Chiara Romagnani

In adaptive immunity, antigen receptor specificity and affinity drive clonal selection, but what enables individual innate lymphocytes to gain a selective advantage without rearranged antigen receptors remains unknown. We traced somatic mutations in human natural killer (NK) cells, revealing extensive clonal dominance in cytomegalovirus-associated memory, with individual genetically diversified clones comprising up to 95% of the population, originating early in life and expanding over years. Their genetic evolution demonstrated positive selection of acquired coding mutations, linking somatic variation to NK cell clonal fitness. Among candidate drivers, we identified mutations in the chromatin regulator KMT2D, whose induced deletion in mature NK cells conferred a competitive advantage following infection in vivo. These findings establish somatic evolution as a fundamental mechanism shaping innate immune memory.

Chronic forms of autoimmune arthritis, including juvenile idiopathic arthritis (JIA), are characterised by persistent synovial inflammation. While adaptive mechanisms are well-studied, the innate networks driving early joint disease remain poorly defined. Here, through unbiased profiling of innate lymphoid cells from the joints of children with JIA at disease onset, we identify activated natural killer (NK) cells as key orchestrators of adaptive immune niches. The selective upregulation of lymphotactin (XCL1) by activated NK cells is accompanied by the synovial enrichment of XCR1 ⁺ type 1 conventional dendritic cells (cDC1s), directly correlating with disease severity. In situ, NK cells spatially anchor with recruited cDC1s to assemble discrete, multicellular effector niches alongside T cells. Finally, we demonstrate this NK-cDC1 spatial architecture represents a conserved pathogenic module across adult arthropathies, including rheumatoid arthritis. Our findings establish the XCL1-XCR1 axis as a fundamental innate feature of chronic joint inflammation, defining a targetable mechanism that precedes and promotes downstream adaptive responses.

Multicellular cytokine networks drive intestinal inflammation and colitis-associated cancer (CAC). Interleukin-22 (IL-22) exerts both protective and pathogenic effects in the intestine, but the mechanisms that regulate this balance remain unclear. Here, we identify that IL-22 directly induces responsiveness to the IL-6 family cytokine oncostatin M (OSM) in intestinal epithelial cells (IECs) by activating STAT3 and upregulating the OSM receptor. In turn, OSM synergizes with IL-22 to sustain STAT3 activation in IECs and promote proinflammatory epithelial adaptation and immune cell chemotaxis to the inflamed intestine. Conditional deletion of the OSM receptor in IECs protects mice from both colitis and CAC, and pharmacological blockade of OSM attenuates established CAC. Thus, IL-22 and OSM form a pathogenic circuit that drives inflammation and tumorigenesis. Our findings reveal a previously unknown mechanism by which OSM supports intestinal pathology and highlight the IL-22–OSM axis as a promising therapeutic target for inflammatory bowel disease and CAC.

Multicellular cytokine networks orchestrate the onset and progression of intestinal inflammation and colitis-associated cancer (CAC). Interleukin 22 (IL-22), a member of the IL-10 superfamily, is known for its ability to promote epithelial cell recovery and repair. However, dysregulation of IL-22 can inadvertently fuel intestinal inflammation and tumorigenesis through mechanisms that are not completely understood. Here we show that IL-22, derived from group 3 innate lymphoid cells (ILC3), directly triggers oncostatin M (OSM) responsiveness in intestinal epithelial cells by activating the transcription factor STAT3 and upregulating OSM receptor (OSMR) expression. OSM, a member of the IL-6 cytokine family implicated in inflammatory bowel disease, collaborates with IL-22 to sustain STAT3 activation in epithelial cells, thereby orchestrating proinflammatory epithelial adaptations and immune cell chemotaxis to the intestine. Conditional deletion of OSMR in epithelial cells protects mice from both colitis and CAC. Furthermore, pharmacological blockade of OSM reduces the progression of established CAC. Our study reveals a novel mechanism by which OSM sustains intestinal inflammation and CAC and identifies the IL-22-OSM axis as a promising therapeutic target for intestinal inflammation and tumorigenesis.

