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Caspase enzyme activity is not essential for apoptosis during thymocyte development
P Doerfler1, K A Forbush, R M Perlmutter
1Department of Immunology and Rheumatology, Merck Research Laboratories, Rahway, NJ 07065, USA. Petra_Doerfler@merck.com
Abstract:
Caspases, a family of cysteine proteases, are critical mediators of apoptosis. To address the importance of caspases in thymocyte development, we have generated transgenic mice that express the baculovirus protein p35, a viral caspase inhibitor, specifically in the thymus. p35 expression inhibited Fas (CD95)-, CD3-, or peptide-induced caspase activity in vitro and conferred resistance to Fas-induced apoptosis. However, p35 did not block specific peptide-induced negative selection in OT1 and HY TCR transgenic mouse models. Even the potent pharmacological caspase inhibitor zVAD-FMK (benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl-ketone) could not prevent peptide-induced deletion of OT1 thymocytes, although it improved basal thymocyte survival in vitro. Moreover, the developmental block observed in rag1-/- thymocytes, which lack pre-TCR signaling, was also not rescued by p35 expression. These results indicate that caspase-independent signal transduction pathways can mediate thymocyte death during normal T cell development.
Insights
Caspase inhibitors like p35 and zVAD-FMK did not prevent thymocyte deletion during T cell development. This suggests that thymocyte death can occur independently of caspases, highlighting alternative cell death pathways.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Caspases are key proteases regulating apoptosis.
- Thymocyte development involves programmed cell death (apoptosis).
Purpose of the Study:
- To investigate the role of caspases in thymocyte development and T cell selection.
- To determine if caspase inhibition affects thymocyte apoptosis during T cell development.
Main Methods:
- Generated transgenic mice expressing the baculovirus caspase inhibitor p35 in the thymus.
- Utilized Fas, CD3, and peptide stimulation in vitro and in vivo.
- Administered the pharmacological caspase inhibitor zVAD-FMK.
- Analyzed thymocyte apoptosis and selection in OT1, HY TCR, and rag1-/- transgenic models.
Main Results:
- p35 expression inhibited Fas- and CD3-induced caspase activity and conferred resistance to Fas-induced apoptosis.
- p35 and zVAD-FMK failed to block peptide-induced negative selection in OT1 and HY TCR thymocytes.
- zVAD-FMK improved basal thymocyte survival but did not rescue developmental block in rag1-/- thymocytes.
Conclusions:
- Thymocyte death during T cell development can be mediated by caspase-independent pathways.
- Suggests alternative signal transduction pathways are involved in thymocyte deletion.