Cloning and expression of rat pancreatic beta-cell malonyl-CoA decarboxylase

To gain insight into the function and regulation of malonyl-CoA decarboxyla se (MCD) we have cloned rat MCD cDNA from a differentiated insulin-secretin g pancreatic beta-cell-line cDNA library. The full-length cDNA sequence sho ws 69 % identity with the cDNA cloned previously from the goose uropygial g land, and predicts a 492 amino acid protein of 54.7 kDa. The open reading f rame contains an N-terminal mitochondrial targeting sequence and the C-term inal part of the enzyme ends with a peroxisomal (Ser-Lys-Leu) targeting mot if. Since the sequence does not reveal hydrophobic domains, MCD is most lik ely expressed in the mitochondrial matrix and inside the peroxisomes, A sec ond methionine residue, located 3' of the mitochondrial presequence, might be the first amino acid of a putative cytosolic MCD, since the nucleotide s equence around it fits fairly well with a consensus Kozak site for translat ion initiation. However, primer extension detects the presence of only one transcript initiating upstream of the first ATG, indicating that the major, if not exclusive, transcript expressed in the pancreatic beta-cell encodes MCD with its mitochondrial presequence. The sequence also shows multiple p ossible sites of phosphorylation by casein kinase II and protein kinase C. mRNA tissue-distribution analysis indicates a transcript of 2.2 kb, and tha t the MCD gene is expressed over a wide range of rat tissues. The distribut ion of the enzyme shows a broad range of activities from very low in the br ain to elevated in the liver and heart. The results provide the foundations for further studies of the role of MCD in lipid metabolism and metabolic s ignalling in various tissues.