Alternative mechanisms of natural killer cell activation during herpes simplex virus infection

Although NK cells can kill both malignant cells and virus-infected cells without prior sensitization, it has remained unclear whether the mechanism by which an NK cell is activated in the presence of a tumor cell is similar to that induced by the presence of a virus-infected cell. In our experimental system using homogeneous populations of cloned human CD16+ NK cells, we found that HSV-infected target cells do not induce in the NK cells the same pharmacologically-active second messengers elicited by NK-sensitive tumor cells. Although phosphoinositide turnover and calcium signaling were generated in NK cells exposed to NK-sensitive tumor cells, the recognition of HSV-infected cells by NK cells did not result in similar transmembrane signaling. Furthermore, depending on the cell type infected by HSV, alternative mechanisms of cytotoxicity were employed. HSV-infected foreskin fibroblasts were rapidly and selectively killed by cloned NK cells without a requirement for IFN or accessory cells. In contrast to this direct cytotoxicity against HSV-infected foreskin fibroblasts, NK cell-mediated cytotoxicity against an HSV-infected fibrosarcoma cell line (1591) was dependent on IFN-alpha production by accessory cells. Importantly, in both systems of cytotoxicity, IFN-alpha activation of NK cells resulted in augmented killing against both infected and uninfected targets. These results suggest that NK cell activation induced during antiviral immunity is distinct from activation elicited during an antitumor response. These differences include the utilization of alternative forms of signal transduction and alternative mechanisms of cytotoxicity.     

Human cytomegalovirus strain-dependent changes in NK cell recognition of infected fibroblasts

NK cells play a key role in the control of CMV infection in mice, but the mechanism by which NK cells can recognize and kill CMV-infected cells is unclear. In this study, the modulation of NK cell susceptibility of human CMV (hCMV)-infected cells was examined. We used a human lung and a human foreskin fibroblast cell line infected with clinical isolates (4636, 13B, or 109B) or with laboratory strains (AD169, Towne). The results indicate that all three hCMV clinical isolates confer a strong NK resistance, whereas only marginal or variable effects in the NK recognition were found when the laboratory strains were used. The same results were obtained regardless of the conditions of infection, effector cell activation status, cell culture conditions, and/or donor-target cell combinations. The NK cell inhibition did not correlate with HLA class I expression levels on the surface of the target cell and was independent of the leukocyte Ig-like receptor-1, as evaluated in Ab blocking experiments. No relevant changes were detected in the adhesion molecules ICAM-I and LFA-3 expressed on the cell surface of cells infected with hCMV clinical and laboratory strains. We conclude that hCMV possesses other mechanisms, related neither to target cell expression of HLA-I or adhesion molecules nor to NK cell expression of leukocyte Ig-like receptor-1, that confer resistance to NK cell recognition. Such mechanisms may be lost during in vitro passage of the virus. These results emphasize the differences between clinical hCMV isolates compared with laboratory strains.     

Unexpected role for granzyme K in CD56bright NK cell-mediated immunoregulation of multiple sclerosis

Functional NK cell deficiencies are associated with autoimmune diseases, including multiple sclerosis. NK cells can promote or inhibit adaptive immunity via either cytokine production or cytotoxicity toward immature dendritic cells and activated T cells. In humans, this immunoregulatory role resides in the CD56(bright) NK cell subset, which is selectively expanded by daclizumab, a CD25-blocking Ab that suppresses multiple sclerosis-associated inflammation. The objective of this study was to investigate the molecular mechanisms underlying the cytotoxicity of NK cells toward activated T cells. We demonstrated that NK cells induce caspase-independent apoptosis that requires NK cell degranulation and causes mitochondrial dysfunction in activated T cells. Although both granzyme A and granzyme K (GrK) can mediate this form of apoptosis, quantitatively we observed preferential transfer of GrK to target cells. Consequently, gene silencing of GrK in the NK-92 cell line, which retains functional characteristics of CD56(bright) NK cells, profoundly inhibited the ability of NK-92 cells to kill activated syngeneic T cells. Finally, we demonstrated that daclizumab treatment significantly enhanced this newly defined mechanism of cytotoxicity by CD56(bright) NK cells. Our study describes the important physiological role that GrK plays in immunoregulation of adaptive immunity in humans and indicates that therapeutic exploitation of this pathway is beneficial in controlling autoimmunity.     

