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p53-independent elevation of p21 expression by PMA results from PKC-mediated mRNA stabilization
1Department of Immunology, School of Medicine, Keimyung University, 194 DongSan-Dong, Jung-Gu, Taegu, 700-712, South Korea.
Abstract:
The p21 (cip1/waf1) protein induces cell cycle arrest through inhibition of the activity of cdk (cyclin dependent kinase)/cyclin complexes. Expression of p21 is induced in a p53-dependent manner by DNA damage. p21 can also be induced independently of p53 by phorbol ester or okadaic acid. In this study, we have addressed the role of the PKC (protein kinase C) signaling pathway in the induction of p21 in response to PMA (phorbol myristate acetate) and okadaic acid. Levels of p21 (protein and mRNA) rapidly increased (within approximately 4 h) in U937 cells treated with PMA. The PKC-specific inhibitors RO 31-8220 and GF109203X down-regulated PMA or okadaic acid-induced p21 expression. Following persistent PKC activation, p21 mRNA levels remained elevated, indicating an enhanced stability of the mRNA. Using actinomycin D to measure mRNA stability and p21 promoter luciferase assays to measure activity, we provide evidence to support a role for the PKC signaling pathway in p21 mRNA stability. Thus, PKC regulates the amount of p21 in U937 cells at the level of mRNA accumulation and translation.
Insights
Protein kinase C (PKC) signaling regulates p21 protein levels by increasing mRNA stability and translation. This pathway influences cell cycle arrest independent of p53, impacting cancer research.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- The p21 protein (cip1/waf1) is a key regulator of cell cycle arrest, inhibiting cyclin-dependent kinase/cyclin complexes.
- p21 expression is typically induced by DNA damage in a p53-dependent manner, but can also be triggered by other stimuli like phorbol ester and okadaic acid.
Purpose of the Study:
- To investigate the role of the protein kinase C (PKC) signaling pathway in the induction of p21 expression.
- To elucidate the mechanisms by which PKC influences p21 levels in response to phorbol myristate acetate (PMA) and okadaic acid.
Main Methods:
- Treatment of U937 cells with PMA and okadaic acid.
- Assessment of p21 protein and mRNA levels.
- Utilized PKC-specific inhibitors (RO 31-8220, GF109203X).
- Measured mRNA stability using actinomycin D.
- Assessed p21 promoter activity via luciferase assays.
Main Results:
- PMA treatment rapidly increased p21 protein and mRNA levels in U937 cells within approximately 4 hours.
- PKC inhibitors significantly reduced PMA or okadaic acid-induced p21 expression.
- Persistent PKC activation led to elevated p21 mRNA levels, suggesting enhanced mRNA stability.
- Evidence supports PKC signaling's role in p21 mRNA stability and promoter activity.
Conclusions:
- The protein kinase C (PKC) signaling pathway plays a crucial role in regulating p21 expression.
- PKC influences p21 levels through mechanisms involving both mRNA accumulation (stability) and translation.
- This regulation occurs independently of p53, highlighting an alternative pathway for cell cycle control.