p53-mediated regulation of cell cycle progression: pronounced impact of cellular microenvironment

Abstract

Data on the biological effects of some overexpressed oncogenes and their cooperation with cellular factors are, at least partially, contradictory. There are reports on the strong pro-apoptotic action of temperature-sensitive (ts) p53(135val) in transformed cells at permissive temperature. However, in our experience very high levels of p53(135val) induce in transformed rat cells at permissive temperature cell cycle arrest but not apoptosis. Comparison of the experimental protocols reveals that cells used for transfection strongly differ. Therefore, we decided to explore the impact of primary cells used for generation of cell clones on the biological effects evoked by p53 and c-Ha-Ras. In the present study, we used primary rat cells (RECs) isolated from rat embryos of different age: at 13.5 gd (y) and 15.5 gd (o). We immortalized rat cells using ts p53(135val) mutant and additionally generated transformed cells after co-transfection with oncogenic Ras. The RECs were transfected with a constitutively activated Ha-Ras protein, a mutation that is found in a wide variety of human tumors. The ts p53(135Val) mutant, switching between wild-type (wt) and mutant conformation, offers the possibility to study the escape from p53-mediated cell cycle control in a model of malignant transformation in cells with the same genetic background. Surprisingly, the kinetics of cell proliferation at non-permissive temperature and that of cell cycle arrest at 32 degrees C strongly differed between cell clones established from yRECs and oRECs, thereby indicating that overexpression of genes such as ts p53(135Val) mutant and oncogenic-Ha-Ras does not fully override the intrinsic cellular program.