Potential role of NK cells in future anti-tumor immunotherapies

Natural Killer (NK) cells are no longer considered as relatively unimportant bystander cells having the capacity to kill certain tumor and virus-infected cells in a mysterious way. During the last decade a significant progress has been made in understanding biology of NK cells in particular their mechanisms of recognition and killing of target cells. This progress has led to novel knowledge-based clinical applications of NK cells as immunotherapeutic tools in various disease settings, especially in bone marrow transplantation for leukemia patients. The potential of NK cell therapy for eradicating solid tumors has not been fully exploited. In this mini-review, we examine the rationale behind these therapies and discuss the problems confronting researchers in their usage as therapeutic agents.     

自然杀伤细胞受体及其配体表达的转录和表观遗传调控

自然杀伤(natural killer cell,NK)细胞受体及其配体在NK细胞发挥抗病毒、抗肿瘤和免疫调节作用中起重要作用.NK细胞功能的发挥取决于NK细胞受体及其配体的表达水平和其所传递信号的综合.病毒、肿瘤和热休克等刺激可以通过激活相应的转录调节因子,提高启动子活性而上调NKG2家族受体及其配体的表达,而启动子区DNA的甲基化状态、组蛋白的乙酰化和甲基化等表观遗传调控,在NK细胞受体及其配体的表达方面亦起重要作用,并决定NK细胞受体的克隆性分布.深入探讨NK细胞受体及其配体的表达调控机制,将为提高NK.细胞抗肿瘤和抗感染疗效提供新的策略.     

Immunoproteasome-deficiency has no effects on NK cell education, but confers lymphocytes into targets for NK cells in infected wild-type mice

Natural killer (NK) cells are part of the innate immune system and contribute to the eradication of virus infected cells and tumors. NK cells express inhibitory and activating receptors and their decision to kill a target cell is based on the balance of signals received through these receptors. MHC class I molecules are recognized by inhibitory receptors, and their presence during NK cell education influences the responsiveness of peripheral NK cells. We here demonstrate that mice with reduced MHC class I cell surface expression, due to deficiency of immunoproteasomes, have responsive NK cells in the periphery, indicating that the lower MHC class I levels do not alter NK cell education. Following adoptive transfer into wild-type (wt) recipients, immunoproteasome-deficient splenocytes are tolerated in naive but rejected in virus-infected recipients, in an NK cell dependent fashion. These results indicate that the relatively low MHC class I levels are sufficient to protect these cells from rejection by wt NK cells, but that this tolerance is broken in infection, inducing an NK cell-dependent rejection of immunoproteasome-deficient cells.     

HSV-2 Specifically Down Regulates HLA-C Expression to Render HSV-2-Infected DCs Susceptible to NK Cell Killing

Both NK cells and CTLs kill virus-infected and tumor cells. However, the ways by which these killer cells recognize the infected or the tumorigenic cells are different, in fact almost opposite. CTLs are activated through the interaction of the TCR with MHC class I proteins. In contrast, NK cells are inhibited by MHC class I molecules. The inhibitory NK receptors recognize mainly MHC class I proteins and in this regard practically all of the HLA-C proteins are recognized by inhibitory NK cell receptors, while only certain HLA-A and HLA-B proteins interact with these receptors. Sophisticated viruses developed mechanisms to avoid the attack of both NK cells and CTLs through, for example, down regulation of HLA-A and HLA-B molecules to avoid CTL recognition, leaving HLA-C proteins on the cell surface to inhibit NK cell response. Here we provide the first example of a virus that through specific down regulation of HLA-C, harness the NK cells for its own benefit. We initially demonstrated that none of the tested HSV-2 derived microRNAs affect NK cell activity. Then we show that surprisingly upon HSV-2 infection, HLA-C proteins are specifically down regulated, rendering the infected cells susceptible to NK cell attack. We identified a motif in the tail of HLA-C that is responsible for the HSV-2-meduiated HLA-C down regulation and we show that the HLA-C down regulation is mediated by the viral protein ICP47. Finally we show that HLA-C proteins are down regulated from the surface of HSV-2 infected dendritic cells (DCs) and that this leads to the killing of DC by NK cells. Thus, we propose that HSV-2 had developed this unique and surprising NK cell-mediated killing strategy of infected DC to prevent the activation of the adaptive immunity.     

