Selective Protein Phosphorylation in Heterogeneous Subpopulations of Human Colon Carcinoma Cells

Endogenous membrane and cytosolic and nuclear protein phosphorylations were compared among three well-characterized subpopulations of human colonic carcinoma cells that were originally isolated from a single human primary colon tumor. These intratumoral subpopulations of cells were found to differ significantly in their biological properties. Analysis of phosphoproteins by two-dimensional electrophoresis following ³²P phosphorylation of subcellular fractions in a cell-free system or labeling intact cells in vivo revealed significant differences in the selective phosphorylation of membrane, cytosol, and nuclear proteins. The two-dimensional membrane, cytosol, and nuclear phospho-protein profiles distinguished the three subpopulations of colonic carcinoma cells from each other. Silver-staining proteins from the three subpopulations were also compared. The two-dimensional, silver-stained electrophoretic profiles of nuclear proteins were essentially the same for all three subpopulations. The silver-stained electrophoretic profile of membrane and cytosolic proteins revealed only minor differences in the expression of polypeptides. Nevertheless, these changes could also distinguish the three subpopulations. The results of this study suggest that minor differences in the expression of cytosolic and membrane proteins exist in intratumoral subpopulations of colonic cells. However, a significantly greater degree of heterogeneity was found to be associated with posttranslational modification of proteins by phosphorylation and/or dephosphorylation. These modifications could play an important role in determining the expression of different biological properties among subpopulations of malignant cells.