Ly49 Receptors Research Articles

Expression of a single inhibitory Ly49 receptor is sufficient to license NK cells for effector functions.

Natural killer (NK) cells recognize target cells through germline-encoded activation and inhibitory receptors enabling effective immunity against viruses and cancer. The Ly49 receptor family in the mouse and killer immunoglobin-like receptor family in humans play a central role in NK cell immunity through recognition of MHC class I and related molecules. Functionally, these receptor families are involved in licensing and rejection of MHC-I-deficient cells through missing-self. The Ly49 family is highly polymorphic, making it challenging to detail the contributions of individual Ly49 receptors to NK cell function. Herein, we showed mice lacking expression of all Ly49s were unable to reject missing-self target cells in vivo, were defective in NK cell licensing, and displayed lower KLRG1 on the surface of NK cells. Expression of Ly49A alone on a H-2Dd background restored missing-self target cell rejection, NK cell licensing, and NK cell KLRG1 expression. Thus, a single inhibitory Ly49 receptor is sufficient to license NK cells and mediate missing-self in vivo.

Collaboration between a cis-interacting natural killer cell receptor and membrane sphingolipid is critical for the phagocyte function.

Inhibitory natural killer (NK) cell receptors recognize MHC class I (MHC-I) in trans on target cells and suppress cytotoxicity. Some NK cell receptors recognize MHC-I in cis, but the role of this interaction is uncertain. Ly49Q, an atypical Ly49 receptor expressed in non-NK cells, binds MHC-I in cis and mediates chemotaxis of neutrophils and type I interferon production by plasmacytoid dendritic cells. We identified a lipid-binding motif in the juxtamembrane region of Ly49Q and found that Ly49Q organized functional membrane domains comprising sphingolipids via sulfatide binding. Ly49Q recruited actin-remodeling molecules to an immunoreceptor tyrosine-based inhibitory motif, which enabled the sphingolipid-enriched membrane domain to mediate complicated actin remodeling at the lamellipodia and phagosome membranes during phagocytosis. Thus, Ly49Q facilitates integrative regulation of proteins and lipid species to construct a cell type-specific membrane platform. Other Ly49 members possess lipid binding motifs; therefore, membrane platform organization may be a primary role of some NK cell receptors.

MHC-I upregulation safeguards neoplastic T cells in the skin against NK cell-mediated eradication in mycosis fungoides

Cancer-associated immune dysfunction is a major challenge for effective therapies. The emergence of antibodies targeting tumor cell-surface antigens led to advancements in the treatment of hematopoietic malignancies, particularly blood cancers. Yet their impact is constrained against tumors of hematopoietic origin manifesting in the skin. In this study, we employ a clonality-supervised deep learning methodology to dissect key pathological features implicated in mycosis fungoides, the most common cutaneous T-cell lymphoma. Our investigations unveil the prominence of the IL-32β–major histocompatibility complex (MHC)-I axis as a critical determinant in tumor T-cell immune evasion within the skin microenvironment. In patients’ skin, we find MHC-I to detrimentally impact the functionality of natural killer (NK) cells, diminishing antibody-dependent cellular cytotoxicity and promoting resistance of tumor skin T-cells to cell-surface targeting therapies. Through murine experiments in female mice, we demonstrate that disruption of the MHC-I interaction with NK cell inhibitory Ly49 receptors restores NK cell anti-tumor activity and targeted T-cell lymphoma elimination in vivo. These findings underscore the significance of attenuating the MHC-I-dependent immunosuppressive networks within skin tumors. Overall, our study introduces a strategy to reinvigorate NK cell-mediated anti-tumor responses to overcome treatment resistance to existing cell-surface targeted therapies for skin lymphoma.

