The nuclear protein and phosphoprotein profiles from 3 subpopulations of human colonic carcinoma cells which expressed different levels of neoplastic properties were characterized by two-dimensional electrophoresis. The silver stained nuclear protein profiles were found to be remarkably similar among the subpopulations. However, 2 types of nuclear proteins were found to be selectively modified by phosphorylation/dephosphorylation reactions. The dephosphorylation of type I and the phosphorylation of type II nuclear proteins were found to be associated with the HCT 116a subpopulation which expressed a high level of neoplastic properties. Conversely, the phosphorylation of type I and the dephosphorylation of type II nuclear proteins were found to be associated with the HCT 116b subpopulation, which expressed a low level of neoplastic properties. The HCT 116 subpopulation, which expressed an intermediate level of neoplastic properties, was found to possess an intermediate phosphoprotein profile relative to that of the other two subpopulations. Selective modification of cellular proteins by phosphorylation/dephosphorylation reactions may be involved in the generation of tumor cell diversity and heterogeneity.