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Phosphorylation of initiation factor 2 alpha subunit and apoptosis in Ca2+ ionophore-treated cultured neuronal cells
Abstract:
The initial step of protein synthesis is regulated by the eukaryotic initiation factor 2 (eIF-2) whose phosphorylation in the alpha subunit by specific kinases, as double-stranded RNA-dependent protein kinase (PKR), produces an inhibition of the translational rates. Besides, intracellular Ca2+ mobilization has been associated with an increase in the PKR activity. Our results show, in primary neuronal cultures, that the treatment with the Ca2+ ionophore A23187 induces an increase in the phosphorylation of the alpha subunit of eIF-2 (eIF-2 alpha) and inhibition of protein synthesis. Those biochemical changes run parallel to the appearance of specific apoptosis characteristics such as cell shrinkage, segmentation of chromatin into small round bodies, and cleavage of DNA into 180 bp multimers. These results indicate that one of the targets during the process of apoptosis induced by the rise in the intracellular Ca2+, could be eIF-2 factor.
Insights
Increased intracellular calcium triggers apoptosis by inhibiting protein synthesis via eIF-2 alpha phosphorylation. This study reveals eukaryotic initiation factor 2 (eIF-2) as a key target in calcium-induced cell death.
Area of Science:
- Neuroscience
- Molecular Biology
- Cell Biology
Background:
- Protein synthesis initiation is regulated by eukaryotic initiation factor 2 (eIF-2).
- Phosphorylation of eIF-2 alpha subunit by kinases like PKR inhibits translation.
- Intracellular calcium (Ca2+) mobilization can enhance PKR activity.
Purpose of the Study:
- To investigate the effect of increased intracellular Ca2+ on eIF-2 phosphorylation and protein synthesis in neuronal cultures.
- To determine if eIF-2 is a target in Ca2+-induced apoptosis.
Main Methods:
- Primary neuronal cultures were treated with the Ca2+ ionophore A23187.
- Analysis of eIF-2 alpha phosphorylation levels.
- Assessment of protein synthesis rates.
- Observation of apoptosis markers (cell shrinkage, chromatin segmentation, DNA fragmentation).
Main Results:
- A23187 treatment increased eIF-2 alpha phosphorylation and inhibited protein synthesis.
- Biochemical changes correlated with the onset of apoptosis.
- Apoptosis was characterized by cell shrinkage, chromatin segmentation, and DNA laddering.
Conclusions:
- Elevated intracellular Ca2+ leads to eIF-2 alpha phosphorylation and protein synthesis inhibition in neurons.
- These events are linked to the induction of apoptosis.
- eIF-2 factor is identified as a potential target in Ca2+-induced apoptotic pathways.