PRO-TRANSFORMING GROWTH-FACTOR-ALPHA PROCESSING IN HUMAN COLON-CARCINOMA CELLS - ROLE OF PROTEIN-KINASE-C

The human colon cancer cell lines HCT 116 (poorly differentiated) and GEO (well differentiated) express the mitogenic peptide transforming g rowth factor alpha (TGF-alpha). The secretion of TGF-alpha was enhance d by phorbol 12-myristate 13-acetate (PMA), indicating the possible ro le of protein kinase C (PKC) in the formation of mature TGF-alpha. Cel ls were metabolically labeled with S-35-cysteine and the formation of the mature 6 kDa TGF-alpha polypeptide from the 17 kDa pro-TCF-alpha p recursor was determined. The conversion of pro-TGF-alpha was complete in 2-4 hr with the HCT 116 cells showing faster kinetics of TGF-alpha formation than GEO cells. HCT 116 cells secreted more TGF-alpha than C EO cells and the rate and extent of formation of TGF-alpha was enhance d by PMA in both cell lines. The expression of several PKC isozymes by HCT 116 and CEO cells was examined by immunoblotting. The expression of all isozymes examined was higher in HCT 116 cells compared with GEO cells. Calphostin C, an inhibitor of PKC, reduced the enzyme activity and significantly inhibited the PMA-induced secretion of TGF-alpha by both cell lines. Two agonists of PKC that act on specific PKC isozyme s, thymeleatoxin and 12-deoxyphorbol 13-phenylacetate 20-acetate (dPPA ), stimulated the release of TGF-alpha into the medium to the same ext ent as PMA. Since dPPA has been reported to stimulate PKC-BI specifica lly, our results suggest a potential role for PKC-B in the processing of pro-TGF-alpha by these 2 human colon carcinoma cell lines. (C) 1995 Wiley-Liss, Inc.