Characterization of the cDNA and Gene for the Heart/Muscle Isoform of Cytochrome C Oxidase Subunit VIII
Brown, Jessica Sutherland
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The mouse heart cDNA library in Lambda ZAP ll was screened for the heart/muscle isoform of cytochrome c oxidase (COX) subunit VIII. Two clones were isolated and sequence data showed a 100% nucleotide sequence identity that encoded a protein 70 amino acids long with a 24 amino acid presequence and a 46 amino acid mature protein. Comparisons of the mouse cDNA sequence with the rat and bovine cDNAs helped to identify the C-terminal hydrophilic region as exhibiting the greatest conservation. A stretch of 6 completely conserved amino acids was identified that could later be used to prepare degenerate probes for evolutionary comparisons among primates. Humans have been shown to lack the heart isoform of COX subunit VIII and this study may help identify when in evolutionary history the gene was lost. The 5' flanking region of the mouse COX8 heart gene was also sequenced and proximal promoter and enhancer elements were discovered by comparing conserved sequences with known bovine enhancer elements. The transcription start site in the bovine sequence was highly conserved with a region in the mouse sequence though the start site has not yet been mapped in the mouse. A TATAA box and two E-box elements, one on either side of the TATAA box, were identified in the mouse gene that were conserved with sequences in the bovine promoter demonstrated to increase transcription in myotubes. Therefore, these elements may play a role in tissue specific expression of the gene in skeletal muscle in mice. Moreover, a potential CArG box or NRF-2 site was also identified in the 5' untranslated region of the gene that may also regulate expression.With honors.