Human keratin 18 (K18) and its mouse form Endo B are type I keratin intermediate filament proteins that polymerize with their co- expressed, complementary, type 11 keratin protein partners, K8 and Endo A, respectively. Endo B is first expressed at the 48 cell stage of mouse development and appears to be an excellent marker of the first differentiated tissues to form. In adults, K18 and Endo B expression is restricted to a variety of simple epithelial cell types. The tissue-specific expression of human and mouse K18 can be considered intermediate between constitutively expressed """"""""house-keeping"""""""" functions and very specialized differentiation products such as beta-globins or even epidermal keratins. The objectives of this proposal are to determine the molecular mechanisms responsible for the tissue specific expression of K18 and the function of K18 intermediate filaments during development. The DNA elements of the coding K18 and Endo B genes necessary for tissue-specific transcription of the genes in differentiated somatic tissues will be identified by testing recombinant DNA constructions in cultured cells, experimental mouse teratocar- cinomas and transgenic mice. Possible differences in the expression of KI8 in preimplantation human and mouse embryos will be investigated in teratocarcinoma stem cell lines and the embryos of transgenic mice. Proteins which bind to regulatory regions of the genes will be identified. The role of DNA methylation in suppressing the expression of K18 and Endo B will be examined. The biological function(s) of K18 and Endo B will be investigated by expressing mutant proteins in cultured cells and mice and by generating transgenic mouse strains deficient in Endo B by experimental homologous recombination. These studies should provide a paradigm for the developmental regulation of genes which have an intermediate type of restricted expression in somatic tissues and help elucidate how differential gene expression of the first differentiated cell types of the mammalian embryo is controlled. Knowledge of how K18 is regulated is important because of the increasing use of intermediate filaments as indicators of the tissue origin of human tumors and to our general understanding of how genetic information is translated into animal form.
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