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HLAs, TCRs, and KIRs, a Triumvirate of Human Cell-Mediated Immunity
Abstract
In all human cells, human leukocyte antigen (HLA) class I glycoproteins assemble with a peptide and take it to the cell surface for surveillance by lymphocytes. These include natural killer (NK) cells and γδ T cells of innate immunity and αβ T cells of adaptive immunity. In healthy cells, the presented peptides derive from human proteins, to which lymphocytes are tolerant. In pathogen-infected cells, HLA class I expression is perturbed. Reduced HLA class I expression is detected by KIR and CD94:NKG2A receptors of NK cells. Almost any change in peptide presentation can be detected by αβ CD8 ⁺ T cells. In responding to extracellular pathogens, HLA class II glycoproteins, expressed by specialized antigen-presenting cells, present peptides to αβ CD4 ⁺ T cells. In comparison to the families of major histocompatibility complex (MHC) class I, MHC class II and αβ T cell receptors, the antigenic specificity of the γδ T cell receptors is incompletely understood.
... Natural killer (NK) cells are innate lymphocytes that develop in the bone marrow from common lymphoid progenitor cells. NK-cell maturity and function are critically dependent upon the transcription factors T-BET and EOMES [1] and following maturation they express a diverse repertoire of germline-encoded, non-rearranged surface receptors [2,3]. This diversity in the expression of activating and inhibitory receptors allows for the recognition of infected or tumour cells through complementary mechanisms in a combinatorial manner. ...
... In addition, the diverse family of inhibitory killer cell immunoglobulin-like receptors (KIR) predominantly engage with HLA-A, -B, and -C alleles. Thus, the loss of HLA class I ligands for inhibitory receptors such as NKG2A or the inhibitory KIR proteins on cancerous cells leads to loss-of-self inhibitory signals and NK-cell activation [2]. The interaction of KIR and NKG2A with their ligands on healthy cells is also necessary for the production of functionally competent NK cells (termed NK-cell education). ...
... There are many activating receptors expressed by NK cells, and some examples of relevance to cancer immunotherapy include NKp46, NKp30, DNAM-1 (CD226), 2B4 (CD244), activating KIR, NKG2D, NKG2C, and NKp44 [2,3,13,14]. In cancer, the ligands for NKG2D and NKp46 are stressinduced ligands such as MICA/B, ULBPs, and ecto-calreticulin [3,15]. ...
Natural killer (NK) cells are cytotoxic innate lymphoid cells that participate in anti-tumour and anti-viral immune responses. Their ability to rapidly destroy abnormal cells and to enhance the anti-cancer function of dendritic cells, CD8+ T cells and macrophages makes them an attractive target for immunotherapeutic strategies. The development of approaches which augment NK cell activation against cancer is currently under intense preclinical and clinical research and strategies include chimeric antigen receptor (CAR) NK cells, NK cell engagers, cytokines, and immune checkpoint inhibitors. In this review, we highlight recent advances in NK cell therapeutic development and discuss their potential to add to our armamentarium against cancer.
... In mammals, the major histocompatibility complex (MHC) is a large genomic region with much recombination and with hundreds of genes, among which are a few that encode highly polymorphic transplantation antigens, now known as classical class I and class II molecules (Trowsdale and Knight 2013). The classical class I molecules bind peptides which originate primarily from proteins in the cytoplasm and nucleus where viruses (and a few cytoplasmic bacteria) replicate and present them at the cell surface to CD8-bearing cytotoxic T lymphocytes (CTLs) as well as natural killer (NK) cells, which can kill infected cells acting as viral factories (as well as cells with other differences from normal self, including allogeneic cells in transplants) (Blum et al. 2013;Parham and Moffett 2013;Djaoud and Parham 2020). ...
