Transcriptional control of the mouse prealbumin (transthyretin) gene: both promoter sequences and a distinct enhancer are cell specific.

The mouse genomic clone for the prealbumin (transthyretin) gene was cloned, and its upstream regulatory regions were analyzed. The 200 nucleotides 5' to the cap site when placed within a recombinant plasmid were sufficient to direct transient expression in HepG2 (human hepatoma) cells, but this DNA region did not support expression in HeLa cells. The sequence of the 200-nucleotide region is highly conserved between mouse and human DNA and can be considered a cell-specific promoter. Deletions of this promoter region identified a crucial element for cell-specific expression between 151 and 110 nucleotides 5' to the RNA start site. A region situated at about 1.6 to 2.15 kilobases upstream of the RNA start site was found to stimulate expression 10-fold in HepG2 cells but not in HeLa cells. This far upstream element was invertible and increased expression from the beta-globin promoter in HepG2 cells. Unlike the simian virus 40 enhancer, the prealbumin enhancer would not stimulate beta-globin synthesis in HeLa cells, and even the simian virus 40 enhancer did not stimulate the prealbumin promoter in HeLa cells. Thus, we identified in the prealbumin gene two DNA elements that respond in a cell-specific manner: a proximal promoter including a crucial sequence between -108 and -151 nucleotides and a distant enhancer element located between 1.6 and 2.15 kilobases upstream.     

Structural analysis of human pyruvate kinase L-gene and identification of the promoter activity in erythroid cells.

The human pyruvate kinase (PK) L-gene is organized in 12 exons over 9.5 kilobases, and the first and second exons are specifically transcribed to the R- and L-type PK mRNA. The 5'-flanking region upstream the first exon has two CAC boxes and four GATA motifs within 250 bp from the translational initiation codon. Comparison with the rat L-gene revealed four well-conserved elements in the region. The transient transfection demonstrated that the 270-bp upstream region was a powerful promoter in K562 cells, whereas deletion of the distal 150-bp sequences, which included three GATA motifs, resulted in drastic reduction of the activity. When the hypersensitive site 2 of the human beta-globin gene locus was joined to the promoter, the activity of the proximal 120-bp region was enhanced. We concluded that the proximal 120-bp region had a basal promoter activity and that the distal 150-bp region acted as an enhancer in erythroid cells.