Bruton tyrosine kinase (BTK) is present in a wide variety of cells and may thus have important non–B cell functions. Here, we explored the function of this kinase in macrophages with studies of its regulation of the NLR family, pyrin domain–containing 3 (NLRP3) inflammasome. We found that bone marrow–derived macrophages (BMDMs) from BTK-deficient mice or monocytes from patients with X-linked agammaglobulinemia (XLA) exhibited increased NLRP3 inflammasome activity; this was also the case for BMDMs exposed to low doses of BTK inhibitors such as ibrutinib and for monocytes from patients with chronic lymphocytic leukemia being treated with ibrutinib. In mechanistic studies, we found that BTK bound to NLRP3 during the priming phase of inflammasome activation and, in doing so, inhibited LPS- and nigericin-induced assembly of the NLRP3 inflammasome during the activation phase of inflammasome activation. This inhibitory effect was caused by BTK inhibition of protein phosphatase 2A–mediated (PP2A-mediated) dephosphorylation of Ser5 in the pyrin domain of NLRP3. Finally, we show that BTK-deficient mice were subject to severe experimental colitis and that such colitis was normalized by administration of anti–IL-β or anakinra, an inhibitor of IL-1β signaling. Together, these studies strongly suggest that BTK functions as a physiologic inhibitor of NLRP3 inflammasome activation and explain why patients with XLA are prone to develop Crohn’s disease.
Liming Mao, Atsushi Kitani, Eitaro Hiejima, Kim Montgomery-Recht, Wenchang Zhou, Ivan Fuss, Adrian Wiestner, Warren Strober
Submitter: Alexander Weber | email@example.com
Authors: Alexander Weber, Xiao Liu, Ana Tapia Abellán, Sangeetha Shankar, Zsofía Bittner and Rimpei Morita
Eberhard Karls University of Tübingen, Germany
Published March 23, 2020
Work by our labs recently identified Bruton’s tyrosine kinase (BTK) (Weber Front Immunol 2017) as a positive regulator of NLRP3 inflammasome function (Liu J Allergy Clin Immunol 2017, Ito Nat Commun 2015): We observed reduced IL-1β release in BTK-deficient mouse cells, whole animals, cell lines, and primary cells from BTK-deficient (X-linked agammaglobulinemia) and BTK-inhibitor-treated patients. Here, Mao et al. propose that BTK acts conversely, as a negative NLRP3 regulator. This is based on results from human and mouse BTK-deficient cells primed with 200 ng/ml LPS, which secreted more mature IL-1β than WT cells. Conversely, at LPS concentrations <100 ng/ml – which are more physiological (Zweigner Blood 2001, Copeland Clin Diagn Lab Immunol 2005) – , they observed lower IL-1β under BTK ablation, in actual agreement with our data. Mao et al. suggest that, in the absence of BTK, low LPS may insufficiently prime the inflammasome. However, under the LPS concentrations we used, IL1B mRNA induction, an important consequence of priming, and secretion of inflammasome-independent cytokines, were always comparable in WT and BTK-deficient murine or human samples. Additionally, in their work NLRP3 oligomerization and adaptor recruitment occur by LPS alone, i.e. without an NLRP3-specific stimulus, indicating that the excessive LPS stimulation used may blur the lines between TLR priming and NLRP3 activation. Mechanistically, Mao et al. implicate BTK-mediated deactivation of the NLRP3-activating phosphatase PP2A under these conditions. It will be interesting to explore this novel mechanism in relationship to direct tyrosine phosphorylation of NLRP3 by BTK which we recently investigated (Bittner bioRxiv 2019). Finally, an aggravated colitis phenotype observed by Mao et al. in BTK-deficient (and hence B cell-depleted) animals in vivo appears to argue for a negative inflammasome role and to parallel increased colitis in XLA and BTK inhibitor-treated leukemia patients. However, a genetically- or BTK inhibitor-induced lack of (regulatory) B cells is well known to exacerbate colitis in mice and humans (Yanaba Am J Pathol 2011, Wang Mucosal Immunol 2015, Kondo Leukemia 2018) and thus a likely and critical confounder in their experiments. In our opinion even the presented in vivo evidence therefore does not unequivocally support the conclusion of BTK as an exclusively negative regulator of NLRP3 inflammasome activity. Nevertheless, this recent work points to a possible influence of TLR signaling in ‘tuning’ BTK for inflammasome regulation. Certainly, our and their studies warrant the further exploration of a possible ‘rheostat role’ of BTK that may tune NLRP3 activation.