Figure 3.

Characterization of reporter activity and spindle cell cycle checkpoint induced by a transcription-deficient p53 mutant protein. (A) NHF3 cells were transiently cotransfected with 5 μg of MDR1 CAT reporter plasmid and 10 μg of either p53–281G or p53–281G,22,23 using Lipofectamine. After 48 h, cells were lysed and CAT activity was assayed using ¹⁴C-labeled chloramphenicol (31). CAT activity was observed only in cells cotransfected with the reporter alone nor cells cotransfected with p53-281G,22,23, and MDR1 CAT (lanes 1 and 3). (B) Flow cytometric analysis of NHF stably transfected with either p53–281G or p53–281G,22,23 in the presence (b and d) or absence (a and c) of colcemid. Colonies expressing mutant p53 were pooled and processed for cell cycle distribution of DNA content after colcemid treatment for 48 h as described in Fig. 1. Flow cytometry documented a decrease in apoptotic cells but no increase in polyploid cells.