The Eukaryotic Linear Motif resource for
Functional Sites in Proteins
Functional site class:
IRF-3 interaction and dimerisation motif
Functional site description:
The recognition of pathogen-associated molecular patterns (PAMPs) involves different pathways that can trigger convergent immune responses. Following different microbial and viral infections, various innate adaptor proteins like STING, MAVS and TRIF trigger IRF-3 activation and production of type I interferons (IFNs) that are essential for host protection. IRF-3 protein is activated by binding to a conserved motif, referred to as pLxIS present in the adaptor proteins that are phosphorylated by TBK1 or IKKε. Once phosphorylated, the motif binds to the transcription factor IRF-3 resulting in TBK1-dependent phosphorylation of an additional motif pLxIS in IRF-3. Phosphorylated IRF-3 then forms a homodimer that activates the protein and positively regulates the transcription of IFN-β. The rotavirus NSP1 protein also contains the pLxIS motif which binds to the same binding region in IRF-3 and is able to escape innate immune recognition by interfering with the IRF-3-dependent pathway.
ELM Description:
The innate immune response is mainly initiated by pathogen-responsive activation of the transcription factor IRF-3. IRF-3 activation is initiated when it binds to innate adaptor proteins like STING, MAVS and TRIF through a conserved phosphorylated motif named pLxIS, where p represents a hydrophilic residue followed by two or three hydrophilic residues and then S represents the phosphorylated serine. Moreover, IRF-3 itself contains the pLxIS motif where it is important for IRF-3 dimerization (including heterodimerization with other IRFs) and activation. A variant of the motif is found in rotavirus NSP1 protein which can bind weakly in the unphosphorylated state, and more strongly after phosphorylation. In the cellular examples, mutation of the serine residue abolishes the interaction.

A five-residue binding model is seen in NSP1-IRF-3 binding and for IRF-3 dimerization (Zhao,2016). All motif instances interact with the same ligand-binding surface of IRF-3. There are interactions involving hydrophilic residues in the motif but they are presently too variable and the number of instances too few for these to be captured in the ELM pattern. However, there is always a semi-conserved hydrophobic residue, with variable spacing, preceding the perfectly conserved positions: This is indicated by ϕ in the pattern ϕx{1,3}LxI(S) entered into ELM. At the C-terminus, the positively charged cluster around Arg285 recognizes the phosphoserine of the pLxIS motif. Subsets of the variably binding residues are involved in pSTING, pMAVS, pTRIF, and NSP1 binding and are likely account for their different affinities towards IRF-3.

Pattern: [VILPF].{1,3}L.I(S)
Pattern Probability: 0.0001590
Present in taxons: Eukaryota Viruses
Interaction Domain:
IRF-3 (PF10401) Interferon-regulatory factor 3 (Stochiometry: 1 : 1)
o See 7 Instances for LIG_IRF3_LxIS_1
o Abstract
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).
o 5 selected references:

o 25 GO-Terms:

o 7 Instances for LIG_IRF3_LxIS_1
(click table headers for sorting; Notes column: =Number of Switches, =Number of Interactions)
Acc., Gene-, NameStartEndSubsequenceLogic#Ev.OrganismNotes
436 442 SPFSGCFEDLAISASTSLGM TP 7 Homo sapiens (Human)
203 210 TGSPASLASNLEISQSPTMP TP 6 Homo sapiens (Human)
Q3TBT3 Tmem173
360 365 PSVLSQEPRLLISGMDQPLP TP 2 Mus musculus (House mouse)
P70671 Irf3
383 388 GASSLKTVDLHISNSQPISL TP 1 Mus musculus (House mouse)
Q86WV6 TMEM173
361 366 TSTMSQEPELLISGMEKPLP TP 7 Homo sapiens (Human)
Q99FX5 Non-structural protein 1
484 489 SGTLTEEFELLISNSEDDNE TP 4 Simian rotavirus A/SA11-4F
Q14653 IRF3
391 396 ASSLENTVDLHISNSHPLSL TP 5 Homo sapiens (Human)
Please cite: The Eukaryotic Linear Motif resource: 2022 release. (PMID:34718738)

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