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Bcl-xL overexpression inhibits taxol-induced Yama protease activity and apoptosis
1Department of Medicine, Winship Cancer Center, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Abstract:
Intracellularly, the anticancer drug taxol induces tubulin polymerization and mitotic arrest, followed by apoptosis. The DNA repair enzyme poly(ADP-ribose) polymerase (PARP) and lamins are known to be degraded during apoptosis. PARP is a substrate for the Yama protease, which is encoded by the CPP32 beta/ Yama gene, whereas lamins are degraded by the Yama and lamin proteases. In the present studies, we determined the effects of enforced overexpression of the antiapoptosis Bcl-xL protein on taxol-mediated microtubule and cell cycle perturbations, as well as on taxol-induced apoptosis and associated Yama protease activity in human myeloid leukemia HL-60 cells. Our data demonstrate that high Bcl-xL levels do not affect the microtubular bundling or mitotic arrest due to taxol but significantly inhibit the morphological, flow cytometric, and DNA fragmentation features associated with taxol-induced apoptosis. This resulted in a significant improvement in the survival of taxol-treated cells that possess high Bcl-xL levels. In the control HL-60 cells, following taxol treatment, whereas the mRNA of Yama was not induced, taxol-induced apoptosis was associated with Yama activation and PARP as well as lamin B1 degradation. These features were blocked by coculture of these cells with the cysteine protease inhibitor YVAD-cmk as well as in cells with overexpression of Bcl-xL. These results suggest that Bcl-xL antagonizes taxol-induced apoptosis by a mechanism that interferes with the activation of a key protease involved in the execution of apoptosis.
Insights
The anti-apoptosis protein Bcl-xL inhibits taxol-induced apoptosis in leukemia cells by blocking key protease activation. This improves cell survival, offering potential therapeutic insights for cancer treatment.
Area of Science:
- Cell Biology
- Molecular Biology
- Pharmacology
Background:
- Taxol (paclitaxel) is an anticancer drug that induces apoptosis by arresting cells in mitosis.
- Apoptosis involves the degradation of proteins like poly(ADP-ribose) polymerase (PARP) and lamins by specific proteases.
- The Yama protease, encoded by CPP32 beta/Yama, is crucial for cleaving PARP and lamins during apoptosis.
Purpose of the Study:
- To investigate the effect of overexpressing the anti-apoptotic protein Bcl-xL on taxol-induced apoptosis in human myeloid leukemia HL-60 cells.
- To determine if Bcl-xL influences taxol-mediated microtubule and cell cycle effects.
- To elucidate the mechanism by which Bcl-xL modulates taxol-induced apoptosis and protease activity.
Main Methods:
- Overexpression of Bcl-xL in HL-60 leukemia cells.
- Treatment of cells with taxol.
- Analysis of microtubule polymerization and cell cycle progression using flow cytometry.
- Assessment of apoptosis-associated DNA fragmentation and morphological changes.
- Measurement of Yama protease activity and degradation of PARP and lamin B1.
- Inhibition studies using the cysteine protease inhibitor YVAD-cmk.
Main Results:
- High levels of Bcl-xL did not alter taxol's effects on microtubule bundling or mitotic arrest.
- Bcl-xL significantly inhibited taxol-induced apoptosis, evidenced by reduced DNA fragmentation and altered cell morphology.
- Overexpression of Bcl-xL led to improved survival rates in taxol-treated cells.
- Taxol-induced Yama protease activation and subsequent degradation of PARP and lamin B1 were blocked by Bcl-xL overexpression or YVAD-cmk treatment.
- Yama mRNA levels were not induced by taxol, indicating post-translational regulation of protease activity.
Conclusions:
- Bcl-xL antagonizes taxol-induced apoptosis by interfering with the activation of key executioner proteases, such as Yama.
- The anti-apoptotic function of Bcl-xL is independent of its effects on microtubule dynamics or cell cycle arrest.
- These findings suggest that targeting Bcl-xL or related pathways could enhance the efficacy of taxol chemotherapy.