Advantages and clinical applications of natural killer cells in cancer immunotherapy

The past decade has witnessed a burgeoning of research and further insight into the biology and clinical applications of natural killer (NK) cells. Once thought to be simple innate cells important only as cytotoxic effector cells, our understanding of NK cells has grown to include memory-like responses, the guidance of adaptive responses, tissue repair, and a delicate paradigm for how NK cells become activated now termed “licensing” or “arming.” Although these cells were initially discovered and named for their spontaneous ability to kill tumor cells, manipulating NK cells in therapeutic settings has proved difficult and complex in part due to our emerging understanding of their biology. Therapies involving NK cells may either activate endogenous NK cells or involve transfers of exogenous cells by hematopoietic stem cell transplantation or adoptive cell therapy. Here, we review the basic biology of NK cells, highlighting characteristics which make NK cells particularly useful in cancer therapies. We also explore current treatment strategies that have been used for cancer as well as discuss potential future directions for the field.     

NK cells use perforin rather than granulysin for anticryptococcal activity

Cytotoxic lymphocytes have the capacity to kill microbes directly; however, the mechanisms involved are poorly understood. Using Cryptococcus neoformans, which causes a potentially fatal fungal infection in HIV-infected patients, our previous studies showed that granulysin is necessary, while perforin is dispensable, for CD8 T lymphocyte fungal killing. By contrast, the mechanisms by which NK cells exert their antimicrobial activity are not clear, and in particular, the contribution of granulysin and perforin to NK-mediated antifungal activity is unknown. Primary human NK cells and a human NK cell line YT were found to constitutively express granulysin and perforin, and possessed anticryptococcal activity, in contrast to CD8 T lymphocytes, which required stimulation. When granulysin protein and mRNA were blocked by granulysin small interfering RNA, the NK cell-mediated antifungal effect was not affected in contrast to the abrogated activity observed in CD8 T lymphocytes. However, when perforin was inhibited by concanamycin A, and silenced using hairpin small interfering RNA, the anticryptococcal activities of NK cells were abrogated. Furthermore, when granulysin and perforin were both inhibited, the anticryptococcal activities of the NK cells were not reduced further than by silencing perforin alone. These results indicate that the antifungal activity is constitutively expressed in NK cells in contrast to CD8 T lymphocytes, in which it requires prior activation, and perforin, but not granulysin, plays the dominant role in NK cell anticryptococcal activity, in contrast to CD8 T lymphocytes, in which granulysin, but not perforin, plays the dominant role in anticryptococcal activity.     

Natural killer cells kill extracellular Pseudomonas aeruginosa using contact-dependent release of granzymes B and H

Pseudomonas aeruginosa is an opportunistic pathogen that often infects individuals with the genetic disease cystic fibrosis, and contributes to airway blockage and loss of lung function. Natural killer (NK) cells are cytotoxic, granular lymphocytes that are part of the innate immune system. NK cell secretory granules contain the cytolytic proteins granulysin, perforin and granzymes. In addition to their cytotoxic effects on cancer and virally infected cells, NK cells have been shown to play a role in an innate defense against microbes, including bacteria. However, it is not known if NK cells kill extracellular P. aeruginosa or how bacterial killing might occur at the molecular level. Here we show that NK cells directly kill extracellular P. aeruginosa using NK effector molecules. Live cell imaging of a co-culture of YT cells, a human NK cell line, and GFP-expressing P. aeruginosa in the presence of the viability dye propidium iodide demonstrated that YT cell killing of P. aeruginosa is contact-dependent. CRISPR knockout of granulysin or perforin in YT cells had no significant effect on YT cell killing of P. aeruginosa. Pre-treatment of YT and NK cells with the serine protease inhibitor 3,4-dichloroisocoumarin (DCI) to inhibit all granzymes, resulted in an inhibition of killing. Although singular CRISPR knockout of granzyme B or H had no effect, knockout of both in YT cells completely abrogated killing of P. aeruginosa in comparison to wild type YT cell controls. Nitrocefin assays suggest that the bacterial membrane is damaged. Inhibition of killing by antioxidants suggest that ROS are required for the bactericidal mode-of-action. Taken together, these results identify that NK cells kill P. aeruginosa through a membrane damaging, contact-dependent process that requires granzyme induced ROS production, and moreover, that granzyme B and H are redundant in this killing process.     