Effects of acupuncture on RT-BM1 and maternal-fetal immune tolerance in rats with ovarian hyperstimulation: 针刺对超排卵大鼠RT-BM1及母胎免疫耐受的影响

ObjectiveThis study aimed to evaluate the effects of acupuncture on embryo implantation and maternal–fetal immune tolerance in rats with ovarian hyperstimulation, identify the mechanisms of acupuncture treatment, and promote application of acupuncture in the field of assisted reproduction. MethodsEighty female Wistar rats were randomized into normal (N), model (M), acupuncture treatment (A), and progesterone treatment (P) groups, with 20 rats in each group. An ovarian hyperstimulation model was established in groups M, A, and P by the peritoneal injection of pregnant mare serum gonadotropin (PMSG) combined with human chorionic gonadotropin (HCG). Bilateral “Sanyinjiao (SP6)”, “Zusanli (ST36)” and “Taichong (LR3)” as acupoints were needled in rats in group A. Rats in group P received an intramuscular injection of progesterone (4 mg/kg). All pregnant rats were treated for seven days. On the 8th day, the pregnant rats were sacrificed for testing. The expression levels of estrogen (E2) and progesterone (P4) in serum and their receptors in the endometrium, Th1 and Th2 cytokines in serum and endometrium, RT-BM1 (the counterpart of HLA-G in rats) mRNA, Ly49 activating receptors (Ly49s3, Ly49s5) and inhibitory receptors (Ly49i3, Ly49i4, Ly49i5) on immunocytes in the maternal–fetal interface were detected using ELISA, Western blot, and real-time polymerase chain reaction (PCR). ResultsCompared with group N, the pregnancy rate and the number of implanted embryos were significantly decreased in group M (P < 0.05); E2 and P4 levels in serum, and the protein levels of ER-α and Progesterone Receptor (PGR) in the implantation site of endometrial tissue were significantly reduced in group M (P < 0.05); the expression of INF-γ and IL-2 in serum and endometrium were significantly increased, while the expression of IL-4, IL-6, and IL-10 in serum and endometrium were significantly decreased in group M(P < 0.05). The mRNA levels of RT-BM1, Ly49i3, and Ly49i4 receptors in group M were significantly higher than those in group N, whereas the mRNA levels of Ly49i5 and Ly49s5 receptors in group M were significantly lower than those in group N at the maternal–fetal interface (P < 0.05). The pregnancy rate and number of implanted embryos were significantly higher in group A than in group M (P < 0.05). Serum E2 and P4 levels were significantly higher in groups A and P than in group M (P < 0.05). The expression of ER-α and PGR in the implantation site of endometrial tissue in group A were significantly higher than those in group M (P < 0.05). The serum levels of INF-γ and IL-2 in group A and group P were significantly lower than those in group M (P < 0.05). The serum expression of IL-4, IL-6, IL-10 in group A and group P were significantly higher than those in group M (P < 0.05). The endometrium expression of INF-γ and IL-2 in group A were significantly lower than those in group M (P < 0.05). The endometrium expression of IL-4, IL-6, IL-10 in group A were significantly higher than those in group M (P < 0.05). The endometrium expression of INF-γ in group P were significantly lower than those in group M (P < 0.05). The endometrium expression of IL-6 and IL-10 in group P were significantly higher than those in group M (P < 0.05). The mRNA levels of RT-BM1 and these Ly49 receptors (except the Ly49s3 receptor) at the maternal–fetal interface of groups A and P were higher than those of group M (P < 0.05). ConclusionAcupuncture can significantly increase the pregnancy rate and average number of implanted blastocysts in rats with ovarian hyperstimulation. This mechanism is related to the effects of acupuncture in upregulating the expression of RT-BM1, regulating the balance of Th1/Th2 cytokines, and upregulating the expression of inhibitory receptors in immunocytes.

Two developmental pathways generate functionally distinct populations of natural killer cells

Abstract Natural killer (NK) cells function by eliminating virus-infected cells or tumor cells during early defenses. However, the early development of NK cells and lineage relationships between NK cells and helper innate lymphoid cells (ILCs) remain elusive. Common precursors for ILCs (ILCPs) can differentiate into both helper ILCs and NK cells. Here, we identified a NK lineage-restricted progenitor population, early NK progenitor (ENKP), which does not develop from ILCPs, thus ENKP may represent the ILCP-independent pathway of NK cell development. Competitive chimera experiment shows ENKPs generate NK cells more efficiently than ILCPs, suggesting that ENKP-dependent pathway is the major pathway for NK cell development. scRNA-seq shows ENKP-derived NK cells express Ly49 receptors and higher levels of cytotoxic genes whereas ILCP-derived NK cells have very low expression of Ly49 receptors and express higher levels of genes implicated in tissue residency such as CD69 and CD200R. Furthermore, Ly49H+ NK cells which response to MCMV infection mostly develop from ENKPs but not ILCPs. Consistently, ENKP-derived NK cells but not ILCP-derived NK cells expanded dramatically after MCMV infection. Our findings establish the existence of two pathways of NK cell development that generate functionally distinct NK cell subsets.

