Pro-apoptotic apoptosis protease-activating factor 1 (Apaf-1) has a cytoplasmic localization distinct from Bcl-2 or Bcl-x(L)

How Bcl-2 and its pro-survival relatives prevent activation of the caspases that mediate apoptosis is unknown, but they appear to act through the caspase activator apoptosis protease-activating factor 1 (Apaf-1). According to the apoptosome model, the Bcl-2-like proteins preclude Apaf-1 activity by...

Full description

Saved in:
Bibliographic Details
Published in:The Journal of cell biology 2000-05, Vol.149 (3), p.623-634
Main Authors: Hausmann, G, O'Reilly, L A, van Driel, R, Beaumont, J G, Strasser, A, Adams, J M, Huang, D C
Format: Article
Language:English
Subjects:
Animals
Antibodies, Monoclonal - immunology
Apoptosis
Apoptotic Protease-Activating Factor 1
bcl-X Protein
Caspases - metabolism
Cell Line
Cytochrome c Group - pharmacology
Cytoplasm - metabolism
Cytoplasm - ultrastructure
Enzyme Activation
Fluorescent Antibody Technique
Humans
Microscopy, Confocal
Microscopy, Immunoelectron
Proteins - immunology
Proteins - metabolism
Proto-Oncogene Proteins c-bcl-2 - metabolism
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:How Bcl-2 and its pro-survival relatives prevent activation of the caspases that mediate apoptosis is unknown, but they appear to act through the caspase activator apoptosis protease-activating factor 1 (Apaf-1). According to the apoptosome model, the Bcl-2-like proteins preclude Apaf-1 activity by sequestering the protein. To explore Apaf-1 function and to test this model, we generated monoclonal antibodies to Apaf-1 and used them to determine its localization within diverse cells by subcellular fractionation and confocal laser scanning microscopy. Whereas Bcl-2 and Bcl-x(L) were prominent on organelle membranes, endogenous Apaf-1 was cytosolic and did not colocalize with them, even when these pro-survival proteins were overexpressed or after apoptosis was induced. Immunogold electron microscopy confirmed that Apaf-1 was dispersed in the cytoplasm and not on mitochondria or other organelles. After the death stimuli, Bcl-2 and Bcl-x(L) precluded the release of the Apaf-1 cofactor cytochrome c from mitochondria and the formation of larger Apaf-1 complexes, which are steps that presage apoptosis. However, neither Bcl-2 nor Bcl-x(L) could prevent the in vitro activation of Apaf-1 induced by the addition of exogenous cytochrome c. Hence, rather than sequestering Apaf-1 as proposed by the apoptosome model, Bcl-2-like proteins probably regulate Apaf-1 indirectly by controlling upstream events critical for its activation.
ISSN:0021-9525
DOI:10.1083/jcb.149.3.623