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Caspase-12 processing and fragment translocation into nuclei of tunicamycin-treated cells
1Divisions of Development and Differentiation, National Institute of Neuroscience, NCNP, Kodaira, Tokyo, Japan.
Abstract:
Excess endoplasmic reticulum (ER) stress induces processing of caspase-12, which is located in the ER, and cell death. However, little is known about the relationship between caspase-12 processing and cell death. We prepared antisera against putative caspase-12 cleavage sites (anti-m12D318 and anti-m12D341) and showed that overexpression of caspase-12 induced autoprocessing at D(318) but did not induce cell death. Mutation analysis confirmed that D(318) was a unique autoprocessing site. In contrast, tunicamycin, one of the ER stress stimuli, induced caspase-12 processing at the N-terminal region and the C-terminal region (both at D(318) and D(341)) and cell death. Anti-m12D318 and anti-m12D341 immunoreactivities were located in the ER of the tunicamycin-treated cells, and some immunoreactivities were located around and in the nuclei of the apoptotic cells. Thus, processing at the N-terminal region may be necessary for the translocation of processed caspase-12 into nuclei and cell death induced by ER stress. Some of the caspase-12 processed at the N-terminal and C-terminal regions may directly participate in the apoptotic events in nuclei.
Insights
Endoplasmic reticulum (ER) stress triggers caspase-12 processing. Specific processing at N-terminal and C-terminal regions, not just autoprocessing, is linked to cell death and nuclear translocation during ER stress.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Endoplasmic reticulum (ER) stress is implicated in cellular dysfunction and death.
- Caspase-12, an ER-resident protease, is activated during ER stress but its precise role in cell death remains unclear.
- Understanding caspase-12 processing is crucial for elucidating ER stress-induced apoptosis pathways.
Purpose of the Study:
- To investigate the relationship between specific caspase-12 processing events and cell death induced by ER stress.
- To determine the cleavage sites involved in caspase-12 activation and their functional consequences.
- To explore the subcellular localization of processed caspase-12 during ER stress-induced apoptosis.
Main Methods:
- Generation of specific antibodies against caspase-12 cleavage sites (D318 and D341).
- Overexpression of wild-type and mutated caspase-12 in cells.
- Induction of ER stress using tunicamycin.
- Immunohistochemical analysis to detect caspase-12 processing and localization.
Main Results:
- Caspase-12 overexpression led to autoprocessing at D318 but did not induce cell death.
- Tunicamycin treatment induced caspase-12 processing at both D318 and D341, correlating with cell death.
- Processed caspase-12 fragments were detected in the ER and translocated to the nucleus in apoptotic cells.
Conclusions:
- Caspase-12 autoprocessing at D318 alone is insufficient for ER stress-induced cell death.
- Processing at both N-terminal and C-terminal regions, including D341, is essential for caspase-12's role in ER stress-induced apoptosis.
- Nuclear translocation of processed caspase-12 suggests its direct involvement in apoptotic events within the nucleus.