Post-transcriptional regulation of rat carnitine octanoyltransferase

Carnitine octanoyltransferase (COT) produces three different transcripts in rat through cis- and trans-splicing reactions, which can lead to the synth esis of two proteins. The occurrence of the three COT transcripts in rat ha s been found in all tissues examined and does not depend on sex, fat feedin g, peroxisome proliferators or hyperinsulinaemia. Rat COT exon 2 contains a putative exonic splicing enhancer (ESE) sequence. Mutation of this ESE (GA AGAAG) to AAAAAAA decreased trans-splicing in vitro, from which it is deduc ed that this ESE sequence is partly responsible for the formation of the th ree transcripts. The protein encoded by cis-spliced mRNA of rat COT is inhi bited by malonyl-CoA and etomoxir. cDNA species encoding full-length wild-t ype COT and one double mutant COT were expressed in Saccharomyces cerevisia e. The recombinant enzymes showed full activity towards both substrates, ca rnitine and decanoyl-CoA. The activity of the doubly mutated H131A/H340A en zyme was similar to that of the rat peroxisomal enzyme but was completely i nsensitive to malonyl-CoA and etomoxir. These results indicate that the his tidine residues His-131 and His-340 are the sites responsible for the inter action of these two inhibitors, which inhibit COT by interacting with the s ame sites.