The long-term objective of the proposed research is to identify and characterize trans-acting cellular factors that determine the tissue-specific transcription of genes, and that therefore determine mammalian cell type. My approach to this problem is to study the trans-acting factors that permit the mouse serum albumin gene to be transcribed in liver hepatocyte cells but not in other mammalian cell types. The research proposal addresses this issue with the following three goals: 1. To identify cis-acting DNA sequences that confer liver-specific transcription of the mouse serum albumin gene. 2. To fuse the cis-acting sequences to genetically selectable markers and to use the gene fusions to select for cultured cell mutants that are defective in putative trans-acting determinants of liver-specific transcription. We will perform selections for liver-derived cell mutants that fail to express albumin fusion genes and for nonliver-derived cell mutants that express the fusion genes inappropriately. 3. To identify and characterize nuclease hypersensitive sites and protein footprints in the chromatin of the serum albumin gene as an independent means to identify and monitor putative trns-acting factors. By comparing these sites in different tissues of fetal and adult mice and in clutured cells, we may detect the activity of factors in the intact organism that are missing in cell culture. The proposed experiments are designed so that in the future we can readily clone the genes encoding liver-specific trans-acting factors. Understanding the action of factors that determine tissue-specific gene expression and mammalian cell type is fundamental to the study of biological development in general, and to the treatment of human disease and developmental defects in particular.