Cell surface glycoconjugates as possible target structures for human natural killer cells: evidence against the involvement of glycolipids and N-linked carbohydrate chains

Natural killer (NK) cells can spontaneously kill various malignantcells, but the susceptibility towards NK cells differs greatlyamong different types of tumour cells. The molecules, whichare recognized by NK cells, have not yet been identified, butthere is ample evidence that target cell surface glycoconjugatesare involved in the interaction with NK cells. In this report,we show that the recognition of K562 target cells by human NKcells depends on the presence of protein-bound determinants,implying that glycolipids are not the primary target structureson K562 cells. The NK susceptibility of K562 cells was not alteredby enzymic removal of various cell surface carbohydrates oroligosaccharides, mostly related to N-linked carbohydrate chains.Treatment of K562 cells with 1-deoxynojirimycin and 1-deoxymannojirimycin,inhibitors of N-glycan processing, resulted in drastic alterationsin the carbohydrate phenotype of the cell surface, as couldbe shown by flow cytometric analysis of the lectin-binding propertiesof the cells. Despite these clear changes in N-glycosylation,the NK susceptibility of K562 cells remained unaffected. Summarizing,the results described in this report show that potential targetstructures for NK cells are protein bound, but the involvementof a specific (N-linked) carbohydrate determinant in the interactionbetween NK cells and target cells could not be established. cell adhesion molecules cell—cell interaction cell surface glycoconjugates natural killer cells target structures     

Lysis of endogenously infected CD4+ T cell blasts by rIL-2 activated autologous natural killer cells from HIV-infected viremic individuals

Understanding the cellular mechanisms that ensure an appropriate innate immune response against viral pathogens is an important challenge of biomedical research. In vitro studies have shown that natural killer (NK) cells purified from healthy donors can kill heterologous cell lines or autologous CD4+ T cell blasts exogenously infected with several strains of HIV-1. However, it is not known whether the deleterious effects of high HIV-1 viremia interferes with the NK cell-mediated cytolysis of autologous, endogenously HIV-1-infected CD4+ T cells. Here, we stimulate primary CD4+ T cells, purified ex vivo from HIV-1-infected viremic patients, with PHA and rIL2 (with or without rIL-7). This experimental procedure allows for the significant expansion and isolation of endogenously infected CD4+ T cell blasts detected by intracellular staining of p24 HIV-1 core antigen. We show that, subsequent to the selective down-modulation of MHC class-I (MHC-I) molecules, HIV-1-infected p24(pos) blasts become partially susceptible to lysis by rIL-2-activated NK cells, while uninfected p24(neg) blasts are spared from killing. This NK cell-mediated killing occurs mainly through the NKG2D activation pathway. However, the degree of NK cell cytolytic activity against autologous, endogenously HIV-1-infected CD4+ T cell blasts that down-modulate HLA-A and -B alleles and against heterologous MHC-I(neg) cell lines is particularly low. This phenomenon is associated with the defective surface expression and engagement of natural cytotoxicity receptors (NCRs) and with the high frequency of the anergic CD56(neg)/CD16(pos) subsets of highly dysfunctional NK cells from HIV-1-infected viremic patients. Collectively, our data demonstrate that the chronic viral replication of HIV-1 in infected individuals results in several phenotypic and functional aberrancies that interfere with the NK cell-mediated killing of autologous p24(pos) blasts derived from primary T cells.     

Natural killer cells and malignant haemopathies: a model for the interaction of cancer with innate immunity

Despite recent progress in the therapeutic approach of malignant haemopathies, their prognoses remain frequently poor. Immunotherapy offers an alternative of great interest in this context but defect or abnormal expression of human leukocyte antigens (HLA), frequently observed in cancer cells, limits its efficiency. Natural killer (NK) cells, which are able to kill target cells in a HLA-independent way, represent a novel tool in the treatment of haematological malignancies. Abnormal NK cytolytic function is observed in all the haematological malignancies studied, such as acute leukaemia, myelodysplastic syndromes or chronic myeloid/lymphoid leukaemia. Several mechanisms are involved in the alterations of NK cytotoxicity: decreased expression of activating receptors, increased expression of inhibitory receptors or defective expression of NK ligands on target cells. Further studies are needed to identify how each type of haematological malignancy escapes from the innate immune response. Attempts to increase the expression of activating receptors, to counteract inhibitory receptors expression, or to increase NK cell cytotoxic capacities could overcome tumour escape from innate immunity. These therapies are based on monoclonal antibodies or culture of NK cells in presence of cytokines or dendritic cells. Moreover, many novel drugs used in haematological malignancies [tyrosine kinase inhibitors, IMIDs^®, proteasome inhibitors, demethylating agents, histone deacetylase inhibitors (HDACis), histamine dihydrochloride] display interesting immunomodulatory properties that affect NK cells. These data suggest that combined modalities associating cytotoxic drugs with innate immunity modulators may represent a major breakthrough in tumour eradication.     

