Type 2 inflammation is found in most forms of asthma, which may co-exist with recurrent viral infections, bacterial colonization, and host cell death. These processes drive the accumulation of intracellular cyclic-di-nucleotides such as cyclic-di-GMP (CDG). Group 2 innate lymphoid cells (ILC2s) are critical drivers of type 2 lung inflammation during fungal allergen exposure in mice; however, it is unclear how CDG regulates lung ILC responses during lung inflammation. Here, we show that intranasal CDG induced early airway type 1 interferon (IFN) production and dramatically suppressed CD127+ST2+ ILC2s and type 2 lung inflammation during Alternaria and IL-33 exposure. Further, CD127–ST2–Thy1.2+ lung ILCs, which showed a transcriptomic signature consistent with ILC1s, were expanded and activated by CDG combined with either Alternaria or IL-33. CDG-mediated suppression of type 2 inflammation occurred independent of IL-18R, IL-12, and STAT6 but required the stimulator of interferon genes (STING) and type 1 IFN signaling. Thus, CDG potently suppresses ILC2-driven lung inflammation and promotes ILC1 responses. These results suggest potential therapeutic modulation of STING to suppress type 2 inflammation and/or increase anti-viral responses during respiratory infections.

Innate lymphoid cells (ILC) promote lung inflammation in asthma through cytokine production. RNA-binding proteins (RBPs) are critical post-transcriptional regulators, although less is known about RBPs in ILC biology. Here, we demonstrate that RNA-binding motif 3 (RBM3) is highly expressed in lung ILCs and is further induced by alarmins TSLP and IL-33. Rbm3−/− and Rbm3−/−Rag2−/− mice exposed to asthma-associated Alternaria allergen develop enhanced eosinophilic lung inflammation and ILC activation. IL-33 stimulation studies in vivo and in vitro show that RBM3 suppressed lung ILC responses. Further, Rbm3−/− ILCs from bone marrow chimeric mice display increased ILC cytokine production suggesting an ILC-intrinsic suppressive function of RBM3. RNA-sequencing of Rbm3−/− lung ILCs demonstrates increased expression of type 2/17 cytokines and cysteinyl leukotriene 1 receptor (CysLT1R). Finally, Rbm3−/−Cyslt1r−/− mice show dependence on CysLT1R for accumulation of ST2+IL-17+ ILCs. Thus, RBM3 intrinsically regulates lung ILCs during allergen-induced type 2 inflammation that is partially dependent on CysLT1R.

Type 2 inflammation is found in most forms of asthma, which may co-exist with recurrent viral infections, bacterial colonization, and host cell death. These processes drive the accumulation of intracellular cyclic-di-nucleotides such as cyclic-di-GMP (CDG). Group 2 innate lymphoid cells (ILC2s) are critical drivers of type 2 lung inflammation during fungal allergen exposure in mice; however, it is unclear how CDG regulates lung ILC responses during lung inflammation. Here, we show that CDG induced early airway type 1 interferon (IFN) production and dramatically suppressed both lung proliferating CD127+ST2+ ILC2s and Alternaria- and IL-33-induced lung inflammation. Further, transcriptomic analysis of CD127−ST2−Thy1.2+ ILCs, which were expanded and activated by CDG, revealed an ILC1 signature. CDG administration led to accumulation of IFNγ+ and T-bet+ ILC1s, as well as neutrophilia, independent of IL-18R, IL-12, and STAT6 but dependent on stimulator of interferon genes (STING) and partially dependent on type 1 IFN signaling. Thus, CDG potently suppresses ILC2-driven lung inflammation and promotes ILC1 responses. These results suggest potential therapeutic modulation of STING to suppress type 2 inflammation and/or increase anti-viral responses during respiratory infections.

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Innate lymphoid cells (ILCs) promote lung inflammation through cytokine production in diseases such as asthma. RNA-binding proteins (RBPs) are critical post-transcriptional regulators of cellular function, including inflammatory responses, though the role of RBPs in innate lymphoid cells is unknown. Here, we demonstrate that RNA-binding motif 3 (RBM3) is one of the most highly expressed RBPs in Thy1.2+ lung ILCs after fungal allergen challenge and is further induced by epithelial cytokines TSLP and IL-33 in both human and mouse ILCs. Single (rbm3−/−) and double (rbm3−/−rag2−/−) knockout mice exposed via the airway to the asthma-associated fungal allergen Alternaria alternata displayed increases in eosinophilic lung inflammation and ILC activation compared to control mice. In addition to increased Th2 cytokine production, rbm3−/− ILCs produced elevated IL-17A. The negative regulation by RBM3 in ILC responses was direct as purified rbm3−/− ILCs were hyperinflammatory in vitro and in vivo after stimulation with IL-33. Transcriptomic analysis by RNA-sequencing of rbm3−/− lung ILCs showed increased type 2 and 17 cytokines as well as global expression differences in critical cytokines, receptors, transcription factors, and survival transcripts compared with WT ILCs. Intriguingly, these transcript changes did not correlate with the presence of AU-rich elements (AREs), which RBM3 is known to bind. Thus, regulation of ILC responses by RNA-binding proteins offers novel mechanistic insight into lung ILC biology and ILC-driven inflammatory diseases.