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Bcl-xS and Bax induce different apoptotic pathways in PC12 cells

Oncogene volume 19, pages 1783–1793 (2000)Cite this article

Abstract

Apoptosis is regulated by the action of the Bcl-2 family of proteins, which includes anti- and pro-apoptotic members such as Bcl-xS and Bax. These proteins may differ from each other in structure, mechanism of action and interactions with anti-apoptotic signaling. The mechanism whereby Bax induces cell death has been studied in some cellular systems, but the mechanism of Bcl-xS-induced apoptosis is largely unknown. In this study we investigated and compared the apoptotic effects of Bcl-xS and Bax in the pheochromocytoma cell line, PC12 (a useful model system for studying neuronal apoptosis), and the extent to which they are protected by the survival factor, nerve growth factor (NGF). PC12 cells express endogenous Bcl-xS, Bax and Bcl-xL proteins. Subcellular fractionation revealed that Bax is presented mainly in the cytosolic and the heavy membrane fractions, Bcl-xS is present only in the cytosol, and the anti-apoptotic protein Bcl-xL is located mainly in the heavy membrane fraction. In contrast to the cytosolic localization of endogenous Bcl-xS, the exogenously overexpressed Bcl-xS is localized to the mitochondria. Overexpression of Bcl-xS or Bax induces cell death in the transfected cells. The cell death induced by overexpression of Bcl-xS was inhibited by co-expression of Bcl-xS with Bcl-2 or Bcl-xL, or by treatment with the broad-spectrum caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoro-methylketone (Z-VAD-FMK) or with NGF. The Bcl-2 mutants ΔC22, which lacks the transmembrane domain, and G145A (mI-3) were able to inhibit the death-inducing effect of Bcl-xS. These results therefore suggest that the apoptotic pathway induced by overexpression of Bcl-xS in PC12 cells can be controlled by Bcl-2 and Bcl-xL, is mediated by caspases, and can be inhibited by the NGF signaling pathway. The Bax-induced cell death was inhibited by co-expression of Bax with Bcl-2 or Bcl-xL, but was not inhibited by Z-VAD-FMK, NGF, or the Bcl-2 mI-3 or ΔC22 mutants. These results therefore suggest that Bax induces a caspase-independent cell death pathway which is blocked by Bcl-2 but not by the NGF signaling pathway. They further suggest that Bcl-xS and Bax induce different cell death pathways in PC12 cells.

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Acknowledgements

We thank Dr Craig Thompson for providing the Bcl-xL and Bcl-xS vectors, Dr Stanley Korsmeyer for providing the Bax, Bcl-2 and Bcl-2 mutant vectors and Dr Ron Kopito for providing anti-calnexin antibodies. We are also grateful to Ms Shirley Smith for excellent editorial assistance. This work was supported by the United States Israel–Binational Science Foundation.

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  1. Department of Neurobiochemistry, George S. Wise Faculty of Life Sciences, Tel Aviv University, 69978, Ramat Aviv, Israel

    Liora Lindenboim & Reuven Stein

  2. Department of Cell Biology, Harvard Medical School, Boston, 02115, Massachusetts, MA, USA

    Junying Yuan

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Lindenboim, L., Yuan, J. & Stein, R. Bcl-xS and Bax induce different apoptotic pathways in PC12 cells. Oncogene 19, 1783–1793 (2000). https://doi.org/10.1038/sj.onc.1203495

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