... In humans, there are three classical class I loci, each with thousands of alleles. HLA-A and HLA-B are the primary CTL ligands and are well expressed on most cells (but not on red blood cells), although a few HLA-A alleles and about half of HLA-B alleles can be recognised by NK cells (Trowsdale and Knight 2013;Parham and Moffett 2013;Djaoud and Parham 2020). HLA-C alleles are less well expressed on most cells (although they are very well expressed on placental trophoblasts, but for tissue remodelling rather than as cytotoxic targets) and all alleles act primarily as NK ligands (Trowsdale and Knight 2013;Parham and Moffett 2013;Djaoud and Parham 2020), although they can present peptides to CTLs (Das and Khakoo 2015;Saunders et al. 2015). ...
... HLA-A and HLA-B are the primary CTL ligands and are well expressed on most cells (but not on red blood cells), although a few HLA-A alleles and about half of HLA-B alleles can be recognised by NK cells (Trowsdale and Knight 2013;Parham and Moffett 2013;Djaoud and Parham 2020). HLA-C alleles are less well expressed on most cells (although they are very well expressed on placental trophoblasts, but for tissue remodelling rather than as cytotoxic targets) and all alleles act primarily as NK ligands (Trowsdale and Knight 2013;Parham and Moffett 2013;Djaoud and Parham 2020), although they can present peptides to CTLs (Das and Khakoo 2015;Saunders et al. 2015). ...
The chicken major histocompatibility complex (MHC, also known as the BF-BL region of the B locus) is notably small and simple with few genes, most of which are involved in antigen processing and presentation. There are two classical class I genes, of which only BF2 is well and systemically expressed as the major ligand for cytotoxic T lymphocytes (CTLs). The other class I gene, BF1, is believed to be primarily a natural killer (NK) cell ligand. Among most standard chicken MHC haplotypes examined in detail, BF1 is expressed tenfold less than BF2 at the RNA level due to defects in the promoter or in a splice site. However, in the B14 and typical B15 haplotypes, BF1 RNA was not detected, and here, we show that a deletion between imperfect 32 nucleotide direct repeats has removed the BF1 gene entirely. The phenotypic effects of not having a BF1 gene (particularly on resistance to infectious pathogens) have not been systematically explored, but such deletions between short direct repeats are also found in some BF1 promoters and in the 5′ untranslated region (5′UTR) of some BG genes found in the BG region of the B locus. Despite the opposite transcriptional orientation of homologous genes in the chicken MHC, which might prevent the loss of key genes from a minimal essential MHC, it appears that small direct repeats can still lead to deletion.
Supplementary Information
The online version contains supplementary material available at 10.1007/s00251-023-01313-9.
... Despite increased CD8 Treg numbers in populations with CD4-driven autoimmune diseases, the expansion of pathogenic CD4 T cells is not sufficiently inhibited to prevent pathology and/or disease progression. The lack of productive engagement with pathogenic CD4 T cells may be due partly to the presence of inhibitory receptors on the CD8 Treg surface that regulate their threshold for activation, including inhibitory KIRs, which engage major histocompatibility complex (MHC) class I on target cells (14). Inhibitory KIRs compete with the TCRs for MHC class I/peptide binding and send inhibitory signals that counteract activating stimuli (15). ...
... The association of genomic expression of KIRs with various autoimmune diseases (46,47), coupled with the expression of inhibitory KIRs by CD8 Tregs, suggests that KIRs may also regulate CD8 Treg functions. KIRs compete with TCRs for binding to the MHC class I/peptide complex, and they are autoimmune checkpoints initiating an inhibitory signaling cascade that counteracts activating stimuli (14). Based on collective observations by us and others in animal models and patients with autoimmune diseases, we hypothesized that CD8 Treg functions can be restored by reducing the inhibitory signals delivered by KIRs, leveraging the selective reduction of pathogenic CD4 T cells, and mitigating the risk of broad immune suppression or off-target toxicities. ...
