Gut microbiota is increasingly linked to the development of various pulmonary diseases through a gut–lung axis. However, the mechanisms by which gut commensal microbes impact trafficking and functional transition of immune cells remain largely unknown. Using integrated microbiota dysbiosis approaches, we uncover that the gut microbiota directs the migration of group 2 innate lymphoid cells (ILC2s) from the gut to the lung through a gut–lung axis. We identify Proteobacteria as a critical species in the gut microbiome to facilitate natural ILC2 migration, and increased Proteobacteria induces IL-33 production. Mechanistically, IL-33–CXCL16 signaling promotes the natural ILC2 accumulation in the lung, whereas IL-25–CCL25 signals augment inflammatory ILC2 accumulation in the intestines upon abdominal infection, parabiosis, and cecum ligation and puncture in mice. We reveal that these two types of ILC2s play critical but distinct roles in regulating inflammation, leading to balanced host defense against infection. Overall results delineate that Proteobacteria in gut microbiota modulates ILC2 directional migration to the lung for host defense via regulation of select cytokines (IL-33), suggesting novel therapeutic strategies to control infectious diseases.

Influenza A Virus (IAV) infection triggers an exuberant host response that promotes acute lung injury. However, the host response factors that promote the development of a pathologic inflammatory response to IAV remain incompletely understood. In this study, we identify an interferon (IFN)-γ-regulated subset of monocytes, CCR2⁺ monocytes, as a driver of lung damage during IAV infection. IFN-γ regulates the recruitment and inflammatory phenotype of CCR2⁺ monocytes, and mice deficient in CCR2 (CCR2^−/−) or IFN-γ (IFN-γ^−/−) exhibit reduced lung inflammation, pathology, and disease severity. Adoptive transfer of WT (IFN-γR1^+/+), but not IFN-γR1⁻/⁻ CCR2⁺ monocytes, restore the wild-type (WT)-like pathological phenotype of lung damage in IAV-infected CCR2^−/− mice. CD8⁺ T cells are the main source of IFN-γ in IAV-infected lungs. Collectively, our data highlights the requirement of IFN-γ signaling in the regulation of CCR2⁺ monocyte-mediated lung pathology during IAV infection.

The dual role of CD8 ⁺ T cells in influenza control and lung pathology is increasingly appreciated. To explore whether protective and pathological functions can be linked to specific subsets, we dissected CD8 ⁺ T responses in influenza-infected murine lungs. Our single-cell RNA-sequencing (scRNA-seq) analysis revealed notable diversity in CD8 ⁺ T subpopulations during peak viral load and infection-resolved state. While enrichment of a Cxcr3 ^hi CD8 ⁺ T effector subset was associated with a more robust cytotoxic response, both CD8 ⁺ T effector and central memory exhibited equally potent effector potential. The scRNA-seq analysis identified unique regulons regulating the cytotoxic response in CD8 ⁺ T cells. The late-stage CD8 ⁺ T blockade in influenza-cleared lungs or continuous CXCR3 blockade mitigated lung injury without affecting viral clearance. Furthermore, adoptive transfer of wild-type CD8 ⁺ T cells exacerbated influenza lung pathology in Cxcr3 ^−/− mice. Collectively, our data imply that CXCR3 interception could have a therapeutic effect in preventing influenza-linked lung injury.

Bitter receptors function primarily in sensing taste, but may also have other functions, such as detecting pathogenic organisms due to their agile response to foreign objects. The mouse taste receptor type-2 member 138 (TAS2R138) is a member of the G-protein-coupled bitter receptor family, which is not only found in the tongue and nasal cavity, but also widely distributed in other organs, such as the respiratory tract, gut, and lungs. Despite its diverse functions, the role of TAS2R138 in host defense against bacterial infection is largely unknown. Here, we show that TAS2R138 facilitates the degradation of lipid droplets (LDs) in neutrophils during Pseudomonas aeruginosa infection through competitive binding with PPARG (peroxisome proliferator-activated receptor gamma) antagonist: N-(3-oxododecanoyl)-l-homoserine lactone (AHL-12), which coincidently is a virulence-bound signal produced by this bacterium (P. aeruginosa). The released PPARG then migrates from nuclei to the cytoplasm to accelerate the degradation of LDs by binding PLIN2 (perilipin-2). Subsequently, the TAS2R138–AHL-12 complex targets LDs to augment their degradation, and thereby facilitating the clearance of AHL-12 in neutrophils to maintain homeostasis in the local environment. These findings reveal a crucial role for TAS2R138 in neutrophil-mediated host immunity against P. aeruginosa infection.

The majority of outer membrane (OM) lipoproteins in Gram-negative bacteria are tethered to the membrane via an attached lipid moiety and oriented facing in toward the periplasmic space; a few lipoproteins have been shown to be surface exposed. The outer membrane lipoprotein P6 from the Gram-negative pathogenic bacterium nontypeable Haemophilus influenzae (NTHi) is surface exposed and a leading vaccine candidate for prevention of NTHi infections. However, we recently found that P6 is not a transmembrane protein as previously thought (L. V. Michel, B. Kalmeta, M. McCreary, J. Snyder, P. Craig, M. E. Pichichero, Vaccine 29:1624–1627, 2011). Here we pursued studies to show that P6 has a dual orientation, existing infrequently as surface exposed and predominantly as internally oriented toward the periplasmic space. Flow cytometry using three monoclonal antibodies with specificity for P6 showed surface staining of whole NTHi cells. Confocal microscopy imaging confirmed that antibodies targeted surface-exposed P6 of intact NTHi cells and not internal P6 in membrane-compromised or dead cells. Western blots of two wild-type NTHi strains and a mutant NTHi strain that does not express P6 showed that P6 antibodies do not detect a promiscuous epitope on NTHi. Depletion of targets to nonlipidated P6 significantly decreased bactericidal activity of human serum. Protease digestion of surface-exposed P6 demonstrated that P6 is predominantly internally localized in a manner similar to its homologue Pal in Escherichia coli. We conclude that P6 of NTHi is likely inserted into the OM in two distinct orientations, with the predominant orientation facing in toward the periplasm.