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Activin A induction of cell-cycle arrest involves modulation of cyclin D2 and p21CIP1/WAF1 in plasmacytic cells
K Yamato1, T Koseki, M Ohguchi
1Department of Oral Science, The National Institute of Infectious Diseases, Tokyo, Japan.
Abstract:
Activins, members of the transforming growth factor-beta family, have been implicated in the regulation of growth and differentiation of various types of cells. We have recently found that activin A induces apoptotic cell death of plasmacytic cells including B cell hybridoma cells and myeloma cells. In the present study, we demonstrated that activin A caused cell-cycle arrest in the G1 phase before appearance of apoptotic cells in mouse B cell hybridoma cells. Phosphorylation of retinoblastoma protein (Rb) and in vitro Rb kinase activity of cyclin-dependent kinase (CDK)4 was inhibited in activin A-treated cells. Analysis of expression of genes regulating Rb phosphorylation revealed that activin A suppressed cyclin D2, the sole D-type cyclin gene expressed in the hybridoma cells, and activated p21CIP1/WAF1 but had no effect on expression of cyclin-dependent kinases (CDK2, CDK4, CDK6) and other CDK inhibitors (p27KIP1, p16INK4a, p15INK4b). Modulation of cyclin D2 and p21CIP1/WAF1 expression resulted in a decrease in level of cyclin D2-CDK4 complex and an increase in level of CDK4 complexed with p21CIP1/WAF1. Moreover, overexpression of cyclin D2 partially abrogated inhibition of Rb phosphorylation and G1 arrest in the hybridoma cells.
Insights
Activin A induces G1 cell-cycle arrest in B cell hybridoma cells by inhibiting retinoblastoma protein phosphorylation. This occurs through decreased cyclin D2 and increased p21CIP1/WAF1 expression, preceding apoptosis.
Area of Science:
- Cell Biology
- Molecular Biology
- Cancer Research
Background:
- Activins, part of the transforming growth factor-beta superfamily, regulate cell growth and differentiation.
- Activin A has been shown to induce apoptosis in plasmacytic cells, including B cell hybridoma and myeloma cells.
Purpose of the Study:
- To investigate the mechanism by which activin A induces cell death in B cell hybridoma cells.
- To determine the effects of activin A on cell-cycle progression and key regulatory proteins.
Main Methods:
- Treatment of mouse B cell hybridoma cells with activin A.
- Cell-cycle analysis (flow cytometry).
- Western blotting to assess protein levels and phosphorylation (Rb, cyclins, CDKs, CDK inhibitors).
- Analysis of gene expression (RT-PCR).
- Overexpression studies of cyclin D2.
Main Results:
- Activin A induced G1 cell-cycle arrest in hybridoma cells prior to apoptosis.
- Activin A inhibited retinoblastoma protein (Rb) phosphorylation and CDK4 kinase activity.
- Activin A suppressed cyclin D2 expression and increased p21CIP1/WAF1 expression.
- These changes led to reduced cyclin D2-CDK4 complex formation and increased p21CIP1/WAF1-CDK4 complex formation.
- Overexpression of cyclin D2 partially reversed the G1 arrest and Rb phosphorylation inhibition.
Conclusions:
- Activin A-mediated G1 arrest in B cell hybridoma cells is dependent on the modulation of cyclin D2 and p21CIP1/WAF1.
- The findings elucidate a novel mechanism of activin A action on cell-cycle regulation in B cells.
- This provides insights into potential therapeutic strategies targeting B cell malignancies.