Introduction
Regulatory CD8 T cells (CD8 Treg) are responsible for the selective killing of self-reactive and pathogenic CD4 T cells. In autoimmune disease, CD8 Treg may accumulate in the peripheral blood but fail to control the expansion of pathogenic CD4 T cells that subsequently cause tissue destruction. This CD8 Treg dysfunction is due in part to the expression of inhibitory killer immunoglobulin-like receptors (KIR; KIR2DL isoforms [KIR2DL1, KIR2DL2, and KIR2DL3]); these molecules serve as autoimmune checkpoints and limit CD8 Treg activation.
Methods
Here we describe the pre-clinical characterization of MTX-101, a bispecific antibody targeting inhibitory KIR and CD8. Using human peripheral blood mononuculear cells (PBMC) derived from healthy donors and autoimmune patients, humanized mouse models, and human derived tissue organoids, we evaluated the molecular mechanisms and functional effects of MTX-101.
Results
By binding to KIR, MTX-101 inhibited KIR signaling that can restore CD8 Treg ability to eliminate pathogenic CD4 T cells. MTX-101 bound and activated CD8 Treg in human peripheral blood mononuclear cells (PBMC), resulting in increased CD8 Treg cytolytic capacity, activation, and prevalence. Enhancing CD8 Treg function with MTX-101 reduced pathogenic CD4 T cell expansion and inflammation, without increasing pro-inflammatory cytokines or activating immune cells that express either target alone. MTX-101 reduced antigen induced epithelial cell death in disease affected tissues, including in tissue biopsies from individuals with autoimmune disease (i.e., celiac disease, Crohn’s disease). The effects of MTX-101 were specific to autoreactive CD4 T cells and did not suppress responses to viral and bacterial antigens. In a human PBMC engrafted Graft versus Host Disease (GvHD) mouse model of acute inflammation, MTX-101 bound CD8 Treg and delayed onset of disease. MTX-101 induced dose dependent binding, increased prevalence and cytolytic capacity of CD8 Treg, as well as increased CD4 T cell death. MTX-101 selectively bound CD8 Treg without unwanted immune cell activation or increase of pro-inflammatory serum cytokines and exhibited an antibody-like half-life in pharmacokinetic and exploratory tolerability studies performed using IL-15 transgenic humanized mice with engrafted human lymphocytes, including CD8 Treg at physiologic ratios.
Conclusion
Collectively, these data support the development of MTX-101 for the treatment of autoimmune diseases.
... Interestingly, the HLA-A23 and HLA-A24 serotypes contain the Bw4 epitope, an amino acid sequence in the α1 domain responsible for the interaction with the inhibitory KIR3DL1 receptor that is also present in other HLA-A and HLA-B serotypes. 32,33 Analysis of the frequencies of the Bw4 epitope within the different elder groups revealed an equal distribution (Suppl. Table 2). ...
... 46 Second, KIR3DL1 alleles have the potential to inhibit the effector activities of NK cells and CD8+ T cells that express it, after interaction with HLA class I molecules containing the Bw4 epitope that is present on many HLA-B and some HLA-A alleles, including HLA-A23 and HLA-A24. 32,33 Third, KIR3DL1 alleles have been associated with protection from PD symptoms, implying a role for KIR3DL1-expressing cells in delaying PD progression. 47 Thus, in line with what was referred in Subsection 4.2, it can be proposed that the presence of higher levels of sHLA class I molecules (i.e., sHLA-A23 and sHLA-A24) in plasma of cognitively impaired elderly subjects with dementia could be part of the immunomodulatory mechanisms elicited by the ongoing neurodegenerative process (Figure 4). ...
