Organization of the 5' end of the rat gamma-glutamyl transpeptidase gene: structure of a promoter active in the kidney.

ABSTRACT

Two gamma-glutamyl transpeptidase mRNAs (mRNAI and mRNAII), with alternate 5'-untranslated regions, are expressed in the rat kidney. Oligonucleotides were designed based upon these two alternate 5' sequences and used as primers to amplify GGT genomic DNA sequences. The genomic organization of the mRNAI and mRNAII 5'-untranslated sequences reveals that the mRNAs are encoded from two separate promoters which are 2.1 kbp apart on the single GGT gene. A 2775 base pair genomic sequence, which contains the proximal GGT promoter, was cloned from two overlapping amplified fragments. S1 mapping analysis shows that the kidney GGT mRNAI is transcribed from several start sites on this promoter which displays neither a classical TATA box nor Sp1 binding sites. Chimeric plasmids, including the GGT promoter region for mRNAI, associated with the chloramphenicol acetyltransferase (CAT) reporter gene, were transiently expressed in a kidney (LLCPK) and in a hepatoma (HTC) cell line. A sequence extending 308 bases upstream from the major GGT mRNAI start site drives a promoter activity which is 5-fold higher in LLCPK than in HTC cells and is sufficient to confer cell specificity to the GGT proximal promoter.