The goals of this project are to determine the diverse factors controlling the expression of mammalian alcohol dehydrogenase genes (Adh). Alcohol dehydrogenase (ADH) is one of the key enzymes of ethanol metabolism in liver and excessive ethanol metabolism via this pathway is hypothesized to lead to liver disease. A genetic and molecular system exists in the mouse to explore Adh gene regulation. Regulatory genes modulating level of expression of Adh-1 in liver and kidney and Adh-3 in stomach, lung and reproductive tissues have been identified. cDNA and genomic clones of Adh-1 have been sequenced. The Adh-1 is transcriptionally activated by androgen in kidney. Adh-1 mRNA changes in content during liver development and it will be determined if this is under transcriptional control. Adh-1 mRNA deficient deermice are available as are Mus species (saxicola and cookii) which are not androgen inducible. Sequence comparisons of the 5'-end of the Adh-1 gene between strains and closely related species will be used as a way of defining sequences important to the regulation of Adh-1. The 5'- end of Adh-1 will be sequenced from M. cookii and M. saxicola, C57BL/6 and the B6.Adh-1a congenic (high kidney Adh-1 mRNA relative to other strains). Specific sequences involved in the androgen respone will be further delineated by transfecting in vitro mutated genes into androgen-respnosive cell lines, and using tissue specific transcription systems. Proteins interacting with the Adh-1 gene during development and after androgen stimulation will be identified, their binding sites determined, and their significance will be studied by mutating those sites. In order to more fully identify sequences and factors in the Adh-1 gene essential for correct expression, genes mutated in vitro will be used to produce transgenic mice. Tissue expression, developmental regualtion, and androgen inducibility will be studied in transgenic mice possessing mutating Adh-1 genes. Promising prospects for cloning Adh-3 and/or Adh-2 appear on the horizon, and these may already be available. These genes are expressed differently than Adh-1 and comparisons of presumptive regulatory sequences in these related genes should provide useful information regarding tissue specific expression of mammalian genes important in ethanol metabolism.
