REGULATION OF GLUT4 GENE-EXPRESSION BY ARACHIDONIC-ACID - EVIDENCE FOR MULTIPLE PATHWAYS, ONE OF WHICH REQUIRES OXIDATION TO PROSTAGLANDIN E(2)

We have previously described the ability of arachidonic acid (AA) to r egulate GLUT4 gene expression (Tebbey, P. W., McGowan, K. M., Stephens , J. M., Buttke, T. M., and Pekala, P. H. (1994) J. Biol. Chem. 269, 6 39-644). Chronic exposure (48 h) of fully differentiated 3T3 L1 cells to AA resulted in an similar to 90% suppression of GLUT4 mRNA accumula tion. This decrease was demonstrated to be due to a 50% decrease in GL UT4 gene transcription as well as a destabilization of the GLUT4 messa ge (t(1/2) decreased from 8.0 to 4.6 h). In the current study we have identified, at least in part, the mechanism by which AA exerts its eff ects on GLUT4 expression. Compatible with a cyclooxygenase mediated ev ent, the AA-induced suppression of GLUT4 mRNA was abolished by pretrea ting the cells with the inhibitor, indomethacin. Consistent with this observation, exposure of the cells to 10 mu M PGE(2) mimicked the effe ct of AA, in contrast to products of the lipoxygenase pathway which we re unable to suppress GLUT4 mRNA content. Quantification of the conver sion of AA to PGE(2) demonstrated a 50-fold increase in PGE(2) release d into the media within 7 h of AA addition. Cyclic AMP levels were als o increased 50-fold with AA treatment consistent with PGE(2) activatio n of adenylate cyclase. Various long chain fatty acids, including the nonmetabolizable analog of AA, eicosatetraenoic acid (ETYA), also decr eased GLUT4 mRNA levels. The effect of ETYA, a potent inhibitor of bot h lipo- and cyclooxygenases and a potent activator of peroxisome proli ferator activated receptors (PPARs), suggested the presence of a secon d pathway where nonmetabolized fatty acid functioned to suppress GLUT4 mRNA levels. Further support for a PPAR-mediated mechanism was obtain ed by exposure of the cells to the classic PPAR activator, clofibrate, which resulted in a similar to 75% decrease in GLUT4 mRNA content. Nu clear extracts prepared from the adipocytes contained a protein comple x that bound to the PPAR responsive element (PPRE) found in the promot er of the fatty acyl-CoA oxidase gene. When the adipocytes were treate d with either AA or ETYA, binding to the PPRE was disrupted, consisten t with an ability of these fatty acids to control gene expression by a ltering the occupation of a PPRE. However, a perfect PPRE has yet to b e identified in the GLUT4 promoter, but this does not rule the possibi lity of a PPAR playing an indirect role in the AA-induced GLUT4 mRNA s uppression.