MHC class I molecules regulate brain development and plasticity in mice and HLA class I molecules are associated with brain disorders in humans. We investigated the relationship between plasma‐derived soluble human HLA class I molecules (sHLA class I), HLA class I serotypes and dementia. A cohort of HLA class I serotyped elderly subjects with no dementia/pre‐dementia (NpD, n = 28), or with dementia (D, n = 28) was studied. Multivariate analysis was used to examine the influence of dementia and HLA class I serotype on sHLA class I levels, and to compare sHLA class I within four groups according to the presence or absence of HLA‐A23/A24 and dementia. HLA‐A23/A24 and dementia, but not age, significantly influenced the level of sHLA class I. Importantly, the concurrent presence of HLA‐A23/A24 and dementia was associated with higher levels of sHLA class I (p < 0.001). This study has shown that the simultaneous presence of HLA‐A23/HLA‐A24 and dementia is associated with high levels of serum sHLA class I molecules. Thus, sHLA class I could be considered a biomarker of neurodegeneration in certain HLA class I carriers.
... 18,20,21 There are up to fteen KIR genes found on chromosome 19 that have activating (usually denoted by "S" for short tail, e.g., KIR2DS1 ) or inhibitory functions (usually denoted by "L" for long tail, e.g., KIR2DL5) for NK cells. 18,26,28,29 Most mature NK cells express KIR, although the total number of KIR genes observed and expressed as well as the type of KIR ligands produced by the genes (i.e., inhibitory and activating ligands) varies by individual. 18,20, 21 We have recently shown that the KIR genes KIR2DL5, KIR2DS5, and KIR2DS1 are more frequently found in individuals with AD. 22 This is also true for the HLA-C2 epitope and HLA-B*-21T leader sequence. ...
... This nding is consistent with previous tissue based reports. 11 Our study adds to the growing literature on the genetic basis of the immune dysregulation of AD and, speci cally, that KIR allelic variation is associated with an increased risk of AD. 20,27,29,61,62 Further a paradigm shift has occurred in understanding of the immunology of AD suggesting that NK de ciency may be real, may interplay with barrier dysfunction and that treatment studies speci cally examining NK induction and modulation are now warranted. 11 Speci cally, studies of NK cell immunologic function in those with AD may help explain disease variability, different disease immunologic endotypes, differences in genetic predilection for AD by race, as well as treatment failure. ...
Natural killer cells (NK) have been associated with the pathophysiology of atopic dermatitis (AD). NK function is regulated by killer cell Ig-like receptor family (KIR) receptors that interact with HLA ligands. The goal of this study was to focus on allelic variation in genes KIR2DL5 , KIR2DS5 , and KIR2DS1 with respect to AD. This was a case-control study of individuals with (n = 313) and without (n = 176) AD. Associations were estimated using logistic regression. Evaluations included interactions between KIR and known HLA ligand pairs. The prevalence of KIR2DL5 was 52.5% (95% CI: 48.0,57.0), KIR2DS5 was 33.0% (28.8,37.3), and KIR2DS1 was 33.6% (29.4,38.0). When compared to those who did not have KIR2DL5 , homozygote individuals for KIR2DL5*001:01 were more likely to have AD (OR: 2.16 (95% CI:1.31,3.53) p = 0.0023). The effect of KIR2DL5*001:01 was similar in Whites and Blacks. The alleles from the other KIR genes of interest were not associated with AD. There is no known HLA ligand for KIR2DL5 . However, the effect of KIR2DL5*001:01 increases in the presence of HLA-B *-21TT leader sequence (2.46(1.37,4.41) p = 0.0025) and HLA-C2 ligand (2.07 (1.37,4.41, p = 0.000002). This is the first study to explore KIR allelic variation in AD. KIR2DL5*001:01 allele is independently associated with an increased risk of AD.
... NKs are lymphoid innate immune cells and are the key effector cells in immunotherapy [100], which are related to the occurrence, progression and recurrence of AML [101]. The killer cell immunoglobulin-like receptor (KIR) expressed on the surface of NKs can be divided into two types, inhibitory and activating [102], which can specifically recognize and bind target cell surface molecules and play important antitumor roles [103]. NKs express different immunophenotypes at the immature, mature, and hypermature stages and can migrate, release cytokines, and destroy target cells [104]. ...
