Imaging Studies Reveal Order in Programmed Cell Death
ScienceDaily (Mar. 3, 2010) — Every day, about 10 billion cells in a human body commit suicide. Cells infected by virus, that are transformed or otherwise dysfunctional altruistically sacrifice themselves for the greater good. Now, new imaging experiments have revealed a previously unseen order to this process, showing closely related cells dying in synchrony as a wave of destruction sweeps across their mitochondria, snuffing out the main source of energy that keeps cells alive.
Death wave. New imaging research finds order in what was thought to be the random timing or sudden collapse in apoptosis. A mitochondrial protein, cytochrome-c, is pictured dissipating in an orderly wave around the nucleus (black center) in a cascade that ends in cell death.
In experiments published recently inThe Journal of Cell Science and Biophysical Journal, researchers inSanford M. Simon's Laboratory of Cellular Biophysics at Rockefeller University photographed the deaths of individual cells, showing an orderly series of events in the staged shut-down of the cell. The experiments revealed that the likelihood of death, as well as the timing, depends on how closely cells are related, not on their proximity to one another or their stage in the cell cycle. The findings rule out, for instance, the hypothesis that cells die in a localized cascade accelerated by the secretion of toxic molecules from dying cells nearby.
"What we saw is that, regardless of their location, only the sister cells remained linked in the timing of their deaths," says Simon. "It suggests that there is not some nonspecific toxic effect here, but that the variability is in the molecular makeup of the cells -- the variability in the population."
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Volume 97, Issue 8, 21 October 2009, Pages 2222-2231
Spatial and Temporal Dynamics of Mitochondrial Membrane Permeability Waves during Apoptosis
Patrick D. Bholaa, Alexa L. Mattheysesa and Sanford M. Simon, a,
aDepartment of Cellular Biophysics, Rockefeller University, New York, New York
aDepartment of Cellular Biophysics, Rockefeller University, New York, New York
Received 11 May 2009;
accepted 7 July 2009.
Editor: Joshua Zimmerberg..
Available online 19 October 2009.
Abstract
Change in the permeability of the mitochondrial membrane to proteins (cytochrome c and Smac) and protons is a critical step in apoptosis. Although the time from the induction of apoptosis to the change of mitochondrial permeability is variable over a period of hours, the release of proteins is an “all or none” phenomenon that is completed in an individual cell within minutes. Here, using single-cell fluorescence microscopy, we show that the release of cytochrome c from a single mitochondrion occurs in a single step. However, this increased permeability of the outer membrane to cytochrome c propagates throughout the cell as a slower, spatially coordinated wave. The permeability of the outer membrane to Smac propagates with the same spatial pattern but lagging in time. This is followed by a wave of increased permeability of the inner membrane to protons. Only afterward do the mitochondria fission. The spatial dependence of the permeability wave was inhibited by thapsigargin, an inhibitor of the endoplasmic reticulum calcium pumps, but buffering cytosolic calcium had no effect. These results show that the trigger for apoptosis is spatially localized, initiating at one or only a few mitochondria preceding the loss of mitochondrial energetics, and the subsequent temporal propagation of mitochondrial membrane permeability is calcium-dependent.
accepted 7 July 2009.
Editor: Joshua Zimmerberg..
Available online 19 October 2009.
Abstract
Change in the permeability of the mitochondrial membrane to proteins (cytochrome c and Smac) and protons is a critical step in apoptosis. Although the time from the induction of apoptosis to the change of mitochondrial permeability is variable over a period of hours, the release of proteins is an “all or none” phenomenon that is completed in an individual cell within minutes. Here, using single-cell fluorescence microscopy, we show that the release of cytochrome c from a single mitochondrion occurs in a single step. However, this increased permeability of the outer membrane to cytochrome c propagates throughout the cell as a slower, spatially coordinated wave. The permeability of the outer membrane to Smac propagates with the same spatial pattern but lagging in time. This is followed by a wave of increased permeability of the inner membrane to protons. Only afterward do the mitochondria fission. The spatial dependence of the permeability wave was inhibited by thapsigargin, an inhibitor of the endoplasmic reticulum calcium pumps, but buffering cytosolic calcium had no effect. These results show that the trigger for apoptosis is spatially localized, initiating at one or only a few mitochondria preceding the loss of mitochondrial energetics, and the subsequent temporal propagation of mitochondrial membrane permeability is calcium-dependent.
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Journal of Cell Science 122, 4296-4302 (2009)
Published by The Company of Biologists 2009
Determinism and divergence of apoptosis susceptibility in mammalian cellsPatrick D. Bhola and Sanford M. Simon*
Department of Cellular Biophysics, The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA
* Author for correspondence (simon@rockefeller.edu)
Accepted 15 September 2009
Although the cellular decision to commit to apoptosis is important for organism homeostasis, there is considerable variability in the onset of apoptosis between cells, even in clonal populations. Using live single-cell imaging, we observed that the onset of apoptotic proteolytic activity was tightly synchronized between nearby cells. This synchrony was not a consequence of secreted factors and was not correlated to the cell cycle. The synchrony was only seen amongst related cells and was lost over successive generations. The times of apoptosis also diverged within a generation, but this was blocked by inhibiting protein synthesis before triggering apoptosis. These results suggest that the cell-cell variability of apoptosis times is due to the divergence of the molecular composition of the cell, and that the decision to commit to apoptosis at the time of drug addition is a deterministic decision.
Key words: Fluorescence microscopy, Protease, Caspase, Lineage, Epigenetics
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Department of Cellular Biophysics, The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA
* Author for correspondence (simon@rockefeller.edu)
Accepted 15 September 2009
Although the cellular decision to commit to apoptosis is important for organism homeostasis, there is considerable variability in the onset of apoptosis between cells, even in clonal populations. Using live single-cell imaging, we observed that the onset of apoptotic proteolytic activity was tightly synchronized between nearby cells. This synchrony was not a consequence of secreted factors and was not correlated to the cell cycle. The synchrony was only seen amongst related cells and was lost over successive generations. The times of apoptosis also diverged within a generation, but this was blocked by inhibiting protein synthesis before triggering apoptosis. These results suggest that the cell-cell variability of apoptosis times is due to the divergence of the molecular composition of the cell, and that the decision to commit to apoptosis at the time of drug addition is a deterministic decision.
Key words: Fluorescence microscopy, Protease, Caspase, Lineage, Epigenetics
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