Cloning of the human keratin 18 gene and its expression in nonepithelial mouse cells.
Human keratin 18 (K18) and the homologous mouse protein, Endo B, are intermediate filament subunits of the type I keratin class. Both are expressed in many simple epithelial cell types including trophoblasts, the first differentiated cell type to appear during mouse embryogenesis. The K18 gene was identified and cloned from among the 15 to 20 similar sequences identified within the human genome. The identity of the cloned gene was confirmed by comparing the sequence of the first two exons to the K18 cDNA sequence and transfecting the gene into various murine cell lines and verifying the encoded protein as K18 by immunoprecipitation and partial peptide mapping. The transfected K18 gene was expressed in mouse HR9 parietal endodermal cells and mouse fibroblasts even though the fibroblasts fail to express endogenous Endo B. S1 nuclease protection analysis indicated that mRNA synthesized from the transfected K18 gene is initiated at the same position as authentic K18 mRNA found in both BeWo trophoblastoma cells and HeLa cells. Pulse-chase experiments indicated that the human K18 protein is stable in murine parietal endodermal cells (HR9) which express EndoA, a complementary mouse type II keratin. Surprisingly, however, K18 was degraded when synthesized in cells which lack a type II keratin. This turnover of K18 may be an important mechanism by which epithelial cells maintain equal molar amounts of both type I and II keratins. In addition, the levels of the endogenous type I Endo B in parietal endodermal cells were compensatingly down regulated in the presence of the K18 protein, while the levels of the endogenous type II Endo A were not affected in any of the transfected cell lines.
Kulesh
D A
DA
Cancer Research Center, La Jolla Cancer Research Foundation, California 92037.
Oshima
R G
RG
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D005091:Exons; D005796:Genes; D006801:Humans; D007633:Keratins / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010957:Plasmids; D014158:Transcription, Genetic; D014162:Transfection
D007633:Keratins; D004262:DNA Restriction Enzymes