Fatty acids induce L-CPT I gene expression through a PPARalpha-independent mechanism in rat hepatoma cells.
Liver carnitine palmitoyl transferase (L-CPT) I is a key regulatory enzyme of long-chain fatty acid (LCFA) oxidation that ensures the first step of LCFA import into the mitochondrial matrix. In rat hepatocytes, we showed previously that L-CPT I gene expression was induced by LCFAs as well as by fibrates. The aim of this study was to determine whether LCFA-induced L-CPT I gene expression was mediated by PPARalpha. For this purpose, we constructed a PPARalpha-dominant negative receptor to inhibit endogenous PPARalpha signaling. Highly conserved hydrophobic and charged residues (Leu459 and Glu462) in helix 12 of the ligand-binding domain were mutated to alanine. These mutations led to a total loss of transcriptional activity due to impaired coactivator recruitment. Furthermore, competition studies confirmed that the mutated PPARalpha receptor abolished the wild-type PPARalpha receptor action and thus acted as a powerful dominant negative receptor. When overexpressed in rat hepatoma cells (H4IIE) using a recombinant adenovirus, the mutated PPARalpha receptor antagonized the clofibrate-induced L-CPT I gene expression, whereas it did not affect LCFA-induced L-CPT I. These results provide the first direct demonstration that LCFAs regulate L-CPT I transcription through a PPARalpha-independent pathway, at least in hepatoma cells.
Le May
Cédric
C
Institut Cochin, INSERM U567, CNRS UMR 8104, Université PARIS V, Département d'Endocrinologie, Paris.
Caüzac
Michèle
M
Diradourian
Claire
C
Perdereau
Dominique
D
Girard
Jean
J
Burnol
Anne-Françoise
AF
Pégorier
Jean-Paul
JP
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D019556:COS Cells; D006528:Carcinoma, Hepatocellular; D002334:Carnitine O-Palmitoyltransferase / Q000235:genetics* / Q000378:metabolism*; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D002994:Clofibrate / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D000960:Hypolipidemic Agents / Q000494:pharmacology; D019787:Linoleic Acid / Q000494:pharmacology*; D008113:Liver Neoplasms; D047493:PPAR alpha / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation / Q000187:drug effects
D000960:Hypolipidemic Agents; D047493:PPAR alpha; D011994:Recombinant Proteins; D019787:Linoleic Acid; D002334:Carnitine O-Palmitoyltransferase; D002994:Clofibrate