CHARACTERIZATION OF THE 5'-FLANKING SEQUENCE OF RAT CLASS-I ALCOHOL-DEHYDROGENASE GENE

Expression of the rat class I alcohol dehydrogenase (ADH) gene is high est in the liver and regions of the intestine. We characterized over 3 kilobases of the gene's 5'-flanking region by sequencing and transien t transfection. Alignment of the flanking sequence of the rat gene wit h those of the mouse and human class I genes revealed a cis-acting ele ment, known to be a functional glucocorticoid response element in the human gene and conserved in the mouse, is interrupted in the rat promo ter by a 490-base pair processed retropseudogene of the ribosomal prot ein S25. Southern analysis indicated that this inserted element is pre sent in the class I ADH promoters of multiple strains of rat. Transfec tion analysis of the rat and mouse promoters showed that the mouse, bu t not the rat promoter, is inducible by dexamethasone. Electrophoretic mobility shift assays using nuclear extracts from dexamethasone-treat ed cells confirmed that the mouse's element interacts with the glucoco rticoid receptor. Transient transfection of the 5'-flanking region of the rat gene linked to a human growth hormone reporter demonstrated th e liver and intestinal specificity of the rat promoter. Two positive e lements, one from nucleotides -1,327 to -977 and the other from -241 t o -12, were shown to support high levels of reporter activity. In addi tion, a suppressive element was localized between nucleotides -403 and -241, a region of DNA situated within the domain of the S25 ribosomal protein pseudogene.