In recent years, the relationship between the immunosuppressive niche of the bone marrow and therapy resistance in acute myeloid leukemia (AML) has become a research focus. The abnormal number and function of immunosuppressive cells, including regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs), along with the dysfunction and exhaustion of immunological effector cells, including cytotoxic T lymphocytes (CTLs), dendritic cells (DCs) and natural killer cells (NKs), can induce immune escape of leukemia cells and are closely linked to therapy resistance in leukemia. This article reviews the research progress on the relationship between immune cells in the marrow microenvironment and chemoresistance in AML, aiming to provide new ideas for the immunotherapy of AML.
... A critical receptor family for controlling NK cell activation is the Killer-cell Immunoglobulin-like Receptor (KIR) family which is made up of activating and inhibitory isoforms (12,13). Ligation of inhibitory KIR with self-HLA prevents the killing of healthy host cells by NK cells (14) and HLA downregulation by malignantly transformed cells can unleash NK cell cytotoxicity, a paradigm known as missing-self (15). ...
Strategies to mobilise natural killer (NK) cells against cancer include tumour-targeting antibodies, NK cell engagers (NKCEs) and the adoptive transfer of ex vivo expanded healthy donor-derived NK cells. Genetic and functional studies have revealed that expression of the activating killer immunoglobulin-like receptor KIR2DS2 is associated with enhanced function in NK cells from healthy donors and improved outcome in several different malignancies. The optimal strategy to leverage KIR2DS2+ NK cells therapeutically is however currently unclear. In this study, we therefore evaluated the response of KIR2DS2-expressing NK cells to activation against cancer with clinically relevant tumour-targeting antibodies and following ex vivo expansion. We identified that KIR2DS2high NK cells from patients with chronic lymphocytic leukaemia and hepatocellular carcinoma had enhanced activation in response to tumour-targeting antibodies compared to KIR2DS2- NK cells. However, the superior function of healthy donor derived KIR2DS2high NK cells was lost following ex vivo expansion which is required for adoptive transfer-based therapeutic strategies. These data provide evidence that targeting KIR2DS2 directly in cancer patients may allow for the utilisation of their enhanced effector function, however such activity may be lost following their ex vivo expansion.
... 56 The upregulation of HLA-E in cancer cells suppresses NK cell activation through NKG2A. 57 The interaction of NK cells with HLA class I molecules regulates their overall function by modulating the amount of releasable granzyme B, which is essential for cytotoxic activity. 58 Conversely, upon activation of their receptors, NK cells can induce target cell lysis through antibody-dependent cellular cytotoxicity (ADCC) by releasing granzymes, granulysins and perforins. ...
Background and main body
The anti‐tumour and tumour‐promoting roles of B cells in the tumour microenvironment (TME) have gained considerable attention in recent years. As essential orchestrators of humoral immunity, B cells potentially play a crucial role in anti‐tumour therapies. Chemotherapy, a mainstay in cancer treatment, influences the proliferation and function of diverse B‐cell subsets and their crosstalk with the TME. Modulating B‐cell function by targeting B cells or their associated cells may enhance chemotherapy efficacy, presenting a promising avenue for future targeted therapy investigations.
Conclusion
This review explores the intricate interplay between chemotherapy and B cells, underscoring the pivotal role of B cells in chemotherapy treatment. We summarise promising B‐cell‐related therapeutic targets, illustrating the immense potential of B cells in anti‐tumour therapy. Our work lays a theoretical foundation for harnessing B cells in chemotherapy and combination strategies for cancer treatment.
Key points
Chemotherapy can inhibit B‐cell proliferation and alter subset distributions and functions, including factor secretion, receptor signalling, and costimulation.
Chemotherapy can modulate complex B‐cell–T‐cell interactions with variable effects on anti‐tumour immunity.
Targeting B‐cell surface markers or signalling improves chemotherapy responses, blocks immune evasion and inhibits tumour growth.
