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Plasma membrane and nuclear envelope integrity during the blebbing stage of apoptosis: a time-lapse study
Ricardo Andrade1, Lorena Crisol, Roberto Prado
1Biomedical Analytical and High Resolution Microscopy Facility, University of the Basque Country, Leioa, Vizcaya, Spain.
Background Information:
The execution phase of apoptosis is characterized by extensive blebbing of the plasma membrane, which usually results in secondary lysis in vitro. To analyse the permeability of cellular membranes during this process, we induced apoptosis in human melanoma A375 cells that had been transfected with fluorescently tagged proteins which were targeted to different subcellular locations.
Results:
The dual treatment of resveratrol and butyrate produced a synergistic induction of apoptosis by blocking different phases of the cell cycle. Changes in the plasma membrane, nuclear envelope and nucleoli were monitored by time-lapse confocal microscopy. Fluorescently labelled proteins were not mis-localized from their original locations in any of the cells undergoing blebbing for several hours. Thus the maintenance of karyophilic and nucleolar proteins within the nucleus during the blebbing stage and the accessibility of vital selective chromatin dyes confirmed a functional preservation of the nuclear compartment until the final necrotic blister. The translocation of phosphatidylserine to the outer leaflet of the plasma membrane was not detected during the blebbing period.
Conclusion:
These results show that the functional integrity of the nuclear envelope and plasma membrane may be conserved until the end of the execution phase of apoptosis.
Insights
Apoptosis involves cell membrane blebbing, but this study shows the nucleus and plasma membrane maintain integrity. This functional preservation occurs throughout the late stages of programmed cell death.
Area of Science:
- Cell Biology
- Molecular Biology
Background:
- Apoptosis execution involves plasma membrane blebbing, often leading to secondary lysis in vitro.
- Investigating cellular membrane permeability during apoptosis is crucial for understanding cell death mechanisms.
Purpose of the Study:
- To analyze the permeability of cellular membranes during apoptosis.
- To investigate the functional integrity of the nuclear envelope and plasma membrane during the execution phase of apoptosis.
Main Methods:
- Induced apoptosis in human melanoma A375 cells using dual treatment of resveratrol and butyrate.
- Transfected cells with fluorescently tagged proteins targeted to various subcellular locations.
- Monitored membrane changes using time-lapse confocal microscopy.
Main Results:
- Resveratrol and butyrate synergistically induced apoptosis by blocking cell cycle phases.
- Fluorescently labeled proteins remained localized, indicating nuclear compartment integrity during blebbing.
- Phosphatidylserine translocation was not detected during the blebbing phase.
Conclusions:
- The nuclear envelope and plasma membrane maintain functional integrity until the final stages of apoptosis.
- Cellular membranes show remarkable resilience during the execution phase of programmed cell death.
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