Activation in Human Colon Carcinoma

We measured the in vitro protein-tyrosine kinase activity of pp60C-" from human colon carcinoma cell lines and tumors. The activity of pp6O¢-'Cl from six of nine carcinoma cell lines was higher (on average, fivefold as measured by enolase phosphorylation, or eightfold as measured by autophosphorylation) than that of pp6OC" from normal colonic mucosal cells, or human or rodent fibroblasts. Similarly, the activity of pp60Cc from 13 of 21 primary colon carcinomas was fiveor sevenfold higher than that of pp60-n? from normal colonic mucosa adjacent to the tumor. The increased pp6Ocr activity did not result solely from an increase in the level of pp60'C" protein, suggesting the specific activity of the pp6OC`SC kinase is elevated in the tumor cells. pp60oC from colon carcinoma cells and normal colonic mucosal cells was phosphorylated at similar sites. We used immunoblotting with antibodies to phosphotyrosine to identify substrates of protein-tyrosine kinases in colonic cells. Three phosphotyrosine-containing proteins were detected at significantly higher levels in most colon carcinoma cell lines than in normal colonic mucosal cells or human or rat fibroblasts. All colon carcinoma cell lines with elevated pp60C4DC in vitro kinase activity, showed increased phosphorylation of proteins on tyrosine in vivo, suggesting the presence of an activated protein-tyrosine kinase(s).