IRAK3 modulates downstream innate immune signalling through its guanylate cyclase activity

Abstract

Interleukin-1 receptor associated kinase 3 (IRAK3) is a cytoplasmic homeostatic mediator of inflammatory responses and is potentially useful as a prognostic marker in inflammation. IRAK3 inhibits signalling cascades downstream of myddosome complexes associated with toll like receptors. IRAK3 contains a death domain that interacts with other IRAK family members, a pseudokinase domain and a C-terminus domain involved with tumour necrosis factor receptor associated factor 6 (TRAF6). Previous bioinformatic studies revealed that IRAK3 contained a guanylate cyclase centre in its pseudokinase domain but its role in IRAK3 action is unresolved. We demonstrate that wildtype IRAK3 is capable of producing cGMP. Furthermore, we show that a specific point mutation in the guanylate cyclase centre reduced cGMP production. Cells containing toll like receptor 4 and a nuclear factor kappa-light-chain-enhancer of activated B cells (NFĸB) reporter system were transfected with IRAK3 or mutant IRAK3 proteins. Cell-permeable cGMP treatment of untransfected control cells suppresses downstream signalling through modulation of the NFĸB in the presence of lipopolysaccharides. Cells transfected with wildtype IRAK3 also suppress lipopolysaccharide induced NFĸB activity in the absence of exogenous cGMP. Lipopolysaccharide induced NFĸB activity was not suppressed in cells transfected with the IRAK3 mutant with reduced cGMP-generating capacity. Whereas in the presence of exogenously applied cell-permeable cGMP the IRAK3 mutant was able to retain its function by suppressing lipopolysaccharide induced NFĸB activity. Furthermore, increasing the amount of membrane permeable cGMP did not affect IRAK3’s ability to reduce NFĸB activity. These results suggest that cGMP generated by IRAK3 may be involved in regulatory function of the protein where the presence of cGMP may selectively affect downstream signalling pathway(s) by modulating binding and/or activity of nearby proteins that interact in the inflammatory signalling cascade.

Genome‐wide transcriptome analysis identifies novel dysregulated genes implicated in Alzheimer's pathology

“…This suggests that lower expression of complement genes may have beneficial effects on the brain, aligning with our data showing lower CD46 expression in controls compared to those with AD. IRAK3 , another of the significantly upregulated genes, encodes an interleukin‐1 receptor–associated kinase, which is ubiquitously expressed and functions in the Toll/IL‐R immune signal transduction pathway 47,63 . Although there was no evidence from our literature search associating IRAK3 with AD, other interleukin 1 receptor–associated kinases have been associated with pro‐inflammatory processes in AD, 64,65 suggesting that these molecules play important roles in AD.…”

Moonlighting Proteins Shine New Light on Molecular Signaling Niches

“…Homology modelling indicates that the pseudokinase domain forms similar fold patterns to models of the kinase domain of PSKR1 with the guanylate cyclase center in a similar position [ 153 ]. Phosphorylation status and kinase activity do not contribute to the guanylate cyclase activity although small amounts of cGMP are important in promoting the ability of IRAK3 to suppress nuclear factor kappa B (NFκB) activity [ 159 ]. Potentially, localized small amounts of cGMP allosterically modulate IRAK3 conformation to promote down-stream effects.…”

“…We suggest that proteins containing cryptic enzymatic activities, such as the guanylate cyclase in WAKL10, PSKR1, BRI1, and PEPR1 generating cGMP-enriched nanodomains, are part of the solution to highly spatially differentiated stimulus-specific cellular signaling and form a new paradigm in cellular signaling and homeostatic responses [ 23 ]. Interestingly, auto-generation of a cGMP-enriched nanodomain is part of the mechanism of action of the cytoplasmic protein IRAK3, which is involved in inhibiting animal cell responses to DAMPs [ 153 , 159 ], indicating that this is potentially a universal paradigm in cell signaling. Establishment of a cGMP-enriched niche or nanodomain in the vicinity of the protein is a puzzle as physics suggests that small molecules will rapidly diffuse away unless they are attached in some way [ 172 ].…”

Analysis of interleukin-1 receptor associated kinase-3 (IRAK3) function in modulating expression of inflammatory markers in cell culture models: A systematic review and meta-analysis

“…NF-κB activation then triggers transcription of anti-inflammatory molecules, which generates an overall inhibitory effect on inflammatory responses [ 7 ]. Furthermore, IRAK3 contains a catalytic guanylate cyclase centre which can generate a nano-environment of cyclic guanosine monophosphate surrounding IRAK3 protein that appears to be involved in the modulatory mechanism of IRAK3 effect on NF-κB activation [ 34 ]. The various actions of IRAK3 on inflammation appear to be affected by ligand specificity and dose [ 35 ], or cell types [ 33 ].…”