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oai:figshare.com:article/33315465 2026-09-30T06:56:52Z category_30881 category_31388 category_30884 category_31973 category_31439 category_31925 category_31100 portal_234 item_type_6 month_year_09_2026

Mutations in the Vicinity of the IRAK3 Guanylate Cyclase Center Impact Its Subcellular Localization and Ability to Modulate Inflammatory Signaling in Immortalized Cell Lines Ilona Turek (9478139) Hong Trang Nguyen (9492191) Charles Galea (11212134) Isaiah Abad (24614616) Lubna Freihat (24614619) David T Manallack (24614622) Tony Velkov (1315626) Helen Irving (8310519) Biological sciences Medical biochemistry and metabolomics Biochemistry and cell biology Physical chemistry Medical microbiology Medicinal and biomolecular chemistry Microbiology cyclic guanosine monophosphate (cGMP) guanylate cyclase human Toll-like receptor 4 (hTLR4) inflammatory cytokines interleukin-1 receptor-associated kinase 3 (IRAK3 or IRAK-M) lipopolysaccharide nuclear factor kappa-light-chain-enhancer of activated B (NFκB) Interleukin-1 receptor-associated kinase 3 (IRAK3) modulates the magnitude of cellular responses to ligands perceived by interleukin-1 receptors (IL-1Rs) and Toll-like receptors (TLRs), leading to decreases in pro-inflammatory cytokines and suppressed inflammation. The molecular mechanism of IRAK3’s action remains unknown. IRAK3 functions as a guanylate cyclase, and its cGMP product suppresses lipopolysaccharide (LPS)-induced nuclear factor kappa-light-chain-enhancer of activated B cell (NFκB) activity. To understand the implications of this phenomenon, we expanded the structure–function analyses of IRAK3 through site-directed mutagenesis of amino acids known or predicted to impact different activities of IRAK3. We verified the capacity of the mutated IRAK3 variants to generate cGMP in vitro and revealed residues in and in the vicinity of its GC catalytic center that impact the LPS-induced NFκB activity in immortalized cell lines in the absence or presence of an exogenous membrane-permeable cGMP analog. Mutant IRAK3 variants with reduced cGMP generating capacity and differential regulation of NFκB activity influence subcellular localization of IRAK3 in HEK293T cells and fail to rescue IRAK3 function in IRAK3 knock-out THP-1 monocytes stimulated with LPS unless the cGMP analog is present. Together, our results shed new light on the mechanism by which IRAK3 and its enzymatic product control the downstream signaling, affecting inflammatory responses in immortalized cell lines.<p></p> 2023-05-10T00:00:00Z Text Journal contribution 10.3390/ijms24108572 https://figshare.com/articles/journal_contribution/Mutations_in_the_Vicinity_of_the_IRAK3_Guanylate_Cyclase_Center_Impact_Its_Subcellular_Localization_and_Ability_to_Modulate_Inflammatory_Signaling_in_Immortalized_Cell_Lines/33315465 CC BY 4.0