Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2

Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2

Reportedly, antiapoptotic Bcl-2 family proteins suppress apoptosis by binding to and inhibiting members of the CED-4 family of caspase activators. To explore this question, we used embryonic stem (ES) cells in which one (−/+) or both (−/−) copies of the gene encoding apoptotic protease activating fa...

Full description

Bibliographic Details
Main Authors: Haraguchi, Misako, Torii, Seiji, Matsuzawa, Shu-ichi, Xie, Zhihua, Kitada, Shinichi, Krajewski, Stanislaw, Yoshida, Hiroki, Mak, Tak W., Reed, John C.
Format: Online
Language:English
Published: The Rockefeller University Press 2000
Online Access:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2193150/
id pubmed-2193150
recordtype oai_dc
spelling pubmed-21931502008-04-16 Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2 Haraguchi, Misako Torii, Seiji Matsuzawa, Shu-ichi Xie, Zhihua Kitada, Shinichi Krajewski, Stanislaw Yoshida, Hiroki Mak, Tak W. Reed, John C. Original Article Reportedly, antiapoptotic Bcl-2 family proteins suppress apoptosis by binding to and inhibiting members of the CED-4 family of caspase activators. To explore this question, we used embryonic stem (ES) cells in which one (−/+) or both (−/−) copies of the gene encoding apoptotic protease activating factor 1 (Apaf-1), a CED-4 homologue, were disrupted by homologous recombination. Stable clones of heterozygous (−/+) and homozygous (−/−) Apaf-1 knockout ES cells that overexpressed Bcl-2 were generated. Withdrawal of serum growth factors or stimulation of heterozygous ES cells with staurosporine (STS), ultraviolet (UV)B irradiation, etoposide (VP16), or cisplatin induced apoptosis followed by cell death (determined by failure to exclude propidium iodide dye). These cell death stimuli also induced activation of several types of caspases and loss of mitochondrial membrane potential (ΔΨ) in heterozygous (+/−) Apaf-1 knockout ES cells. In addition, overexpression of Bcl-2 protected against these events in Apaf-1–expressing ES cells. In contrast, STS, UVB, and VP16 induced little or no caspase activation and apoptosis in homozygous (−/−) Apaf-1 knockout ES cells. Nevertheless, Apaf-1–deficient ES cells subjected to these cell death stimuli or deprived of growth factors did eventually die through a nonapoptotic mechanism associated with loss of ΔΨ. Moreover, Bcl-2 overprotection preserved ΔΨ, reduced the percentage of Apaf-1−/− ES cells undergoing cell death, and increased clonigenic survival. The extent of Bcl-2–mediated cytoprotection was not significantly different for heterozygous (−/+) versus homozygous (−/−) Apaf-1 knockout cells. Furthermore, although Bcl-2 could be readily coimmunoprecipitated with Bax, associations with Apaf-1 were undetectable under conditions where Apaf-1 interactions with procaspase-9 were observed. We conclude that Bcl-2 has cytoprotective functions independent of Apaf-1, preserving mitochondrial function through a caspase-independent mechanism. The Rockefeller University Press 2000-05-15 /pmc/articles/PMC2193150/ /pubmed/10811864 Text en © 2000 The Rockefeller University Press This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
repository_type Open Access Journal
institution_category Foreign Institution
institution US National Center for Biotechnology Information
building NCBI PubMed
collection Online Access
language English
format Online
author Haraguchi, Misako
Torii, Seiji
Matsuzawa, Shu-ichi
Xie, Zhihua
Kitada, Shinichi
Krajewski, Stanislaw
Yoshida, Hiroki
Mak, Tak W.
Reed, John C.
spellingShingle Haraguchi, Misako
Torii, Seiji
Matsuzawa, Shu-ichi
Xie, Zhihua
Kitada, Shinichi
Krajewski, Stanislaw
Yoshida, Hiroki
Mak, Tak W.
Reed, John C.
Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2
author_facet Haraguchi, Misako
Torii, Seiji
Matsuzawa, Shu-ichi
Xie, Zhihua
Kitada, Shinichi
Krajewski, Stanislaw
Yoshida, Hiroki
Mak, Tak W.
Reed, John C.
author_sort Haraguchi, Misako
title Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2
title_short Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2
title_full Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2
title_fullStr Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2
title_full_unstemmed Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2
title_sort apoptotic protease activating factor 1 (apaf-1)–independent cell death suppression by bcl-2
description Reportedly, antiapoptotic Bcl-2 family proteins suppress apoptosis by binding to and inhibiting members of the CED-4 family of caspase activators. To explore this question, we used embryonic stem (ES) cells in which one (−/+) or both (−/−) copies of the gene encoding apoptotic protease activating factor 1 (Apaf-1), a CED-4 homologue, were disrupted by homologous recombination. Stable clones of heterozygous (−/+) and homozygous (−/−) Apaf-1 knockout ES cells that overexpressed Bcl-2 were generated. Withdrawal of serum growth factors or stimulation of heterozygous ES cells with staurosporine (STS), ultraviolet (UV)B irradiation, etoposide (VP16), or cisplatin induced apoptosis followed by cell death (determined by failure to exclude propidium iodide dye). These cell death stimuli also induced activation of several types of caspases and loss of mitochondrial membrane potential (ΔΨ) in heterozygous (+/−) Apaf-1 knockout ES cells. In addition, overexpression of Bcl-2 protected against these events in Apaf-1–expressing ES cells. In contrast, STS, UVB, and VP16 induced little or no caspase activation and apoptosis in homozygous (−/−) Apaf-1 knockout ES cells. Nevertheless, Apaf-1–deficient ES cells subjected to these cell death stimuli or deprived of growth factors did eventually die through a nonapoptotic mechanism associated with loss of ΔΨ. Moreover, Bcl-2 overprotection preserved ΔΨ, reduced the percentage of Apaf-1−/− ES cells undergoing cell death, and increased clonigenic survival. The extent of Bcl-2–mediated cytoprotection was not significantly different for heterozygous (−/+) versus homozygous (−/−) Apaf-1 knockout cells. Furthermore, although Bcl-2 could be readily coimmunoprecipitated with Bax, associations with Apaf-1 were undetectable under conditions where Apaf-1 interactions with procaspase-9 were observed. We conclude that Bcl-2 has cytoprotective functions independent of Apaf-1, preserving mitochondrial function through a caspase-independent mechanism.
publisher The Rockefeller University Press
publishDate 2000
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2193150/
_version_ 1611430919459045376

Similar Items