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p57Kip2, a glucocorticoid-induced inhibitor of cell cycle progression in HeLa cells
M K Samuelsson1, A Pazirandeh, B Davani
1Department of Medical Nutrition, Karolinska Institute, Huddinge University Hospital, Sweden.
Abstract:
Glucocorticoids exert antiproliferative effects on a number of cell types, including the HeLa cervical carcinoma cell line. However, the mechanism responsible for the antiproliferative effect is poorly understood. In this report we have investigated the role of the recently identified cyclin-dependent kinase inhibitor (CDI) p57Kip2 in the antiproliferative effect conferred by glucocorticoids. When HeLa cells were treated with the synthetic glucocorticoid dexamethasone (DEX), the doubling time of exponentially growing cells increased 2-fold. Within 11 h of DEX treatment, this was accompanied by an accumulation of cells in the G1 phase of the cell cycle with a corresponding decreased proportion of cells in the S phase and decreased CDK2 activity. DEX treatment of the HeLa cells dramatically induced the protein and mRNA expression of the CDI p57Kip2. This induction was seen within 4 h of DEX treatment, preceding a major DEX-induced accumulation of cells in the G1 phase. DEX-induced mRNA expression of p57Kip2 did not require de novo protein synthesis, and the transcription of the p57Kip2 gene was increased as determined by a run-on transcription assay. Furthermore, DEX induction of p57Kip2 was not a consequence of the cell cycle arrest, since other growth inhibition signals did not result in strong p57Kip2 induction. Overexpression of p57Kip2 using HeLa cells stably transfected with a tetracycline-inducible vector showed that p57Kip2 is sufficient to reconstitute an antiproliferative effect similar to that seen in DEX-treated cells. Selective p57Kip2 expression by the tetracycline analog doxycycline to levels comparable to those observed on DEX induction resulted in a 1.7-fold increase in the doubling time and a shift of HeLa cells to the G1 phase as well as a decrease in CDK2 activity. Taken together, these results suggest that glucocorticoid treatment directly induces transcription of the p57Kip2 gene and that the p57Kip2 protein is involved in the glucocorticoid-induced antiproliferative effect.
Insights
Glucocorticoids like dexamethasone inhibit HeLa cell proliferation by inducing the cyclin-dependent kinase inhibitor p57Kip2. This protein is sufficient to cause cell cycle arrest in the G1 phase, revealing a key mechanism in glucocorticoid action.
Area of Science:
- Molecular Biology
- Cell Biology
- Cancer Research
Background:
- Glucocorticoids are known to inhibit proliferation in various cell types, including HeLa cervical carcinoma cells.
- The precise molecular mechanisms underlying this antiproliferative effect remain incompletely understood.
- The cyclin-dependent kinase inhibitor (CDI) p57Kip2 has been recently identified.
Purpose of the Study:
- To investigate the role of p57Kip2 in mediating the antiproliferative effects of glucocorticoids.
- To elucidate the molecular pathway linking glucocorticoid treatment to cell cycle arrest.
Main Methods:
- HeLa cells were treated with dexamethasone (DEX), a synthetic glucocorticoid.
- Cell cycle progression, CDK2 activity, and p57Kip2 expression (mRNA and protein) were analyzed.
- Gene transcription was assessed using run-on assays, and p57Kip2 overexpression was achieved via a tetracycline-inducible system.
Main Results:
- DEX treatment increased HeLa cell doubling time and induced G1 phase accumulation, decreasing S phase proportion and CDK2 activity.
- DEX significantly upregulated both p57Kip2 mRNA and protein expression, preceding G1 arrest.
- p57Kip2 induction by DEX was transcription-dependent and not a secondary effect of cell cycle arrest; overexpression of p57Kip2 mimicked DEX's antiproliferative effects.
Conclusions:
- Glucocorticoid treatment directly induces p57Kip2 gene transcription in HeLa cells.
- The induced p57Kip2 protein plays a crucial role in mediating the antiproliferative and cell cycle arrest effects of glucocorticoids.