A temperature-sensitive mutant of murine p53 (p53va-135) dimethyl sulfoxide (DMSO) (19). We and others have previously shown that constitutive expression of either c-myc or c-myb prevents MEL cell differentiation (6,7,12,24,42), and expression of c-myb late in the process of differentiation is sufficient to block terminal differentiation (8,34). In many MEL cell lines, there is a lack of expression of the p53 (reviewed in reference 27) tumor suppressor gene or expression of a mutant p53 protein (29, 41). Furthermore, wild-type p53 could not be stably expressed by a p53-negative MEL cell line (22).Myeloid leukemic cell lines frequently fail to express p53, in contrast to lymphoid leukemic cell lines, which characteristically overexpress p53 (9,26,43). However, p53 is often detectable in primary human myeloid leukemic blasts, though the level of expression is low (23,47). p53 expression is also seen in normal human blast cells and increases with maturation (45). The myeloid leukemia cell line ML 1, which has wild-type p53 genes, lacks expression during logarithmic growth but shows increased expression with induced differentiation (23

A temperature-sensitive mutant of murine p53 (p53Val-135) was transfected by electroporation into murine erythroleukemia cells (DP16-1) lacking endogenous expression of p53. While the transfected cells grew normally in the presence of mutant p53 (37.5 degrees C), wild-type p53 (32.5 degrees C) was associated with a rapid loss of cell viability. Genomic DNA extracted at 32.5 degrees C was seen to be fragmented into a characteristic ladder consistent with cell death due to apoptosis. Following synchronization by density arrest, transfected cells released into G1 at 32.5 degrees C were found to lose viability more rapidly than did randomly growing cultures. Following release into G1, cells became irreversibly committed to cell death after 4 h at 32.5 degrees C. Commitment to cell death correlated with the first appearance of fragmented DNA. Synchronized cells allowed to pass out of G1 prior to being placed at 32.5 degrees C continued to cycle until subsequently arrested in G1; loss of viability occurred following G1 arrest. In contrast to cells in G1, cells cultured at 32.5 degrees C for prolonged periods during S phase and G2/M, and then returned to 37.5 degrees C, did not become committed to cell death. G1 arrest at 37.5 degrees C, utilizing either mimosine or isoleucine deprivation, does not lead to rapid cell death. Upon transfer to 32.5 degrees C, these G1 synchronized cell populations quickly lost viability. Cells that were kept density arrested at 32.5 degrees C (G0) lost viability at a much slower rate than did cells released into G1. Taken together, these results indicate that wild-type p53 induces cell death in murine erythroleukemia cells and that this effect occurs predominantly in the G1 phase of actively cycling cells.