Cloning of bovine pyruvate carboxylase and 5′ untranslated region variants

Bovine pyruvate carboxylase (PC; EC 6.4.1.1) cDNA was cloned by reverse transcription (RT) PCR. The coding region plus 3' untranslated region (UTR) of PC mRNA is 3926 bases and encodes 1178 amino acid PC precursor protein. A 5' rapid amplification of cDNA ends protocol was used to clone the 5' end of the mRNA. Six 5'UTR variants ranging from 68 to 363 bp were cloned. Bovine PC 5'UTR (bPC5') variants contain 68 (bPC5'A), 263 (bPC5'B), 363 (bPC5'C), 89 (bPC5'D), 275 (bPC5'E), and 178 bp (bPC5'F). All variants contain a common coding sequence. An RNase protection assay and RT-PCR analysis confirms the presence of the 5'UTR variants. The abundance of PC mRNA, determined by Northern blot analysis, indicates that PC is more abundant in gluconcogenic and lipogenic tissues where all PC variants are expressed compared with tissues that do not possess the full spectrum of PC transcripts. The data suggest that bPC5'A, bPC5'B, and bPC5'F are more abundant in bovine liver than the other variants.