Inhibition of Ly-49/LILRB-MHC-I interactions by anti-MHC-I augments innate and adaptive immunity.

Abstract Mouse NK cells recognize MHC-I antigens via stochastically expressed inhibitory Ly49 receptors. The loss of MHC-I expression on tumor cells (“missing self”) abrogates inhibitory signals, resulting in NK activation. We have identified an anti-mouse pan MHC-I mAb (M1/42), which blocks Ly49-MHC-I interactions but not TCR-MHC-I interactions. Administration of M1/42 in vivo markedly activated IFNg-producing NK cells that further drove the proliferation of NK cells and memory phenotype (MP) T cells and profoundly augmented adaptive immunity against viral infection and cancer metastasis. In contrast to mouse Ly-49 antigens, human killer cell inhibitory receptors bind to HLA at a site overlapping the TCR binding site and blocking of KIR binding to MHC-I would inhibit TCR recognition. However, two anti-pan-human MHC-I abs (DX17 and W6/32) markedly induced NK cell proliferation and IFNg production in cultures of human PBMC. DX17 potently blocked the interaction of leukocyte Ig-like inhibitory receptors LILRB1, B2, B3 and B5 with human MHC-I but had no effect on TCR-MHC-I interactions. The crystal structure of a DX17 Fab/MHC-I complex reveals that the footprint of DX17 overlaps the LILRB1 binding site on MHC-I. Administration of W6/32 Fab to humanized mice (PBMC reconstituted NSG or NK cell reconstituted NOG-IL-15) induced human NK and MP T cell proliferation. Marked reduction of tumor size was seen when NOG-hIL-15Tg mice were reconstituted with CD3 −cells and then treated with W6/32 Fab. DX17 Fc silenced mAb (DX17 LALAPG) activated human tumor infiltrating lymphocytes and monocytes. These results strongly suggest that inhibition of LILRB-MHC-I interactions by anti-MHC-I in humans may result in marked augmentation of anti-tumor immunity. This work was supported by the Intramural Research Program of NIAID, NIH.

MHC Class I-Dependent Shaping of the NK Cell Ly49 Receptor Repertoire Takes Place Early during Maturation in the Bone Marrow.

MHC class I (MHC I) expression in the host influences NK cells in a process termed education. The result of this education is reflected in the responsiveness of NK cells at the level of individual cells as well as in the repertoire of inhibitory MHC I-specific receptors at the NK cell system level. The presence of MHC I molecules in the host environment gives rise to a skewed receptor repertoire in spleen NK cells where subsets expressing few (one or two) inhibitory receptors are expanded whereas subsets with many (three or more) receptors are contracted. It is not known whether this MHC I-dependent skewing is imposed during development or after maturation of NK cells. In this study, we tested the hypothesis that the NK cell receptor repertoire is shaped already early during NK cell development in the bone marrow. We used mice with a repertoire imposed by a single MHC I allele, as well as a C57BL/6 mutant strain with exaggerated repertoire skewing, to investigate Ly49 receptor repertoires at different stages of NK cell differentiation. Our results show that NK cell inhibitory receptor repertoire skewing can indeed be observed in the bone marrow, even during the earliest developmental steps where Ly49 receptors are expressed. This may partly be accounted for by selective proliferation of certain NK cell subsets, but other mechanisms must also be involved. We propose a model for how repertoire skewing is established during a developmental phase in the bone marrow, based on sequential receptor expression as well as selective proliferation.

Influence of Self-MHC Class I Recognition on the Dynamics of NK Cell Responses to Cytomegalovirus Infection.

