A New Domain in the Toll/IL-1R Domain-Containing Adaptor Inducing Interferon-β Factor Protein Amino Terminus Is Important for Tumor Necrosis Factor-α Receptor-Associated Factor 3 Association, Protein Stabilization and Interferon Signaling

Toll/IL-1R domain-containing adaptor inducing interferon-beta (IFN-beta) factor (TRIF) is a key adaptor for Toll-like receptor (TLR) 3 and TLR4 signaling. Using a novel cDNA isolate encoding a TRIF protein with a 21-residue deletion (Delta 60-181) from its amino-terminal half, we investigated the impact of this deletion on TRIF functions. Transfection studies consistently showed higher expression levels of the (Delta 60-181) TRIF compared to wild-type (wt) TRIF, an effect unrelated to apoptosis, cell lines or plasmid amplification. Colocalization of wt and (Delta 60-181) TRIF proteins led to a dramatic reduction of their respective expressions, suggesting that wt/(Delta 60-181) TRIF heterocomplexes are targeted for degradation. We demonstrated that wt TRIF associates with tumor necrosis factor-alpha receptor-associated factor 3 (TRAF3) better than (Delta 60-181) TRIF, culminating in its greater ubiquitination and proteolysis. This explains, in part, the differential expression levels of the two TRIF proteins. Despite higher expression levels in transfected cells, (Delta 60-181) TRIF inefficiently transactivated the IFN pathway, whereas the nuclear factor-kappa B (NE-kappa B) pathway activation remained similar to that by wt TRIF. In coexpression studies, (Delta 60-181) TRIF marginally contributed to the IFN pathway activation, but still enhanced NE-kappa B signaling with wt TRIF. Therefore, this 21 amino acid sequence is crucial for TRAF3 association, modulation of TRIF stability and activation of the IFN pathway. (C) 2014 S. Karger AG, Basel