MCD (malonyl-CoA decarboxylase), which catalyses decarboxylation of malonyl-CoA, is known to play an important role in the regulation of malonyl-CoA concentration. Recently, it has been observed that the expression of MCD is significantly decreased in the hearts of the PPARα (peroxisome-proliferator-activated receptor α) (−/−) mice, where the rate of fatty-acid oxidation is decreased by the increased malonyl-CoA level [Campbell, Kozak, Wagner, Altarejos, Dyck, Belke, Severson, Kelly and Lopaschuk (2002) J. Biol. Chem. 277, 4098–4103]. This suggests that MCD may be transcriptionally regulated by PPARα. To investigate whether PPARα is truly responsible for transcriptional regulation of the rat MCD gene, transient reporter assay was performed in CV-1 cells. The promoter activity was increased by 17-fold in CV-1 cells co-transfected with PPARα/retinoid X receptor α expression plasmid. In sequence analysis of the promoter region, three putative PPREs (PPAR response elements) were identified, and promoter deletion analysis showed that PPRE2 and PPRE3 were functional. Electrophoretic mobility-shift assays revealed that PPARα/retinoid X receptor α heterodimer indeed bound to the two PPREs, and the binding specificity of PPARα on PPRE was also confirmed by experiments with mutated oligonucleotides. These results indicate that the elements behaved as a responsive site to PPARα activation. MCD mRNA levels in WY14643-treated rat hepatoma cells as well as in the liver of fenofibrate-fed Otsuka Long-Evans Tokushima fatty rats were also found to be increased, suggesting that PPARα can activate the rat hepatic MCD transcription by binding to the PPREs in the promoter. We propose that MCD performs an important role in understanding the regulatory mechanism between activated PPARα and fatty-acid oxidation by altering the malonyl-CoA concentration.
Abbreviations used: ACC, acetyl-CoA carboxylase; AMPK, AMP-activated protein kinase; 9-cis-RA, 9-cis-retinoic acid; CPT-1, carnitine palmitoyltransferase-1; EMSA, electrophoretic mobility-shift assay; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; LCFA-CoA, long-chain fatty acyl-CoA; MCD, malonyl-CoA decarboxylase; NEFA, non-esterified fatty acid; OLETF, Otsuka Long-Evans Tokushima fatty; PPARα, peroxisome-proliferator-activated receptor α; PPRE, PPAR response element; RT, reverse transcriptase; RXRα, retinoid X receptor α; TG, triacylglycerol.