In vitro generation of human activated lymphocyte killer cells: separate precursors and modes of generation of NK-like cells and "anomalous" killer cells

The activation of human peripheral blood mononuclear cells (PBM) in culture leads to the generation of nonspecific killer cells. These cells, termed activated lymphocyte killer (ALK) cells, can kill fresh tumor cells and tumor cell lines, in addition to the natural killer (NK) cell sensitive target K562. ALK cells have features in common with both T and NK cells, but their nature and origin are unknown. In the present study, it is shown that ALK cells are in fact heterogeneous and can be generated from both large granular lymphocytes with the same phenotype as NK cells and from T cells. Cell populations enriched for NK cells, when cultured with lymphokines, rapidly acquired a T cell phenotype, enhanced cytolytic activity against K562, and the ability to lyse NK-insensitive target cells such as a melanoma cell line LiBr; these ALK cells were described as NK-like cells. On the other hand, of the cloned cells derived from PBM stimulated with irradiated B lymphoblasts and grown in lymphokines, the major proportion of cytolytic T cells (CTC) able to kill the specific stimulator lymphoblasts were also found to kill LiBr but not K562 cells. These ALK cells, which were derived from the same precursors as CTC, were designated anomalous killer (AK) cells. Consistent with this, the presence of the pan T monoclonal antibody UCHT1 from the beginning of mixed cell cultures inhibited the generation of CTC and of the AK-type of ALK cell, which killed melanoma cells, but not the NK type, which killed K562 targets. By contrast, at the effector cell level, the antibodies UCHT1 and OKT8 only blocked specific killing by CTC but did not block the killing of LiBr or of K562 targets by ALK cells. However, at the effector cell level there was additional evidence for the heterogeneity of ALK cells. Thus, monoclonal antibody 9.1C3, which blocks killing by freshly isolated NK cells, also blocked the killing of K562 targets by NK-like cells, but did not block B lymphoblast killing by CTC or melanoma cell killing by AK cells. It is concluded that after mixed lymphocyte culture, the majority of ALK cells measured by the killing of melanoma target cells arise from the same precursors and are under the same influences as classical CTC (AK cells), whereas cells killing K562 targets are derived from NK cells (NK-like cells). Once generated, the AK cells have a different mechanism of killing from both classical CTC and from NK and NK-like cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

Human killer cells and natural killer cells: distinct subpopulations of Fc receptor-bearing lymphocytes

Unstimulated human peripheral blood lymphocytes were depleted of K cells, which mediate antibody-dependent cellular cytotoxicity (ADCC) without removing NK cells, which mediate natural killing (NK). K cell depletion was achieved by buoyant centrifugation removal of lymphocytes that bound to glutaraldehyde-treated P815-AB cells at high lymphocyte-to-target ratios. Likewise, NK cells were removed with glutaraldehyde-treated K562 cells without removing K cells. Furthermore, both cytotoxic cell populations were observed directly in one agarose single-cell cytotoxic assay (ASCA) using P815-AB and K562 cells simultaneously as target cells. Moreover, the percentage of total cytotoxic cells was equal to the sum of the percentage of K and NK cells observed in separate ASCA. Collectively, these results indicate that K cells and NK cells are distinct subsets of FcR-bearing lymphocytes. One subset, K cells, has more avid Fc receptors (fcR) than NK cells and are 'activated' via thier FcR to kill antibody-coated target cells. The second subset, NK cells, have less avid FcR and are not 'activated' through their FcR to kill antibody-coated target cells.     

NK1.1+ thymocytes. Adult murine CD4-, CD8- thymocytes contain an NK1.1+, CD3+, CD5hi, CD44hi, TCR-V beta 8+ subset.

