressed. e-mail jsaba{at}chori.org

Sphingolipids comprise a complex group of lipids concentrated in membrane rafts and whose metabolites function as signaling molecules. Sphingolipids are conserved in Drosophila, in which their tight regulation is required for proper development and tissue integrity. In this study, we identified a new family of Drosophila sphingolipids containing two double bonds in the long chain base (LCB). The lipids were found at low levels in wild-type flies and accumulated markedly in Drosophila Sply mutants, which do not express sphingosine-1-phosphate lyase and are defective in sphingolipid catabolism. To determine the identity of the unknown lipids, purified whole fly lipid extracts were separated on a C18-HPLC column and analyzed using electrospray mass spectrometry. The lipids contain a LCB of either 14 or 16 carbons with conjugated double bonds at C4,6. The {Delta}4,6-sphingadienes were found as free LCBs, as phosphorylated LCBs, and as the sphingoid base in ceramides. The temporal and spatial accumulation of {Delta}4,6-sphingadienes in Sply mutants suggests that these lipids may contribute to the muscle degeneration observed in these flies.

Supplementary key words high-performance liquid chromatography • sphingolipids • HPLC • sphingadiene • sphingosine-1-phosphate lyase • muscle • Sply

Abbreviations: dsRNA, double-stranded RNA; LCB, long chain base; LCBP, long chain base phosphate; MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium; RNAi, RNA interference; SPL, sphingosine-1-phosphate lyase






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