Sphingolipid catabolites are important signaling molecules present in most eukaryotic cells. The results of sphingolipid signaling are diverse and cell type specific. Some biological effects of sphingolipid signaling include regulation of cell cycle progression, differentiation, proliferation, survival, apoptosis, migration, angiogenesis, and drug resistance. The genetic components of sphingolipid metabolism have recently been identified through the employment of yeast genetics, opening the door to the use of genetic approaches toward elucidating sphingolipidmediated biology. Drosophila melanogaster provides a powerful metazoan model for the study of signal transduction events and their roles in development. The Principal Investigator has identified the Drosophila melanogaster sphingosine phosphate lyase gene, Sply, which encodes an enzyme required for normal degradation of the sphingolipid, sphingosine-l-phosphate. We find that null mutations in this gene lead to a flightless phenotype, caused by defects of indirect flight muscles. Drosophila lace mutants, which lack serine palmitoyl transferase activity, fail to synthesize sphingolipids through the de novo pathway and demonstrate developmental defects distinct from those observed in the Sply mutant. Two cDNAs encoding putative sphingosine kinases, which catalyze the synthesis of sphingosine-l-phosphate from sphingosine, both functionally complement a yeast sphingosine kinase mutant, and for one of these genes a corresponding mutant line exists. These three mutants provide a novel schema by which to elucidate Drosophila sphingolipid metabolism and clarify the role sphingolipids play in specific developmental programs. Therefore, the long-term research goals of this proposal are to further elucidate sphingolipid signaling and the roles of sphingolipids in biology through the use of Drosophila models. Specifically, we propose: 1) to establish the structure of the major sphingolipid signaling molecules of Drosophila melanogaster using high performance liquid chromatography, mass spectrometry, nuclear magnetic resonance and radiolabeling approaches 2) to identify and characterize genes encoding sphingosine phosphate lyase, sphingosine kinase and serine palmitoyl transferase enzymes in Drosophila melanogaster using nucleotide sequence homology based approaches and 3) to determine the role sphingolipids play in Drosophila development, by dissecting phenotypes and analyzing lipid profiles throughout development of Drosophila sphingosine kinase, sphingosine phosphate lyase and serine palmitoyl transferase mutants and the progeny of strategic genetic crosses between new and existing models of sphingolipid metabolism.
Showing the most recent 10 out of 30 publications
