TRANSCRIPTIONAL CONTROL OF A NUCLEAR GENE ENCODING A MITOCHONDRIAL FATTY-ACID OXIDATION ENZYME IN TRANSGENIC MICE - ROLE FOR NUCLEAR RECEPTORS IN CARDIAC AND BROWN ADIPOSE EXPRESSION

Expression of the gene encoding medium-chain acyl coenzyme A dehydroge nase (MCAD), a nuclearly encoded mitochondrial fatty acid beta-oxidati on enzyme, is regulated in parallel with fatty acid oxidation rates am ong tissues and during development. We have shown previously that the human MCAD gene promoter contains a pleiotropic element (nuclear recep tor response element [NRRE-1]) that confers transcriptional activation or repression by members of the nuclear receptor superfamily. Mice tr ansgenic for human MCAD gene promoter fragments fused to a chloramphen icol acetyltransferase gene reporter were produced and characterized t o evaluate the role of NRRE-1 and other promoter elements in the trans criptional control of the MCAD gene in vivo. Expression of the full-le ngth MCAD promoter-chloramphenicol acetyltransferase transgene (MCADCA T371) paralleled the known tissue-specific differences in mitochondria l P-oxidation rates and MCAD expression. MCADCAT.371 transcripts were abundant in heart tissue and brown adipose tissue, tissues with high-l evel MCAD expression. During perinatal cardiac developmental stages, e xpression of the MCADCAT.371 transgene paralleled mouse MCAD mRNA leve ls. In contrast, expression of a mutant MCADCAT transgene, which lacke d NRRE-1 (MCADCAT Delta NRRE-1), was not enriched in heart or brown ad ipose tissue and did not exhibit appropriate postnatal induction in th e developing heart. Transient-transfection studies with MCAD promoter- luciferase constructs containing normal or mutant NRRE-1 sequences dem onstrated that the nuclear receptor binding sequences within NRRE-1 ar e necessary for high-level transcriptional activity in primary rat car diocytes. Electrophoretic mobility shift assays demonstrated that NRRE -1 was bound by several cardiac and brown adipose nuclear proteins and that these interactions required the NRRE-1 receptor binding hexamer sequences. Antibody supershift studies identified the orphan nuclear r eceptor COUP-TF as one of the endogenous cardiac proteins which bound NRRE-1. These results dictate an important role for nuclear receptors in the transcriptional control of a nuclear gene encoding a mitochondr ial fatty acid oxidation enzyme and identify a gene regulatory pathway involved in cardiac energy metabolism.