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CASH, a novel caspase homologue with death effector domains
Y V Goltsev1, A V Kovalenko, E Arnold
1Department of Membrane Research and Biophysics, Weizmann Institute of Science, 76100 Rehovot, Israel.
Abstract:
CASP-8 and CASP-10, members of a cysteine protease family that participates in apoptosis, interact with MORT1/FADD, an adapter protein in the CD120a (p55 tumor necrosis factor receptor), and CD95 (Fas/Apo-1) death-inducing signaling pathways, through a shared N-terminal sequence motif, the death effector domain. We report cloning of two splice variants of a novel protein, CASH, that contain two N-terminal death effector domains and can bind through them to each other, to MORT1/FADD, to CASP-8, and to CASP-10. The unique C-terminal part of the longer variant shows marked sequence homology to the caspase protease region yet lacks several of the conserved caspase active site residues, suggesting that it is devoid of cysteine protease activity. Overexpression of the short CASH splice variant strongly inhibited cytotoxicity induction by CD120a and CD95. Expression of the longer variant, while inhibiting cytotoxicity in HeLa cells, had a marked cytocidal effect in 293 cells that could be shown to involve its protease homology region. The findings suggest that CASH acts as an attenuator and/or initiator in CD95 and CD120a signaling for cell death.
Insights
A novel protein, CASH, interacts with apoptosis pathway proteins. Its variants modulate cell death signaling, with one inhibiting cytotoxicity and the other inducing it via its protease domain.
Area of Science:
- Molecular Biology
- Cell Signaling
- Apoptosis Research
Background:
- Caspase-8 (CASP-8) and Caspase-10 (CASP-10) are cysteine proteases involved in apoptosis.
- These caspases interact with MORT1/FADD, an adapter protein in death-inducing signaling pathways like CD120a (p55 tumor necrosis factor receptor) and CD95 (Fas/Apo-1).
- Interaction occurs via a shared N-terminal motif known as the death effector domain.
Purpose of the Study:
- To clone and characterize novel splice variants of a protein interacting with apoptosis pathway components.
- To investigate the binding properties and functional roles of these CASH variants in cell death signaling.
- To elucidate the mechanism by which CASH influences cytotoxicity mediated by CD120a and CD95.
Main Methods:
- Cloning of two splice variants of the novel protein CASH.
- Analysis of CASH protein interactions with MORT1/FADD, CASP-8, and CASP-10 using their death effector domains.
- Overexpression studies of CASH variants in HeLa and 293 cells to assess effects on cytotoxicity.
- Examination of the protease homology region in the longer CASH variant.
Main Results:
- Two splice variants of CASH were cloned, both containing N-terminal death effector domains enabling self-binding and binding to MORT1/FADD, CASP-8, and CASP-10.
- The longer CASH variant possesses a C-terminal region homologous to caspase proteases but lacks key active site residues, suggesting impaired protease activity.
- Overexpression of the short CASH variant inhibited CD120a and CD95-induced cytotoxicity.
- The longer CASH variant inhibited cytotoxicity in HeLa cells but induced cell death in 293 cells, involving its protease homology region.
Conclusions:
- CASH functions as a novel protein interacting with key apoptosis regulators.
- The distinct splice variants of CASH exhibit differential roles in modulating CD120a and CD95 signaling pathways.
- CASH can act as both an attenuator and an initiator in apoptosis signaling, depending on the variant and cellular context.