Restoration by glycerol of p53-dependent apoptosis in cells bearing the mutant p53 gene

Abstract

Purpose: Effective heat-induced cell death in cultured cells bearing a mutant p53 (mp53) gene was sought by glycerol treatment which led to conformational change from mp53 to wild-type p53 (wtp53) in p53-null murine fibroblasts transfected with mp53. Materials and methods: Heat sensitivity was measured using a colony-forming assay. For heat-induced apoptosis, gel electrophoresis was applied to detect DNA fragmentation and Hoechst33342 staining for apoptotic bodies. Glycerol (0.6 m) was applied to the cultured cells 48 h before heating at 44 C in a water bath. Results: Wtp53 transfectants (MT158/wtp53-1) were sensitive to heat stress compared with mp53 transfectants (MT158/mp53-2 cells), and the combined treatment with glycerol enhanced cell killing only in the MT158/mp53-2 cells. After glycerol pretreatment for 48h, the subsequent heat treatment enhanced DNA fragmentation and apoptosis in MT158/mp53-2 cells, while DNA fragmentation and apoptotic bodies were not enhanced with heat treatment alone in these cells. In contrast, DNA fragmentation or apoptotic bodies were clearly observed in MT158/wtp53-1 cells 3-24h after heat treatment. Treatment with glycerol alone did not induce apoptosis in the transfectants. Conclusions: Glycerol appears to function as a chemical chaperone that restores mp53 to wtp53 function.