The long term goal of our studies is to define the pathways and controls of epidermal protein synthesis in health and disease. During the past grant period there has been a dramatic increase in our knowledge of the structural abnormalities in keratin synthesis and their relationship to diseases such as epidermolysis bullosa simplex and epidermolytic hyperkeratosis. Our goal for the next 5 years is to generate equivalent information regarding the quantitative control of keratin gene expression. During the next project period we shall continue to focus on the mechanisms of transcriptional control involving the three pairs of keratins central to normal and pathologic epidermal keratinization - keratins #5 & 14 of the basal cells, keratins #1 & 10 of the differentiated cells and keratins #6 & 16 of hyperproliferative, activated epidermis. Our focus will be on the sites in the keratin gene promoters which bind to general and specific transcription factors, on the transcription factors themselves, on the alterations which occur in different physiologic and pathologic states and on the interrelationships between the control mechanisms involved in keratin synthesis in comparison to those involved in the synthesis of other unique epidermal proteins such as filaggrin, involucrin and loricrin. Our first two goals are mutually dependent. We will extend our studies of the specific transcription factors we have identified for keratin #5 & 6 - the factors which bind to """"""""site A"""""""" in the K #5 gene and to the """"""""PAL"""""""" site in the K #6 gene. Utilizing the tools of protein chemistry and molecular biology, we will isolate, purify, raise antibodies to and ultimately clone the factors. In parallel, we will define the DNA sites and associated binding proteins involved in control of gene expression of the other members of the epidermal keratin family - K #1, K #10, K #14 and K #16. Beginning during the first year of the next grant period, we will utilize the same biochemical, immunologic and molecular biologic reagents developed for the studies above to analyze the biologic function of the transcription control of keratin gene expression in normal keratinization and in pathologic states including benign and malignant epidermal proliferation. There is growing precedent from several other systems that control at this level is extremely important in the normal and pathologic function of tissues such as bone marrow. Our hypothesis is that an equivalent situation exists in epidermis. Ultimately, we will be prepared to define the relationships among the specific transcription factors and binding sites involved in control of keratin gene expression and other phenotypic epidermal constituents including filaggrin, involucrin, and loricrin.
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