Although interactions between inhibitory Ly49 receptors and their self-MHC class I ligands in C57BL/6 mice are known to limit NK cell proliferation during mouse CMV (MCMV) infection, we created a 36-marker mass cytometry (CyTOF) panel to investigate how these inhibitory receptors impact the NK cell response to MCMV in other phenotypically measurable ways. More than two thirds of licensed NK cells (i.e., those expressing Ly49C, Ly49I, or both) in uninfected mice had already differentiated into NK cells with phenotypes indicative of Ag encounter (KLRG1+Ly6C-) or memory-like status (KLRG1+Ly6C+). These pre-existing KLRG1+Ly6C+ NK cells resembled known Ag-specific memory NK cell populations in being less responsive to IL-18 and IFN-α stimulation in vitro and by selecting for NK cell clones with elevated expression of a Ly49 receptor. During MCMV infection, the significant differences between licensed and unlicensed (Ly49C-Ly49I-) NK cells disappeared within both CMV-specific (Ly49H+) and nonspecific (Ly49H-) responses. This lack of heterogeneity carried into the memory phase, with only a difference in CD16 expression manifesting between licensed and unlicensed MCMV-specific memory NK cell populations. Our results suggest that restricting proliferation is the predominant effect licensing has on the NK cell population during MCMV infection, but the inhibitory Ly49-MHC interactions that take place ahead of infection contribute to their limited expansion by shrinking the pool of licensed NK cells capable of robustly responding to new challenges.

Evidence for Prescribed NK Cell Ly-49 Developmental Pathways in Mice

Previous studies of NK cell inhibitory Ly-49 genes showed their expression is stochastic. However, relatively few studies have examined the mechanisms governing acquisition of inhibitory receptors in conjunction with activating Ly-49 receptors and NK cell development. We hypothesized that the surface expression of activating Ly-49 receptors is nonrandom and is influenced by inhibitory Ly-49 receptors. We analyzed NK cell "clusters" defined by combinatorial expression of activating (Ly-49H and Ly-49D) and inhibitory (Ly-49I and Ly-49G2) receptors in C57BL/6 mice. Using the product rule to evaluate the interdependencies of the Ly-49 receptors, we found evidence for a tightly regulated expression at the immature NK cell stage, with the highest interdependencies between clusters that express at least one activating receptor. Further analysis demonstrated that certain NK clusters predominated at the immature (CD27+CD11b-), transitional (CD27+CD11b+), and mature (CD27-CD11b-) NK cell stages. Using parallel in vitro culture and in vivo transplantation of sorted NK clusters, we discovered nonrandom expression of Ly-49 receptors, suggesting that prescribed pathways of NK cluster differentiation exist. Our data infer that surface expression of Ly-49I is an important step in NK cell maturation. Ki-67 expression and cell counts confirmed that immature NK cells proliferate more than mature NK cells. We found that MHC class I is particularly important for regulation of Ly-49D and Ly-49G2, even though no known MHC class I ligand for these receptors is present in B6 mice. Our data indicate that surface expression of both activating and inhibitory Ly-49 receptors on NK cell clusters occurs in a nonrandom process correlated to their maturation stage.

Functional Characterization of Ly49+CD8 T-Cells in Both Normal Condition and During Anti-Viral Response.

The role of Ly49+CD8 T-cells in the immune system is not clear. Previously, several papers suggested Ly49+CD8 T-cells as immunosuppressors, while multiple studies also suggested their role as potent participants of the immune response. The mechanism of Ly49 expression on CD8 T-cells is also not clear. We investigated phenotype, functions, and regulation of Ly49 expression on murine CD8 T-cells in both normal state and during LCMV infection. CD8 T-cells express different Ly49 receptors compared with NK-cells. In intact mice, Ly49+CD8 T-cells have a phenotype similar to resting central memory CD8 T-cells and do not show impaired proliferation and cytokine production. Conventional CD8 T-cells upregulate Ly49 receptors during TCR-induced stimulation, and IL-2, as well as IL-15, affect it. At the same time, Ly49+CD8 T-cells change the Ly49 expression profile dramatically upon re-stimulation downregulating inhibitory and upregulating activating Ly49 receptors. We observed the expression of Ly49 receptors on the virus-specific CD8 T-cells during LCMV infection, especially marked in the early stages, and participation of Ly49+CD8 T-cells in the anti-viral response. Thus, CD8 T-cells acquire Ly49 receptors during the T-cell activation and show dynamic regulation of Ly49 receptors during stimulation.

Complex variation in the KLRA (LY49) immunity-related genomic region in horses.

