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
Dl. Disch et al., 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, Molecular and cellular biology, 16(8), 1996, pp. 4043-4051
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.