Enhancer elements directing cell-type-specific expression of cytokeratin genes and changes of the epithelial cytoskeleton by transfections of hybrid cytokeratin genes.

摘要

The cytokeratins, which form the intermediate filaments (IFs) characteristic of epithelial cells, are encoded by a large family of genes whose members are differentially expressed in patterns different in the various kinds of epithelia. To identify possible cis-regulatory DNA elements involved in the cell-type-specific expression of these genes, we examined, in transfection assays, 5' upstream sequence intercepts of a certain cytokeratin gene, i.e. that for bovine cytokeratin IV* (CKIV*), in combination with the coding portions of either the chloramphenicol acetyltransferase (CAT) gene or other cytokeratin genes. A 5' upstream region located between the cap-site and nucleotide -605 was found to enhance the specific expression of these reporter genes in bovine mammary gland-derived BMGE + H cells, which express the endogenous gene, but not in bovine kidney epithelium-derived MDBK cells which synthesize cytokeratins other than IV*. This epithelium-type-specific expression was also observed in heterologous combinations, e.g. in murine keratinocytes, but not in other murine cell lines such as 3T3 fibroblasts. When a fragment located between -180 and -605 was coupled to the HSV-TK promoter it stimulated the expression of the reporter gene in a cell-type-specific manner. The enhancer character of this 425 nucleotide long region is also demonstrated. Moreover, the CKIV* promoter/enhancer complex was able to direct the expression of epidermal cytokeratins characteristic for suprabasal differentiation, i.e. bovine cytokeratins Ia and VIb, in cells that normally do not express these genes. We show that the newly synthesized cytokeratins integrate into the pre-existing cytokeratin IF system of the transfected cells and that the forced expression of one of these cytokeratins does not induce the endogenous gene encoding its normal pair partner.