Phorbol acetate enhances the phosphorylation of cytokeratins 8 and 18 in human colonic epithelial cells.
The phosphorylation of epithelial-specific cytokeratin (CK) 8 and 18 was studied in the human colonic cell line HT29. Metabolic labelling of cells with orthophosphate resulted in phosphorylation of cytokeratins 8/18 on serine residues. When phorbol acetate was added to labelled cells, a 2.2-fold increase in CK8/18 phosphate labelling was noted, whereas increasing intracellular cAMP levels using forskolin or 8-Br-cAMP showed no significant change in CK phosphorylation. CKs8/18 were also phosphorylated by added PKC in the presence of [gamma-32P]ATP. Tryptic peptide map analysis of the phosphorylated CK8 species showed that treatment of cells with 8-Br-cAMP or phorbol acetate generated a phosphopeptide not seen in control cells. In contrast, tryptic peptide maps of phosphorylated CK18 showed no discernable differences. Our results support a role for PKC in the phosphorylation of epithelial cytokeratins, with some phosphorylation sites being modulated by cAMP dependent protein kinase.
Chou
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CF
Stanford University School of Medicine, Gastroenterology Division, CA 94305-5100.
Omary
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MB
D015153:Blotting, Western; D005576:Colforsin / Q000494:pharmacology; D003106:Colon / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007633:Keratins / Q000378:metabolism*; D010449:Peptide Mapping; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D005576:Colforsin; D007633:Keratins; D000242:Cyclic AMP; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate