CELL-SPECIFIC FUNCTION OF CIS-ACTING ELEMENTS IN THE REGULATION OF HUMAN ALCOHOL-DEHYDROGENASE-5 GENE-EXPRESSION AND EFFECT OF THE 5'-NONTRANSLATED REGION

The human alcohol dehydrogenase 5 gene (ADH5) differs from all other human alcohol dehydrogenase genes in its ubiquitous expression, although there are tissue-specific differences in the level of expression, To under stand the expression of ADH5, we characterized the structure and function of its 5' region by DNase I footprinting and transient transfection assays. The region from base pair (bp) -34 to +61, flanking the major transcription start site, had strong promoter activity in three different cell lines: HeLa, H4IIE-C3, and CV-1, and could explain the ubiquitous expression, Two Sp1 sites within that region are footprinted by nuclear extracts from all tissues and cells tested. There are sites further upstream that show cell- and tissue-specific differences in both their patterns of occupancy and their effects on promoter activity. The region between bp -34 and -64 strongly increases promoter activity in H4IIE-C3 cells, weakly activates in CV-1 cells, but has no effect in HeLa cells. The region between bp -127 and -163 is a positive element in both HeLa cells and CV-1 cells, but is a negative regulatory element in H4IIE-C3 cells. These differences in part explain the levels of expression of ADH5 in various tissues. Two regions (bp -64 to -127 and bp -163 to -365) contain negative regulatory elements that reduce promoter activity in all three cells. The 5'-nontranslated region of ADH5 contains two upstream ATGs. Insertion of 12 bp within the putative coding region of these upstream ATGs led to a 1.6-2.3-fold increase in activity. This suggests that the 5'-nontranslated region has regulatory significance.