Critical knowledge gaps remain regarding B‐cell interactions in TME, B‐cell chemoresistance mechanisms, TLS biology, heterogeneity, spatial distributions, chemotherapy drug selection and B‐cell targets that future studies should address.
... The changes in the peptide-binding cleft of the HLA molecule caused by these polymorphisms control how the antigens are presented to and bound to T-cell receptors [42,44,45]. Through mediating antigen presentation and cell-mediated immune responses, the HLA genes have an impact on both the innate and adaptive immune systems by making it easier to identify and eliminate virusinfected cells [46,47]. ...
Most cervical cancer cases are a result of persistent high-risk Human papillomavirus (HPV) infections. Cervical cancer is prevalent in LMICs especially in SSA where it is second most prevalent cancer. In South Africa, it is the second most common cancer affecting women aged between 15 and 44 years. The host immune response has been shown as an important factor in controlling the progression or regression of high-risk HPV infection of the cervix to cervical cancer. The risk of cervical cancer is known to be influenced by the host's genetic diversity, particularly by immune response-regulating genes such as the Human Leukocyte Antigen (HLA) class I and II genes. HLA class I genes present viral peptides to CD8+ T cells which are restricted and pre-programmed for cytotoxic functions. There is very little known about the HLA genes of South African women and how these influence outcome of HPV infections. This review aims to understand the role and influence of HLA class I genes in HPV clearance, persistence, and CD8+ T cell mediated immunity in African women by examining existing literature in this area. Understanding this role can influence and inform therapeutic vaccine design that is population specific.
... Studies have demonstrated that lung NK cells are more mature (CD11 brigh CD27 low ), express lower levels of activating co-stimulatory molecules (such as CD1d, CD86, and B220), and express higher levels of inhibitory receptors (CD94-NKG2A) than activating receptors (NKG2D) compared with those in the bone marrow or spleen, which helps maintain pulmonary homeostasis (Wang et al., 2012). The final activity of NK cells is determined by the signals from various inhibitory or activating receptors (Djaoud and Parham, 2020). Killer-cell immunoglobulin-like receptors (KIR) are germ-line encoded receptors that bind to classical human leukocyte antigen (HLA) class I molecules as their ligands, a process named "licensing," which plays an important role in NK cell development. ...
Lung cancer is one of the most prevalent malignancies worldwide, contributing to over a million cancer-related deaths annually. Despite extensive research investigating the genetic factors associated with lung cancer susceptibility and prognosis, few studies have explored genetic predispositions regarding the immune system. This review discusses the most recent genomic findings related to the susceptibility to or protection against lung cancer, patient survival, and therapeutic responses. The results demonstrated the effect of immunogenetic variations in immune system-related genes associated with innate and adaptive immune responses, cytokine, and chemokine secretions, and signaling pathways. These genetic diversities may affect the crosstalk between tumor and immune cells within the tumor microenvironment, influencing cancer progression, invasion, and prognosis. Given the considerable variability in the individual immunegenomics profiles, future studies should prioritize large-scale analyses to identify potential genetic variations associated with lung cancer using highthroughput technologies across different populations. This approach will provide further information for predicting response to targeted therapy and promotes the development of new measures for individualized cancer treatment.
... Peptide binding specificity is determined by anchoring interactions with HLA side chains forming the A-F-pockets, which impose chemical and steric restrictions on peptide docking 6 . Thus, the conformational landscape of pMHC-I structures is defined by both the peptide sequence and HLA groove polymorphisms, which has important ramifications for recognition by antibodies, T-cell receptors, and NK cell receptors [7][8][9][10][11] . Since the surface chemistry of pMHC-I proteins dictates recognition by T-cell receptors (TCRs) and therapeutic monoclonal antibodies [12][13][14] , structure determination of disease-associated complexes is essential for understanding immune responses, and the development of targeted therapies. ...
