The long term goals of this research are to determine the structures of mammalian alcohol dehydrogenase (ADH) genes and the mechanisms which regulate their expression. These studies will contribute to our understanding of genetic factors underlying differences among individuals in the metabolic, pharmacologic and pathological effects of alcohol consumption. This proposal focuses upon control of ADH expression. Specific Aims 1 through 4 involve analysis of the regulation of ADH expression in different tissues of the mouse. ADH enzyme activity, immunoreactive protein, and mRNA will be analyzed in several tissues of inbred strains of mice with high and low liver ADH to determine whether the amount of ADH activity is determined by the steady-state level of its mRNA. In liver and kidney, the relative ADH activity and mRNA content correspond, whereas in stomach we have preliminary evidence that they do not. This suggests an interesting regulatory mechanism in stomach. The effects of chronic ethanol exposure upon ADH expression in liver will also be analyzed. The apparent linkage between a locus controlling ADH expression in liver and restriction fragment polymorphisms in the Adh-1 locus will be tested using recombinant inbred strains of mice. ADH-B cDNA will be isolated. Specific Aims 5 through 7 involve determination of the sequences that control both tissue-specificity and quantitative expression of mouse ADH. The Adh-1 genes of mice with high and low liver ADH will be cloned, and their structures and sequences compared. The sequences important in the control of tissue specificity of Adh-1 expression will be analyzed by constructing chimeric genes with the putative control sequences attached to an easily assayable reporter gene such as chloramphenicol acetyltransferase. The expression of these chimeric genes will be assayed in cultured rodent and human cells of hepatic and non- hepatic origin. Similar analyses will be carried out to determine which sequences are involved in quantitative control of ADH by comparing control regions from high and low activity strains, again using both human and rodent cells to test whether such sequences function across species barriers. Specific Aims 8 and 9 involve analysis of the control of ADH expression in human liver. There are several sized of beta1 mRNA in human livers, which vary in the length of their 3 non- translated regions. The size distributions of human liver mRNA from all three Class I ADH loci will be analyzed. The relative efficiency with which the multiple potential polyadenylation signals in the human beta1 gene are utilized will be determined using chimeric genes, and the potential effects upon expression will be analyzed. Differences in the 3' ends could be a mechanism for control of the quantity of ADH in liver.