Movement of Bax from the Cytosol to Mitochondria during Apoptosis
Movement of Bax from the Cytosol to Mitochondria during Apoptosis
Wolter, Keith G.; Hsu, Yi-Te; Smith, Carolyn L.; Nechushtan, Amotz; Xi, Xu-Guang; Youle, Richard J.
1997-12-01 00:00:00
Bax, a member of the Bcl-2 protein family, accelerates apoptosis by an unknown mechanism. Bax has been recently reported to be an integral membrane protein associated with organelles or bound to organelles by Bcl-2 or a soluble protein found in the cytosol. To explore Bcl-2 family member localization in living cells, the green fluorescent protein (GFP) was fused to the NH 2 termini of Bax, Bcl-2, and Bcl-X L . Confocal microscopy performed on living Cos-7 kidney epithelial cells and L929 fibroblasts revealed that GFP–Bcl-2 and GFP–Bcl-X L had a punctate distribution and colocalized with a mitochondrial marker, whereas GFP–Bax was found diffusely throughout the cytosol. Photobleaching analysis confirmed that GFP–Bax is a soluble protein, in contrast to organelle-bound GFP–Bcl-2. The diffuse localization of GFP–Bax did not change with coexpression of high levels of Bcl-2 or Bcl-X L . However, upon induction of apoptosis, GFP–Bax moved intracellularly to a punctate distribution that partially colocalized with mitochondria. Once initiated, this Bax movement was complete within 30 min, before cellular shrinkage or nuclear condensation. Removal of a COOH-terminal hydrophobic domain from GFP–Bax inhibited redistribution during apoptosis and inhibited the death-promoting activity of both Bax and GFP– Bax. These results demonstrate that in cells undergoing apoptosis, an early, dramatic change occurs in the intracellular localization of Bax, and this redistribution of soluble Bax to organelles appears important for Bax to promote cell death. Footnotes K.G. Wolter was supported by the Howard Hughes Medical Institute-National Institutes of Health Research Scholars Program. Address all correspondence to Richard J. Youle, Bldg 10, Room 5D-37, National Institutes of Health, Bethesda, MD 20892. Tel.: (301) 496-6628. Fax: (301) 402-0380. E-mail: youle@helix.nih.gov Abbreviations used in this paper: ΔCT COOH-terminal truncation DIC differential interference contrast GFP green fluorescent protein STS staurosporine Submitted: 15 July 1997 Revision received 12 September 1997
http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.pngThe Journal of Cell BiologyRockefeller University Presshttp://www.deepdyve.com/lp/rockefeller-university-press/movement-of-bax-from-the-cytosol-to-mitochondria-during-apoptosis-cfBtapx3zp
Movement of Bax from the Cytosol to Mitochondria during Apoptosis
Bax, a member of the Bcl-2 protein family, accelerates apoptosis by an unknown mechanism. Bax has been recently reported to be an integral membrane protein associated with organelles or bound to organelles by Bcl-2 or a soluble protein found in the cytosol. To explore Bcl-2 family member localization in living cells, the green fluorescent protein (GFP) was fused to the NH 2 termini of Bax, Bcl-2, and Bcl-X L . Confocal microscopy performed on living Cos-7 kidney epithelial cells and L929 fibroblasts revealed that GFP–Bcl-2 and GFP–Bcl-X L had a punctate distribution and colocalized with a mitochondrial marker, whereas GFP–Bax was found diffusely throughout the cytosol. Photobleaching analysis confirmed that GFP–Bax is a soluble protein, in contrast to organelle-bound GFP–Bcl-2. The diffuse localization of GFP–Bax did not change with coexpression of high levels of Bcl-2 or Bcl-X L . However, upon induction of apoptosis, GFP–Bax moved intracellularly to a punctate distribution that partially colocalized with mitochondria. Once initiated, this Bax movement was complete within 30 min, before cellular shrinkage or nuclear condensation. Removal of a COOH-terminal hydrophobic domain from GFP–Bax inhibited redistribution during apoptosis and inhibited the death-promoting activity of both Bax and GFP– Bax. These results demonstrate that in cells undergoing apoptosis, an early, dramatic change occurs in the intracellular localization of Bax, and this redistribution of soluble Bax to organelles appears important for Bax to promote cell death. Footnotes K.G. Wolter was supported by the Howard Hughes Medical Institute-National Institutes of Health Research Scholars Program. Address all correspondence to Richard J. Youle, Bldg 10, Room 5D-37, National Institutes of Health, Bethesda, MD 20892. Tel.: (301) 496-6628. Fax: (301) 402-0380. E-mail: youle@helix.nih.gov Abbreviations used in this paper: ΔCT COOH-terminal truncation DIC differential interference contrast GFP green fluorescent protein STS staurosporine Submitted: 15 July 1997 Revision received 12 September 1997
Journal
The Journal of Cell Biology
– Rockefeller University Press
Published: Dec 1, 1997
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