The long range objective of this research is an understanding of the biochemical mechanisms that operate and regulate the expression of human genes during growth and differentiation. The human epidermis has been chosen as a model system, since we are able to serially cultivate these cells in vitro under conditions where many of the differentiative properties, including stratification are retained. The major differentiation-specific proteins in these cells are the keratins, a group of closely related proteins of MW 40-70 K daltons that form 80 Angstrom cytoskeletal filaments. Only a subset of these keratins are ever expressed by an epidermal cell at any one time. During the course of differentiation, this subset changes concomitantly with an increase in keratin synthesis, leaving the fully differentiated epidermal cell with 85% of its total protein as keratins. We already know that early changes in the differentiation involve changes in mRNAs whereas late changes involve proteolytic processing. Recently, we isolated near full-length cDNA clones to the different epidermal keratins mRNAs and we showed that there are two distinct classes of keratins. We would now like to determine the nucleic acid sequence of these cDNAs. Since no amino acid sequence data is available, the amino acid sequence predicted from the cDNA sequence will be essential in determining the structural and functional relationship between these two classes of keratins. We will also isolate and characterize genomic clones for the keratins. This will aid us in determining the complexity and chromosomal organization of the keratin genes. We will then prepare subclones of these sequences to probe posttranscriptional regulation of keratin gene expression. Finally, epidermal cells from patients with genetic skin diseases are currently being cultured and alterations in the regulation and expression of the keratins and other differentiation-specific proteins in these cells will be investigated.