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Restrictin-P/stromal activin A, kills its target cells via an apoptotic mechanism
D Sternberg1, J Honigwachs-sha'anani, N Brosh
1Department of Cell Biology, The Weizmann Institute of Science, Rehovot, Israel.
Abstract:
We have recently found that the inhibitor of plasmacytoma cell growth, restrictin-P, is a stroma derived activin A and that it is an antagonist of interleukin-6 and interleukin-11. The present study was aimed at determining the mode by which this cytokine kills its target cells. On addition of the cytokine there was little or no net increase in cell number, depending on the specific target cells. All plasmacytoma cell lines tested exhibited a similar time dependent inhibition of DNA synthesis and a G0/G1 shift in the cell cycle. Electron microscope examination revealed classical apoptotic features i.e. chromatin condensation and membrane blebbing. DNA fragmentation, measured qualitatively and quantitatively, occurred in all cytokine treated plasmacytoma cell lines. Bovine activin A had an identical capacity to reduce cell viability, to induce G0/G1 shift and to cause DNA fragmentation. X-ray microanalysis of intracellular ions revealed an increase in calcium ions, following exposure of plasmacytoma cells to restrictin-P, accompanied by a decrease in phosphor ions. The cytotoxicity of the inhibitor was augmented in an additive manner by cycloheximide (CHX) indicating that the process did not require de novo protein synthesis. This study thus shows that restrictin-P/stromal activin A kills its target cells by inducing apoptosis. This effect was mediated by subnanogram concentrations and therefore may represent one physiological function of this pleiotropic cytokine.
Insights
Restrictin-P, a stromal activin A, inhibits plasmacytoma cell growth by inducing apoptosis. This cytokine kills target cells via DNA fragmentation and cell cycle arrest, independent of new protein synthesis.
Area of Science:
- Cell Biology
- Immunology
- Molecular Biology
Background:
- Restrictin-P, identified as stromal activin A, antagonizes interleukin-6 and interleukin-11.
- Its role in plasmacytoma cell growth inhibition necessitates understanding its mechanism of action.
Purpose of the Study:
- To elucidate the mechanism by which restrictin-P induces target cell death.
- To investigate the cellular and molecular events following restrictin-P treatment.
Main Methods:
- Cell viability assays and cell cycle analysis (G0/G1 shift).
- Electron microscopy to identify apoptotic features (chromatin condensation, membrane blebbing).
- DNA fragmentation assays, X-ray microanalysis of intracellular ions, and cycloheximide treatment.
Main Results:
- Restrictin-P treatment resulted in minimal cell proliferation and a G0/G1 cell cycle arrest.
- Apoptotic features, including DNA fragmentation, were observed in treated plasmacytoma cells.
- Intracellular calcium increased, while phosphate decreased; cytotoxicity was enhanced by cycloheximide, indicating no de novo protein synthesis.
Conclusions:
- Restrictin-P/stromal activin A induces apoptosis in target cells.
- The observed effects occur at subnanogram concentrations, suggesting a physiological role for this cytokine.