LIPID MEDIATORS ELF INSULIN-RESISTANCE - CERAMIDE SIGNALING DOWN-REGULATES GLUT4 GENE-TRANSCRIPTION IN 3T3-L1 ADIPOCYTES

We have previously demonstrated that chronic exposure of 3T3-L1 adipoc ytes to tumour necrosis factor-alpha. (TNF) resulted in a marked decre ase(similar to 90%)in cellular GLUT4 (insulin-responsive glucose trans porter) mRNA content as a result of a decreased transcription rate of the GLUT4 gene (similar to 75%) and a reduced half-life of its mRNA (9 to 4.5 h). Investigation of the signalling mechanism responsible for this regulation demonstrated that in the 3T3-L1 adipocytes, sphingomye lin levels decreased to 50% of control levels within 40 min of exposur e to TNF, consistent with activation of a sphingomyelinase. In the sam e manner as with TNF, treatment of the adipocytes with 1-3 mu M C-6-ce ramide, a membrane-permeable analogue of ceramide, decreased GLUT4 mRN A content by similar to 60 %. Subsequent investigations revealed that transcription of the GLUT4 gene was reduced by similar to 65% in respo nse to C-6-ceramide, demonstrating that the decrease in mRNA content i s mediated by a reduction in the transcription of the gene. No effect on GLUT4 mRNA stability was observed after exposure of the adipocytes to C-6-ceramide. These observations are interesting in light of our pr evious data demonstrating that TNF affects both GLUT4 transcription an d mRNA stability in the 3T3-L1 adipocytes. In conclusion, the effect o f ceramide on GLUT4 gene expression is at the level of transcription, suggesting that another pathway controls mRNA stability. These data es tablish that ceramide-initiated signal transduction pathways exist wit hin the adipocyte, and provide a potential mechanism for control of GL UT4 gene expression.