Natural killer (NK) cells play important roles in innate and adaptive immunity, as well as in the reproduction of placental mammals. Ly49 (KLRA) molecules represent a lectin-like type of NK cell receptor encoded within a complex genomic region, the NK cell complex. In rodents and horses, an expansion of the genes encoding Ly49 receptors leading to the formation of a gene family was observed. High sequence similarities and frequent high polymorphism of multiple family members represent an obstacle both for their individual identification and for annotation in the reference genomes of their respective species. Here, we focused on resolving complex variation of the KLRA gene family observed in domestic and Przewalski's horses. The KLRA (LY49) genomic region contains six genes (KLRA2-KLRA7) and one putative pseudogene, KLRA1. Two types of polymorphism were observed in the horses analyzed. Copy number variation between haplotypes was documented for the gene KLRA7 by polymerase chain reaction. As expected, the major source of variation of all KLRA genes, including KLRA7, is because of single nucleotide polymorphisms, many of them being nonsynonymous substitutions. Extensive allelic variability of the expanded KLRA (LY49) genes was observed. For four out of the six functional KLRA, high numbers of novel allelic amino acid sequence variants were identified in the genes studied, suggesting that this variation might be of functional importance, especially in the context of high polymorphism of their presumed ligands encoded by major histocompatibility complex class I genes. In fact, polymorphic amino acid sites were mostly found in the ligand-binding C-type lectin-like domain of the putative receptor molecule.

Control of Viral Infection by Natural Killer Cell Inhibitory Receptors

SUMMARYMajor histocompatibility complex class I (MHC-I)-restricted immune responses are largely attributed to cytotoxic T lymphocytes (CTLs). However, natural killer (NK) cells, as predicted by the missing-self hypothesis, have opposing requirements for MHC-I, suggesting that they may also demonstrate MHC-I-restricted effects. In mice, the Ly49 inhibitory receptors prevent NK cell killing of missing-self targets in effector responses, and they have a proposed second function in licensing or educating NK cells via self-MHC-I in vivo. Here we show MHC-I-restricted control of murine cytomegalovirus (MCMV) infection in vivo that is NK cell dependent. Using mice lacking specific Ly49 receptors, we show that control of MCMV requires inhibitory Ly49 receptors and an inhibitory signaling motif and the capacity for MCMV to downregulate MHC-I. Taken together, these data provide definitive evidence that the inhibitory receptors are required for missing-self rejection and are relevant to MHC-I-restricted NK cell control of a viral infection in vivo.

A Central Role for Ly49 Receptors in NK Cell Memory.

In the past decade, the study of NK cells was transformed by the discovery of three ways these "innate" immune cells display adaptive immune behavior, including the ability to form long-lasting, Ag-specific memories of a wide variety of immunogens. In this review, we examine these types of NK cell memory, highlighting their unique features and underlying similarities. We explore those similarities in depth, focusing on the role that Ly49 receptors play in various types of NK cell memory. From this Ly49 dependency, we will build a model by which we understand the three types of NK cell memory as aspects of what is ultimately the same adaptive immune process, rather than separate facets of NK cell biology. We hope that a defined model for NK cell memory will empower collaboration between researchers of these three fields to further our understanding of this surprising and clinically promising immune response.

Evidence for a prescribed NK cell Ly49 developmental pathway in mice

Abstract Previous studies of NK cell Ly49 receptors suggest that the inhibitory receptor expression is stochastic. However, similar studies of activating Ly49 receptors have not been an area of focus. We hypothesize that the expression of activating receptors is not stochastic. We analyzed NK cell “clusters” expressing 16 combinations of Ly49I, Ly49G2, Ly49H and Ly49D in B6 mice. We sorted the 16 clusters, and cultured them with IL-15 for 6 days. In parallel, we also transplanted a select group of clusters in vivo. We discovered a prescribed pathway of cluster transitions in vitro and in vivo, and these data suggest that Ly49 activating receptor acquisition is not stochastic. We found that the majority of clusters that express activating receptors maintain their expression. In addition, Ly49D, which is not reported to have an MHC-I ligand in B6 mice, was upregulated on clusters that transition in the prescribed pathway. Previously, other groups have used the product rule to mathematically predict the independent expression of Ly49 inhibitory receptors. However, the product rule has not been applied to predict activating Ly49 receptor frequencies. Our application of the product rule to our data suggest that Ly49H and Ly49D expression is not independent and cannot be predicted by this mathematical model. This suggests that the biological system that controls the Ly49 activating receptors is distinct from inhibitory receptors. Currently we are investigating whether MHC-I is critical in the prescribed pathway via the transplantation of these clusters in MHC-I-deficient mice. Taken together, these data support the idea that NK cell clusters develop in a sequential, non-random manner that is determined by expression of activating Ly49 receptors.

