Role of FLASH in caspase-8-mediated activation of NF-κB: dominant-negative function of FLASH mutant in NF-κB signaling pathway

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Title
Role of FLASH in caspase-8-mediated activation of NF-κB: dominant-negative function of FLASH mutant in NF-κB signaling pathway
Author(s)
J I Jun; C W Chung; H J Lee; J O Pyo; Kee Nyung Lee; Nam-Soon Kim; Yong Sung Kim; Hyang Sook Yoo; T H Lee; E Kim; Y K Jung
Bibliographic Citation
Oncogene, vol. 24, no. 4, pp. 688-696
Publication Year
2005
Abstract
Caspase-8 is the most receptor-proximal, upstream caspase in the caspase cascade and plays a key role in cell death triggered by various death receptors. Here, we addressed the role of endogenous caspase-8 in tumor necrosis factor (TNF)-α-induced activation of NF-κB. Direct targeting of caspase-8 with siRNA and antisense (AS) approaches abolished TNF-α-induced activation of NF-κB in NIH3T3, HeLa, and HEK293 cells as determined with luciferase reporter gene and cell fractionation assays. Reconstitution of caspase-8-deficient C33A cells with processing-defective (P/D) mutant of caspase-8 sensitized the cells to TNF-α for NF-κB activation. In contrast to wild-type caspase-8, death effector domain mutant replacing Asp73 with Ala (caspase-8 (D73A)) failed to activate NF-κB and to bind FLICE-associated huge protein (FLASH) in vitro and in vivo. Instead, caspase-8 (D73A) mutant bound to caspase-8 and blocked NF-κB activation triggered by TNF-α and caspase-8. In addition, expression of an NF-κB-activating domain-deletion mutant of FLASH or transfection of FLASH AS oligonucleotides abolished TNF-α and caspase-8, but not phorbol 12-myristate 13-acetate, -induced activation of NF-κB. Further, immunoprecipitation assays showed that caspase-8 formed triple complex with TRAF2 and FLASH. Taken together, these results suggest that endogenous caspase-8 mediates TNF-α-induced activation of NF-κB via FLASH.
Keyword
Caspase-8Dominant negativeFLASHNF-κBSiRNA
ISSN
0950-9232
Publisher
Springer-Nature Pub Group
DOI
http://dx.doi.org/10.1038/sj.onc.1208186
Type
Article
Appears in Collections:
Division of Biomedical Research > Rare Disease Research Center > 1. Journal Articles
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