Previous studies have demonstrated that CASPASE 8 can generate a prosurvival signal by inhibiting necroptosis via the cleavage of the deubiquitinating enzyme CYLD. Cleavage of CYLD at D215 results in the generation of a 25 kD N-terminal fragment and degradation of the C-terminal fragment containing the catalytic domain. Since CYLD is required for TNF-induced necroptosis, its proteolysis is necessary and sufficient to suppress necroptosis and generate a survival signal. Here we describe how to visualize CYLD proteolysis by western blot analysis, as a measure of CASPASE 8 activity and inhibition of necroptosis.
CITATION STYLE
Legarda, D., & Ting, A. T. (2018). Analysis of CYLD proteolysis by CASPASE 8 in bone marrow-derived macrophages. In Methods in Molecular Biology (Vol. 1857, pp. 181–188). Humana Press Inc. https://doi.org/10.1007/978-1-4939-8754-2_18
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