TAK1ng aim at mechanisms of allergic inflammation: contribution of TAK1 activity in allergen‐mediated mast cell activation

Abstract

Allergic inflammatory disorders are at epidemic levels worldwide. Mast cells drive this inappropriate immune response via release of a variety of pro‐inflammatory mediators in response to environmental cues detected by the IgE‐FcɛRI complex, but the contributing molecular mechanisms are not fully understood. Transforming growth factor β‐activated kinase 1 (TAK1) is a known participant in related signaling through both the MAPK and NFκB pathways; however, the role of TAK1 in IgE‐FcɛRI mediated signaling and resultant allergic inflammation remains unknown. This study was carried out to assess the role of TAK1 in IgE‐mediated mast cell activation and to determine the effect its inhibition would have on the severity of an allergic inflammatory response. Assessment of the role of TAK1 was conducted by exploiting the characterized inhibitory function of 5Z‐7‐oxozeaenol (OZ), a resorcylic acid lactone established to be an ATP‐competitive selective inhibitor of TAK1. Bone marrow‐derived mast cells were sensitized with allergen‐specific IgE and treated with the corresponding allergen for various times in the presence or absence of OZ. Mast cell signaling was measured by western blotting, induced gene expression by qPCR, pro‐inflammatory mediator release by ELISA, mast cell degranulation by β‐hexosaminidase release assay, and calcium mobilization alterations via Indo‐1 based spectrofluorometry. We detect novel activation of TAK1 at Ser412 in response to IgE‐mediated activation under SCF‐c‐kit potentiation in a mast cell‐driven response characteristic of allergic inflammation, which is potently blocked by TAK1 inhibitor OZ. We therefore interrogated the role of TAK1 in a series of mast cell mediator responses using IgE‐sensitized murine bone marrow‐derived mast cells, stimulated with allergen under several TAK1 inhibition strategies. TAK1 inhibition resulted in significant impairment in the phosphorylation of MAPKs p38, ERK, and JNK; and mediation of the NFκB pathway via IκBα. Impaired gene expression and near abrogation in release of pro‐inflammatory cytokines TNF (p<0.0001), IL‐6 (p<0.0001), IL‐13 (p<0.0001), and chemokines CCL1 (p=0.0002), and CCL2 (p<0.0001) was detected. Finally, a significant inhibition of mast cell degranulation (p<0.0001), accompanied by an impairment in calcium mobilization, was observed in the TAK1‐inhibited cells. These results suggest that TAK1 acts as a signaling node, not only linking the MAPK and NFκB pathways in driving the late‐phase response, but also initiation of the degranulation mechanism of the mast cell early‐phase response following allergen recognition and may warrant consideration in future therapeutic development.Support or Funding InformationFunding: This work was supported by the Natural Sciences and Engineering Research Council (NSERC), the Canada Foundation for Innovation (CFI), the Ontario Research Fund (ORF), and Ontario Graduate Scholarships.