The complex of MCMV proteins and MHC class I evades NK cell control and drives the evolution of virus-specific activating Ly49 receptors.

CMVs efficiently target MHC I molecules to avoid recognition by cytotoxic T cells. However, the lack of MHC I on the cell surface renders the infected cell susceptible to NK cell killing upon missing self recognition. To counter this, mouse CMV (MCMV) rescues some MHC I molecules to engage inhibitory Ly49 receptors. Here we identify a new viral protein, MATp1, that is essential for MHC I surface rescue. Rescued altered-self MHC I molecules show increased affinity to inhibitory Ly49 receptors, resulting in inhibition of NK cells despite substantially reduced MHC I surface levels. This enables the virus to evade recognition by licensed NK cells. During evolution, this novel viral immune evasion mechanism could have prompted the development of activating NK cell receptors that are specific for MATp1-modified altered-self MHC I molecules. Our study solves a long-standing conundrum of how MCMV avoids recognition by NK cells, unravels a fundamental new viral immune evasion mechanism, and demonstrates how this forced the evolution of virus-specific activating MHC I-restricted Ly49 receptors.

Distinct Surface Expression of Activating Receptor Ly49H Drives Differential Expansion of NK Cell Clones upon Murine Cytomegalovirus Infection

Natural killer (NK) cells show some features of adaptive immunity but have not been studied at the clonal level. Here, we used retrogenic color-barcoding and single-cell adoptive transfers to track clonal immune responses to murine cytomegalovirus (MCMV) infection, derived from individual NK cells expressing activating receptor Ly49H. Clonal expansion of single NK cells varied substantially, and this variation could not be attributed to the additional presence or absence of inhibitory Ly49 receptors. Instead, single-cell-derived variability correlated with distinct surface expression levels of Ly49H itself. Ly49Hhi NK cell clones maintained higher Ly49H expression and expanded more than their Ly49Hlo counterparts in response to MCMV. Thus, akin to adaptive processes shaping an antigen-specific Tcell receptor (TCR) repertoire, the Ly49H+ NK cell population adapts to MCMV infection. This process relies on the clonal maintenance of distinct Ly49H expression levels, generating a repertoire of individual NK cells outfitted with distinct reactivity to MCMV.

Predicting Natural Killer Cell Behavior with Mathematical Models

Abstract Natural killer (NK) cells are specialized lymphocytes with an innate ability to eliminate virally infected and cancerous cells, but the mechanisms that control NK cell development and cytotoxicity are incompletely understood. We hypothesize mathematical models can be used to predict how to direct NK cell development toward mature NK cells. We observed Sostdc1-knockout (KO) mice to display a partial NK cell developmental block. Building from these studies, a best fit model was simulated in which rates of transition and differentiation and cell death was determined by ordinary differential equations of deterministic compartmental models. This approach indicated that NK cell proliferation rates are not necessary to predict WT and KO population outcomes. We further hypothesize that clusters of highly cytotoxic NK cells can be identified computationally, using a combination of Ly49 receptor expression, in vitro and in vivo cytotoxicity assays using β2m−/− targets, and viSNE analysis of flow cytometry data. Sostdc1-KO NK cells also display defective cytotoxicity. We identified that KO splenic CD27+CD11b+ transitional NK cells express lower frequencies of inhibitory Ly49G2, but higher frequencies of activating Ly49H+ and D+ cells. Our viSNE results identified 3 NK cell clusters displaying a “superactive” phenotype that were virtually absent in KO mice; consistent with our observations of their hyporesponsiveness. Taken together, these data support a role for Sostdc1 in the regulation of NK cells and could provide insights into novel biological parameters to expand active NK cell numbers with high killing efficiency for immunotherapies.

