DNA damage triggers a prolonged p53-dependent G1 arrest and long-term induction of Cip1 in normal human fibroblasts.

Abstract

The tumor suppressor p53 is a cell cycle checkpoint protein that contributes to the preservation of genetic stability by mediating either a G^ arrest or apoptosis in response to DNA damage. Recent reports suggest that p53 causes growth arrest through transcriptional activation of the cyclin-dependent kinase (Cdk)-inhibitor Cipl. Here, we characterize the p53-dependent Gj arrest in several normal human diploid fibroblast (NDF) strains and p53-deficient cell lines treated with 0.1-6 Gy gamma radiation. DNA damage and cell cycle progression analyses showed that NDF entered a prolonged arrest state resembling senescence, even at low doses of radiation. This contrasts with the view that p53 ensures genetic stability by inducing a transient arrest to enable repair of DNA damage, as reported for some myeloid leukemia lines. Gamma radiation administered in early to mid-, but not late, G^ induced the arrest, suggesting that the p53 checkpoint is only active in Gj until cells commit to enter S phase at the Gi restriction point. A log-linear plot of the fraction of irradiated GQ cells able to enter S phase as a function of dose is consistent with single-hit kinetics. Cytogenetic analyses combined with radiation dosage data indicate that only one or a small number of unrepaired DNA breaks may be sufficient to cause arrest. The arrest also correlated with long-term elevations of p53 protein, Cipl mRNA, and Cipl protein. We propose that p53 helps maintain genetic stability in NDF by mediating a permanent cell cycle arrest through long-term induction of Cipl when low amounts of unrepaired DNA damage are present in G^ before the restriction point.

Twenty years of p53 research: structural and functional aspects of the p53 protein

“…In addition, embryonic ®broblasts from p53-null mice lose the G1 arrest response to IR . It should be noted that in normal diploid ®broblasts, exposure to ionizing irradiation results in long term p53-dependent arrest with prolonged induction of p21WAF1 (Di Leonardo et al, 1994). The irreversibility of this arrest can be attributed to the inability of these cells to repair even a small number of double strand DNA breaks, so that the activating signal persists (Huang et al, 1996).…”

K-ras modulates the cell cycle via both positive and negative regulatory pathways

“…Several studies indicate that stabilization of the p53 protein might be a major mechanism for p53 accumulation in response to DNA damage (Canman et al, 1994;Di Leonardo et al, 1994). Although most studies show that MCF-7 cells contain wild type p53 (Takahashi et al, 1992;Balcer-Kubiczek et al, 1995), there have been some questions as to the functional status of the p53 tumor suppressor gene product present in MCF-7 cells (Casey et al, 1991), the model cell line used in this study.…”

Targeting inhibitors of apoptosis proteins suppresses medulloblastoma cell proliferation via G2/M phase arrest and attenuated neddylation of p21

“…According to several studies, upregulation of p21 transcription can be through p53‐dependent pathways or p53‐independent pathways 49, 50, 51, 52. The promoter of p21 contains two conserved p53‐binding sites required for p53 responsiveness after DNA damage 53.…”