DIETARY POLYUNSATURATED FATTY-ACIDS INTERFERE WITH THE INSULIN GLUCOSE ACTIVATION OF L-TYPE PYRUVATE-KINASE GENE-TRANSCRIPTION/

L-type pyruvate kinase (L-PK) is a key glycolytic enzyme regulating th e flux of metabolites through the pyruvate-phosphoenolpyruvate cycle ( 1). The regulation of L-PK is complex involving both hormones and nutr ients. We have found that feeding rats diets containing polyunsaturate d fatty acids (PUFA) significantly inhibits hepatic pyruvate kinase en zyme activity (>60%) and suppresses mRNA(PK) abundance (>70%). Studies with primary hepatocytes indicate that PUFA act directly on hepatocyt es. Specifically, arachidonic (20:4, omega 6) and eicosapentaenoic (20 :5, omega 3) acid suppressed both mRNA(PK) levels and the activity of a transfected PKCAT (-4300/+12) fusion gene by >70%. This is due to an inhibition of the insulin/glucose-mediated transactivation of L-PKCAT . Deletion analysis localized PUFA-regulated cis-acting elements to a region within the L-PK proximal promoter, i.e. between -197 and -96 ba se pairs. This region binds two transcription factors involved in the hormone/nutrient regulation of L-PK gene transcription, i.e. a major l ate transcription factor-like factor and HNF-4. Linker scanning mutati on analysis localized the PUFA-regulated cis-acting elements to the vi cinity of the HNF-4 binding site. Thus, PUFA-regulated factors abrogat e the insulin/glucose activation of L-PK gene transcription by targeti ng the HNF-4 elements. These studies suggest that PUFA may have signif icant effects on hepatic carbohydrate metabolism by inhibiting the L-P K side of the pyruvate-phosphoenolpyruvate cycle.