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Induction of WAF1/CIP1 by a p53-independent pathway
1Laboratory of Cellular and Molecular Biology, National Cancer Institute, NIH, Bethesda, Maryland 20892.
Abstract:
The p53-inducible gene WAF1/CIP1 encodes a M(r) 21,000 protein (p21) that has been shown to arrest cell growth by inhibition of cyclin-dependent kinases. Induction of WAF1/CIP1 in cells undergoing p53-dependent G1 arrest or apoptosis supports the idea that WAF1/CIP1 is a critical downstream effector of p53. In the present study, we used embryonic fibroblasts from p53 "knock-out" mice to demonstrate p53-independent induction of WAF1/CIP1. We show that serum or individual growth factors such as platelet-derived growth factor, fibroblast growth factor, and epidermal growth factor but not insulin are able to induce WAF1/CIP1 in quiescent p53-deficient cells as well as in normal cells. The kinetics of this transient induction, which is enhanced by cycloheximide, demonstrates that WAF1/CIP1 is an immediate-early gene the transcript of which reaches a peak at approximately 2 h following serum or growth factor stimulation. On the other hand, DNA damage elicited by gamma-irradiation induces WAF1/CIP1 in normal human and mouse fibroblasts but does not affect WAF1/CIP1 expression in p53-deficient cells. These results suggest the existence of two separate pathways for the induction of WAF1/CIP1, a p53-dependent one activated by DNA damage and a p53-independent one activated by mitogens at the entry into the cell cycle. The possible function of p21 at this early stage is discussed.
Insights
The WAF1/CIP1 gene, encoding the p21 protein, is induced independently of p53 by growth factors during cell cycle entry. DNA damage, however, requires p53 for WAF1/CIP1 induction.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- The p53-inducible gene WAF1/CIP1 encodes the p21 protein, known to inhibit cyclin-dependent kinases and arrest cell growth.
- WAF1/CIP1 induction in p53-dependent G1 arrest or apoptosis suggests it's a critical downstream effector of p53.
Purpose of the Study:
- To investigate the induction pathways of WAF1/CIP1.
- To determine if WAF1/CIP1 induction can occur independently of p53.
Main Methods:
- Utilized embryonic fibroblasts from p53 "knock-out" mice.
- Stimulated quiescent cells with serum or individual growth factors (PDGF, FGF, EGF, insulin).
- Examined WAF1/CIP1 expression following gamma-irradiation in normal and p53-deficient cells.
Main Results:
- Demonstrated p53-independent induction of WAF1/CIP1 by serum and growth factors (excluding insulin) in quiescent cells.
- Showed WAF1/CIP1 is an immediate-early gene, with peak transcript levels around 2 hours post-stimulation.
- Confirmed that DNA damage (gamma-irradiation) induces WAF1/CIP1 in normal cells but not in p53-deficient cells.
Conclusions:
- Identified two distinct pathways for WAF1/CIP1 induction: one p53-dependent (DNA damage) and one p53-independent (mitogens at cell cycle entry).
- Suggests a role for p21 in regulating cell cycle entry independent of p53 status.