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Role of caspase‐8 ubiquitination and degradation by the 26S proteasome in regulating extrinsic cell death
Author(s) -
Fiandalo Michael V.,
Schwarze Steven R.
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.528.1
Subject(s) - proteasome , ubiquitin , protein subunit , caspase , microbiology and biotechnology , apoptosis , caspase 3 , chemistry , caspase 8 , protein degradation , caspase 2 , hek 293 cells , biology , biochemistry , programmed cell death , gene
Studying the extrinsic and intrinsic pathways of apoptosis has led to new anticancer strategies. Caspase‐8 activation is critical for extrinsic apoptosis induction. Our data have shown that proteasome inhibition, with the chemotherapeutic agent Velcade, sensitizes prostate cancer cells to apoptosis and stabilizes the caspase‐8 p18 subunit. The goal of this study is to identify the amino acids that regulate caspase‐8 stability. Using a HA‐tagged gene approach, several caspase‐8 fragments were expressed and tested for stability and ubiquitin incorporation to define which region was a target for ubiquitination. Consistent with endogenous pro‐caspase‐8, catalytically inactive unprocessed caspase‐8 was readily expressed and protein levels were not enhanced by proteasome inhibition, while expression of the active processed p18 and p10 subunits required Velcade. Furthermore, only the p18 subunit was covalently associated with polyubiquitin. Site‐directed mutagenesis studies are currently underway to mutate lysine residues to arginine in the p18 subunit in an attempt to generate a degradation‐resistant caspase‐8. This study has demonstrated that active caspase‐8 and more specifically the p18 subunit, is a target for ubiquitination and degradation by the 26S proteasome. Future studies will test the extent to which non‐ubiquitinated caspase‐8 can modulate extrinsic apoptosis.