Sostdc1 Regulates NK Cell Maturation and Cytotoxicity.

NK cells are innate-like lymphocytes that eliminate virally infected and cancerous cells, but the mechanisms that control NK cell development and cytotoxicity are incompletely understood. We identified roles for sclerostin domain-containing-1 (Sostdc1) in NK cell development and function. Sostdc1-knockout (Sostdc1 -/-) mice display a progressive accumulation of transitional NK cells (tNKs) (CD27+CD11b+) with age, indicating a partial developmental block. The NK cell Ly49 repertoire in Sostdc1 -/- mice is also changed. Lower frequencies of Sostdc1 -/- splenic tNKs express inhibitory Ly49G2 receptors, but higher frequencies express activating Ly49H and Ly49D receptors. However, the frequencies of Ly49I+, G2+, H+, and D+ populations were universally decreased at the most mature (CD27-CD11b+) stage. We hypothesized that the Ly49 repertoire in Sostdc1 -/- mice would correlate with NK killing ability and observed that Sostdc1-/- NK cells are hyporesponsive against MHC class I-deficient cell targets in vitro and in vivo, despite higher CD107a surface levels and similar IFN-γ expression to controls. Consistent with Sostdc1's known role in Wnt signaling regulation, Tcf7 and Lef1 levels were higher in Sostdc1 -/- NK cells. Expression of the NK development gene Id2 was decreased in Sostdc1-/- immature NK and tNK cells, but Eomes and Tbx21 expression was unaffected. Reciprocal bone marrow transplant experiments showed that Sostdc1 regulates NK cell maturation and expression of Ly49 receptors in a cell-extrinsic fashion from both nonhematopoietic and hematopoietic sources. Taken together, these data support a role for Sostdc1 in the regulation of NK cell maturation and cytotoxicity, and identify potential NK cell niches.

NKR-P1B expression in gut-associated innate lymphoid cells is required for the control of gastrointestinal tract infections.

Helper-type innate lymphoid cells (ILC) play an important role in intestinal homeostasis. Members of the NKR-P1 gene family are expressed in various innate immune cells, including natural killer (NK) cells, and their cognate Clr ligand family members are expressed in various specialized tissues, including the intestinal epithelium, where they may play an important role in mucosal-associated innate immune responses. In this study, we show that the inhibitory NKR-P1B receptor, but not the Ly49 receptor, is expressed in gut-resident NK cells, ILC, and a subset of γδT cells in a tissue-specific manner. ILC3 cells constitute the predominant cell subset expressing NKR-P1B in the gut lamina propria. The known NKR-P1B ligand Clr-b is broadly expressed in gut-associated cells of hematopoietic origin. The genetic deletion of NKR-P1B results in a higher frequency and number of ILC3 and γδT cells in the gut lamina propria. However, the function of gut-resident ILC3, NK, and γδT cells in NKR-P1B-deficient mice is impaired during gastrointestinal tract infection by Citrobacter rodentium or Salmonella typhimurium, resulting in increased systemic bacterial dissemination in NKR-P1B-deficient mice. Our findings highlight the role of the NKR-P1B:Clr-b recognition system in the modulation of intestinal innate immune cell functions.

Critical role for the Ly49 family of class I MHC receptors in adaptive natural killer cell responses

Adaptive natural killer (NK) cell memory represents a new frontier in immunology. Work over the last decade has discovered and confirmed the existence of NK cells with antigen-specific memories, which had previously been considered a unique property of T and B cells. These findings have shown that antigen-specific NK cells gain their specificity without the use of RAG proteins, representing a novel mechanism for generating antigen specificity, but the details of this mechanism have remained a mystery. We have discovered that members of the Ly49 family of surface receptors are critically involved in both the sensitization and the challenge phases of an NK cell memory response, as is antigen presentation from their binding partner, the class I MHC. Moreover, we demonstrate that the Ly49-interacting component of a presented antigen dictates the specificity of the NK cell memory response, implicating Ly49 receptors themselves in antigen-specific recognition. Finally, we demonstrate that adaptive NK cell memories can protect against an otherwise lethal melanoma without T cell or B cell support. These findings offer insight into the mechanism behind NK cell antigen specificity and demonstrate the clinical potential of this adaptive immune cell.