GENOMIC STRUCTURE OF MURINE METHYLMALONYL-COA MUTASE - EVIDENCE FOR GENETIC AND EPIGENETIC MECHANISMS DETERMINING ENZYME-ACTIVITY

Methylmalonyl-COA mutase (McM) is a nuclear-encoded mitochondrial matr ix enzyme. We have reported characterization of murine MCM and cloning of a murine MCM cDNA and now describe the murine Mut locus, its promo ter and evidence for tissue-specific variation in MCM mRNA, enzyme and holoenzyme levels. The Mut locus spans 30 kb and contains 13 exons co nstituting a unique transcription unit. A B1 repeat element was found in the 3' untranslated region (exon 13). The transcription initiation site was identified and upstream sequences were shown to direct expres sion of a reporter gene in cultured cells. The promoter contains seque nce motifs characteristic of: (1) TATA-less housekeeping promoters; (2 ) enhancer elements purportedly involved in co-ordinating expression o f nuclear-encoded mitochondrial proteins; and (3) regulatory elements including CCAAT boxes, cyclic AMP-response elements and Potential AP-2 -binding sites. Northern blots demonstrate a greater than 10-fold vari ation in steady-state mRNA levels, which correlate with tissue levels of enzyme activity. However, the ratio of holoenzyme to total enzyme v aries among different tissues, and there is no correlation between ste ady-state mRNA levels and holoenzyme activity. These results suggest t hat, although there may be regulation of MCM activity at the level of mRNA, the significance of genetic regulation is unclear owing to the p resence of epigenetic regulation of holoenzyme formation.