TUMOR-NECROSIS-FACTOR-ALPHA - POSTTRANSCRIPTIONAL STABILIZATION OF WAF1 MESSENGER-RNA IN P53-DEFICIENT HUMAN LEUKEMIC-CELLS

The p53 protein directly regulates the expression of the WAF1 (wild-ty pe p53-activated fragment 1) protein which is a cyclin-dependent kinas e inhibitor (CDKI). DNA damaging agents such as ionizing or UV radiati on, and some chemical agents induce WAF1 in wild-type p53 containing c ells, thereby halting cell cycle progression. WAF1 expression is also induced through a p53-independent pathway. Tumor necrosis factor alpha (TNF alpha) is a cytotoxic/cytostatic compound for some human cancer cells. We examined a series of myeloid leukemic cell lines that expres sed either no p53 (HL-60, K562) or mutant inactive p53 (KG-1, KCL22, T HP-1, U937). The KG-1, HL-60, K562, and KCL22 myeloid leukemic cells i ncreased their levels of WAF1 mRNA in the presence of TNF alpha. We fo cused on KG-1 cells to determine how TNF alpha modulated WAF1 expressi on. WAF1 mRNA increased in a dose-dependent manner in the cells after exposure to increasing concentrations of TNF alpha, and this increase occurred in the absence of new protein synthesis. An increase of WAF1 protein and a concominant decrease of cyclin-dependent kinase 2 activi ty also was found in KG-1 cells. Flow cytometry using 5-bromo-2'-deoxy uridine showed an increase in the proportion of TNF alpha-treated KG-1 cells in the G(0)/G(1) phase of the cell cycle. TNF alpha enhanced th e rate of WAF1 transcription only 1.4-fold in TNF alpha-treated KG-1 c ells as compared to untreated cells. Notably, however, the half-life ( t 1/2) of WAF1 mRNA in TNF alpha-treated cells was 2.5 hours as compar ed to 0.5 hours in untreated cells. These results indicate that TNF al pha increases WAF1 levels at least in part via a posttranscriptional s tabilization of the mRNA; and TNF alpha may mediate its cytostatic eff ects through WAF1 in some cell types. (C) 1996 Wiley-Liss, Inc.