LIG_IRF3_LxIS_1

Sensing of pathogenic microbes and tissue damage by the innate immune system triggers immune cells to secrete cytokines that promote host defence. Type I IFNs are the key cytokines mediating innate antiviral immunity (Stetson,2006). Viral RNA, cytosolic DNA, and the bacterial cell wall component lipopolysaccharide all activate defensive signalling pathways through a number of pattern recognition receptor (PRR)–adaptor protein pairs, including RIG-I–MAVS, cGAS-STING and TLR3/4-TRIF (Liu,2015). Microbial double-stranded (ds) DNA in the cytosol is sensed by cGAS which produces the second messenger cGAMP. cGAMP binds to the adaptor protein STING that is located at the endoplasmic reticulum (ER) surface. Viral dsRNA is sensed in the cytosol by Rig-I-like receptors (RLRs) which activate the adaptor protein MAVS that is located at the mitochondrial surface. Membrane anchored toll-like receptors (TLRs) 3 and 4, which recognize viral dsRNA and bacterial LPS respectively, when activated recruit the adaptor protein TRIF. The three adaptor proteins STING, MAVS and TRIF contain a conserved motif referred to as pLxIS that is phosphorylated by TBK1 or IKKε (Liu,2015). Once phosphorylated, these three adaptor proteins bind to the transcription factor interferon regulatory factor 3 (IRF-3) with affinities between 43 and 104 μM, resulting in TBK1-dependent phosphorylation of an additional pLxIS motif in IRF-3. Phosphorylated IRF-3 subsequently dissociates from the adaptor protein and dimerizes through its pLxIS motifs binding same phosphopeptide-binding domain that activates the protein (Zhao,2016). The IRF-3 dimer translocates to the nucleus and positively regulates the transcription of IFN-β (Honda,2006).

The key interaction is between the ϕx{0,2}pLxI(S) motifs, where ϕ is a semi-conserved hydrophobic residue, and the IRF-3 C-terminal region that harbours the positively charged surfaces. In addition, other interactions, primarily electrostatic, upstream of the core motif are also necessary for effective binding.

Interestingly, the rotavirus E3 ubiquitin ligase non-structural protein 1 (NSP1) also contains the pLxIS motif which binds to the same binding region in IRF-3, preventing its activation and promoting its degradation. Phosphorylation of the Ser is not necessary for a low affinity binding with IRF-3 (~200 μM), however, a competitive affinity with respect to the other adaptor proteins (16 μM) is observed when the Ser is phosphorylated. NSP1 then behaves as an E3 ligase, inducing the degradation of IRF-3. In this way, rotavirus is able to escape innate immune recognition by interfering with the IRF-3-dependent pathway (Barro,2005).