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Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes

Bax induces mitochondrial outer membrane permeabilization (MOMP), a critical step in apoptosis in which proteins are released into the cytoplasm. To resolve aspects of the mechanism, we used cryo-electron microscopy (cryo-EM) to visualize Bax-induced pores in purified mitochondrial outer membranes (...

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Autores principales: Gillies, Laura A, Du, Han, Peters, Bjoern, Knudson, C. Michael, Newmeyer, Donald D., Kuwana, Tomomi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Cell Biology 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4294680/
https://www.ncbi.nlm.nih.gov/pubmed/25411335
http://dx.doi.org/10.1091/mbc.E13-11-0638
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author Gillies, Laura A
Du, Han
Peters, Bjoern
Knudson, C. Michael
Newmeyer, Donald D.
Kuwana, Tomomi
author_facet Gillies, Laura A
Du, Han
Peters, Bjoern
Knudson, C. Michael
Newmeyer, Donald D.
Kuwana, Tomomi
author_sort Gillies, Laura A
collection PubMed
description Bax induces mitochondrial outer membrane permeabilization (MOMP), a critical step in apoptosis in which proteins are released into the cytoplasm. To resolve aspects of the mechanism, we used cryo-electron microscopy (cryo-EM) to visualize Bax-induced pores in purified mitochondrial outer membranes (MOMs). We observed solitary pores that exhibited negative curvature at their edges. Over time, the pores grew to ∼100–160 nm in diameter after 60–90 min, with some pores measuring more than 300 nm. We confirmed these results using flow cytometry, which we used to monitor the release of fluorescent dextrans from isolated MOM vesicles. The dextran molecules were released gradually, in a manner constrained by pore size. However, the release rates were consistent over a range of dextran sizes (10–500 kDa). We concluded that the pores were not static but widened dramatically to release molecules of different sizes. Taken together, the data from cryo-EM and flow cytometry argue that Bax promotes MOMP by inducing the formation of large, growing pores through a mechanism involving membrane-curvature stress.
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spelling pubmed-42946802015-03-30 Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes Gillies, Laura A Du, Han Peters, Bjoern Knudson, C. Michael Newmeyer, Donald D. Kuwana, Tomomi Mol Biol Cell Articles Bax induces mitochondrial outer membrane permeabilization (MOMP), a critical step in apoptosis in which proteins are released into the cytoplasm. To resolve aspects of the mechanism, we used cryo-electron microscopy (cryo-EM) to visualize Bax-induced pores in purified mitochondrial outer membranes (MOMs). We observed solitary pores that exhibited negative curvature at their edges. Over time, the pores grew to ∼100–160 nm in diameter after 60–90 min, with some pores measuring more than 300 nm. We confirmed these results using flow cytometry, which we used to monitor the release of fluorescent dextrans from isolated MOM vesicles. The dextran molecules were released gradually, in a manner constrained by pore size. However, the release rates were consistent over a range of dextran sizes (10–500 kDa). We concluded that the pores were not static but widened dramatically to release molecules of different sizes. Taken together, the data from cryo-EM and flow cytometry argue that Bax promotes MOMP by inducing the formation of large, growing pores through a mechanism involving membrane-curvature stress. The American Society for Cell Biology 2015-01-15 /pmc/articles/PMC4294680/ /pubmed/25411335 http://dx.doi.org/10.1091/mbc.E13-11-0638 Text en © 2015 Gillies et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology.
spellingShingle Articles
Gillies, Laura A
Du, Han
Peters, Bjoern
Knudson, C. Michael
Newmeyer, Donald D.
Kuwana, Tomomi
Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes
title Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes
title_full Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes
title_fullStr Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes
title_full_unstemmed Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes
title_short Visual and functional demonstration of growing Bax-induced pores in mitochondrial outer membranes
title_sort visual and functional demonstration of growing bax-induced pores in mitochondrial outer membranes
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4294680/
https://www.ncbi.nlm.nih.gov/pubmed/25411335
http://dx.doi.org/10.1091/mbc.E13-11-0638
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