CD4-, CD8- thymocytes were purified from thymi obtained from normal C57BL/6 mice. By flow cytometry analysis, 5 to 10% of these double negative (DN) thymocytes were found to express NK1.1 on their surface. The NK1.1+ DN thymocytes were demonstrated, by two-color fluorescence, to be CD3lo, CD5hi, CD44hi, J11d-, B220-, MEL 14-, IL2R- with 60% expressing TCR-V beta 8 as determined by the mAb F23.1. In contrast, splenic and peripheral blood NK cells were NK1.1+, CD3-, CD5-, TCR-V beta 8- with 40 to 60% being MEL 14+. Unlike peripheral NK cells, fresh DN thymocytes enriched for NK1.1+ cells were unable to kill YAC-1, the classical murine NK cell target. However, these cells were able to mediate anti-CD3 redirected lysis even when they were assayed immediately after purification, i.e., with no culture or stimulation. These data demonstrate that adult murine thymocytes contain NK1.1+ cells which are distinct, both by function and phenotype, from peripheral NK cells. These data also raise the issue of a possible NK/T bipotential progenitor cell.     

Heat shock protein 70-reactivity is associated with increased cell surface density of CD94/CD56 on primary natural killer cells

Previously we described an involvement of the C-type lectin receptor CD94 and the neuronal adhesion molecule CD56 in the interaction of natural killer (NK) cells with Hsp70-protein and Hsp70-peptide TKD. Therefore, differences in the cell surface density of these NK cell-specific markers were investigated comparatively in CD94-sorted, primary NK cells and in established NK cell lines NK-92, NKL, and YT after TKD stimulation. Initially, all NK cell types were positive for CD94; the CD56 expression varied. After stimulation with TKD, the mean fluorescence intensity (mfi) of CD94 and CD56 was upregulated selectively in primary NK cells but not in NK cell lines. Other cell surface markers including natural cytotoxicity receptors remained unaffected in all cell types. CD3-enriched T cells neither expressing CD94 nor CD56 served as a negative control. High receptor densities of CD94/CD56 were associated with an increased cytolytic response against Hsp70 membrane-positive tumor target cells. The major histocompatibility complex (MHC) class I-negative, Hsp70-positive target cell line K562 was efficiently lysed by primary NK cells and to a lower extent by NK lines NK-92 and NKL. YT and CD3-positive T cells were unable to kill K562 cells. MHC class-I and Hsp70-positive, Cx + tumor target cells were efficiently lysed only by CD94-sorted, TKD-stimulated NK cells with high CD94/CD56 mfi values. Hsp70-specificity was demonstrated by antibody blocking assays, comparative phenotyping of the tumor target cells, and by correlating the amount of membrane-bound Hsp70 with the sensitivity to lysis. Remarkably, a 14-mer peptide (LKD), exhibiting only 1 amino acid exchange at position 1 (T to L), neither stimulated Hsp70-reactivity nor resulted in an upregulated CD94 expression on primary NK cells. Taken together our findings indicate that an MHC class I-independent, Hsp70 reactivity could be associated with elevated cell surface densities of CD94 and CD56 after TKD stimulation.     

Interferon-producing killer dendritic cells provide a link between innate and adaptive immunity

Natural killer (NK) cells and dendritic cells (DCs) are, respectively, central components of innate and adaptive immune responses. We describe here a third DC lineage, termed interferon-producing killer DCs (IKDCs), distinct from conventional DCs and plasmacytoid DCs and with the molecular expression profile of both NK cells and DCs. They produce substantial amounts of type I interferons (IFN) and interleukin (IL)-12 or IFN-gamma, depending on activation stimuli. Upon stimulation with CpG oligodeoxynucleotides, ligands for Toll-like receptor (TLR)-9, IKDCs kill typical NK target cells using NK-activating receptors. Their cytolytic capacity subsequently diminishes, associated with the loss of NKG2D receptor (also known as Klrk1) and its adaptors, Dap10 and Dap12. As cytotoxicity is lost, DC-like antigen-presenting activity is gained, associated with upregulation of surface major histocompatibility complex class II (MHC II) and costimulatory molecules, which formally distinguish them from classical NK cells. In vivo, splenic IKDCs preferentially show NK function and, upon systemic infection, migrate to lymph nodes, where they primarily show antigen-presenting cell activity. By virtue of their capacity to kill target cells, followed by antigen presentation, IKDCs provide a link between innate and adaptive immunity.