Natural killer (NK) cells are innate lymphoid cells (ILCs) contributing to immune responses to microbes and tumors. Historically, their classification hinged on a limited array of surface protein markers. Here, we used single-cell RNA sequencing (scRNA-seq) and cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) to dissect the heterogeneity of NK cells. We identified three prominent NK cell subsets in healthy human blood: NK1, NK2 and NK3, further differentiated into six distinct subgroups. Our findings delineate the molecular characteristics, key transcription factors, biological functions, metabolic traits and cytokine responses of each subgroup. These data also suggest two separate ontogenetic origins for NK cells, leading to divergent transcriptional trajectories. Furthermore, we analyzed the distribution of NK cell subsets in the lung, tonsils and intraepithelial lymphocytes isolated from healthy individuals and in 22 tumor types. This standardized terminology aims at fostering clarity and consistency in future research, thereby improving cross-study comparisons.

Summary: The central nervous system (CNS) contains a pool of innate lymphoid cells (ILCs) of unclear composition and functionality and unknown origin. Here, we demonstrate that group 1 ILCs (inc. ex-ILC3s) and ILC2s are resident cells with low proliferative capacities and subtype specific CNS compartmentalization. We show for the first time that CNS ILC seeding and niche establishment occurs during early life and is initiated by both, ILC progenitors-like PLFZ ⁺ PD- 1 ⁺ cells and lineage committed ILCs. While group 1 ILCs and RORγt ⁺ ILC3s were found within the embryonic and postnatal brain, ILC2s reached the CNS after birth and were predominantly localized within the dura mater, proving early regional distribution. Interestingly, RORγt ⁺ ILC3s were only detected perinatally and vanished from the CNS as an outcome of decreased turnover and in situ ILC3-to-ILC1 conversion. Remarkably, we showed that perinatal RORγt ⁺ ILC3s are required for the correct development of the lymphatic vessels within the dura.

Summary: Clonal expansion of antigen‐specific lymphocytes is the fundamental mechanism enabling potent adaptive immune responses and the generation of immune memory. Accompanied by pronounced epigenetic remodeling, the massive proliferation of individual cells generates a critical mass of effectors for the control of acute infections, as well as a pool of memory cells protecting against future pathogen encounters. Classically associated with the adaptive immune system, recent work has demonstrated that innate immune memory to human cytomegalovirus (CMV) infection is stably maintained as large clonal expansions of natural killer (NK) cells, raising questions on the mechanisms for clonal selection and expansion in the absence of re‐arranged antigen receptors. Here, we discuss clonal NK cell memory in the context of the mechanisms underlying clonal competition of adaptive lymphocytes and propose alternative selection mechanisms that might decide on the clonal success of their innate counterparts. We propose that the integration of external cues with cell‐intrinsic sources of heterogeneity, such as variegated receptor expression, transcriptional states, and somatic variants, compose a bottleneck for clonal selection, contributing to the large size of memory NK cell clones.

Natural killer (NK) cells are innate lymphoid cells (ILCs) that serve as a first line of defense against pathogens and tumors. Historically, their classification hinged on a limited array of surface protein markers. However, recent advancements in single-cell technologies, such as single-cell RNA sequencing (scRNA-seq) and cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq), have unveiled a more complex and nuanced understanding of NK cells. This has led to variations in nomenclature and inconsistencies across scientific literature. In our study, we utilized these technologies to dissect the heterogeneity of NK cells in healthy human blood. We identified three prominent NK cell subsets: NK1, NK2 and NK3, further differentiated into six distinct subgroups. All subsets were present in all donors, irrespective of their human cytomegalovirus (HCMV) status. Our findings delineate the unique molecular characteristics, key transcription factors, principal biological functions, significant metabolic traits, and cytokine responses of each subgroup. These data also suggest two separate ontogenetic origins for NK cells, leading to divergent transcriptional trajectories. We explore these pathways in depth and propose a comprehensive, standardized nomenclature for NK cells. This standardized terminology aims at fostering clarity and consistency in future research, thereby improving cross-study comparisons.

Natural killer (NK) cells are innate cytotoxic lymphocytes with adaptive immune features, including antigen specificity, clonal expansion and memory. As such, NK cells share many transcriptional and epigenetic programs with their adaptive CD8⁺ T cell siblings. Various signals ranging from antigen, co-stimulation and proinflammatory cytokines are required for optimal NK cell responses in mice and humans during virus infection; however, the integration of these signals remains unclear. In this study, we identified that the transcription factor IRF4 integrates signals to coordinate the NK cell response during mouse cytomegalovirus infection. Loss of IRF4 was detrimental to the expansion and differentiation of virus-specific NK cells. This defect was partially attributed to the inability of IRF4-deficient NK cells to uptake nutrients required for survival and memory generation. Altogether, these data suggest that IRF4 is a signal integrator that acts as a secondary metabolic checkpoint to orchestrate the adaptive response of NK cells during viral infection.