The conformational landscapes of peptide/human leucocyte antigen (pHLA) protein complexes encompassing tumor neoantigens provide a rationale for target selection towards autologous T cell, vaccine, and antibody-based therapeutic modalities. Here, using complementary biophysical and computational methods, we characterize recurrent RAS 55-64 Q61 neoepitopes presented by the common HLA-A*01:01 allotype. We integrate sparse NMR restraints with Rosetta docking to determine the solution structure of NRAS Q61K /HLA-A*01:01, which enables modeling of other common RAS 55-64 neoepitopes. Hydrogen/deuterium exchange mass spectrometry experiments alongside molecular dynamics simulations reveal differences in solvent accessibility and conformational plasticity across a panel of common Q61 neoepitopes that are relevant for recognition by immunoreceptors. Finally, we predict binding and provide structural models of NRAS Q61K antigens spanning the entire HLA allelic landscape, together with in vitro validation for HLA-A*01:191, HLA-B*15:01, and HLA-C*08:02. Our work provides a basis to deline-ate the solution surface features and immunogenicity of clinically relevant neoepitope/HLA targets for cancer therapy.
... In addition, CAR-NK cells have promise as an allogeneic "off-the-shelf" product that can be given to multiple patients, allowing for reduced production costs and faster delivery of therapy compared to CAR-T cells. There are currently over 30 active clinical trials evaluating CAR-NK cells in cancer, with donor NK cells sourced from cord blood, iPSC, and peripheral blood [4] . ...
Chimeric antigen receptor (CAR) NK cells are demonstrating promising activity in clinical trials and possess a favorable safety profile compared to CAR-T cells. The Killer cell Immunoglobulin-like Receptors (KIR) have a critical role in the control of NK cell function, and recently, this family of activating and inhibitory receptors have been targeted to improve CAR-NK function. These strategies include the utilisation of inhibitory KIR to reduce trogocytosis-associated NK cell fratricide, the downregulation of inhibitory KIR on CAR-NK cells to alleviate HLA mediated suppression, the selection of CAR-NK cell donors enriched for activating KIR, and the use of activating KIR intracellular domains within novel CAR constructs. These pre-clinical studies demonstrate the potential utility of targeting the KIR to improve CAR-NK cell efficacy and patient outcomes.
... Notably, g-NK cells from both PB and UCB exhibited lower expression levels of CD16 and SYK compared to their c-NK counterparts (Figure 2a Killer cell immunoglobulin-like receptors (KIRs) are essential in dictating the education, maturation, and function of NK cells. 18 Further examination revealed that UCB-NK cells exhibited decreased expression levels of KIR2DL2/L3 but elevated expression of KIR3DL1 when compared to PB-NK cells (Figure 2b and Supplementary Figure S2b). Additionally, UCB-g-NK cells demonstrated higher expression levels of KIR2DL1 than PB-g-NK cells, while the reverse pattern was observed in the c-NK subsets ( Figure 2c). ...
FcRγ-deficient natural killer (NK) cells, designated as g-NK cells, exhibit enhanced antibody-dependent cellular cytotoxicity (ADCC) capacity and increased IFN-γ and TNF-α production, rendering them promising for antiviral and antitumor responses. g-NK cells from peripheral blood (PB) are often associated with prior human cytomegalovirus (HCMV) infection. However, the prevalence, phenotype, and function of g-NK cells in umbilical cord blood (UCB-g-NK) remain unclear. Here, we demonstrate significant phenotypical differences between UCB-g-NK and PB-g-NK cells. Unlike PB-g-NK cells, UCB-g-NK cells did not show heightened cytokine production upon CD16 engagement, in contrast to the conventional NK (c-NK) cell counterparts. Interestingly, following in vitro activation, UCB-g-NK cells also exhibited elevated levels of IFN-γ production, particularly when co-cultured with HCMV and plasma from g-NK⁺ adults. Furthermore, g-NK⁺ plasma from PB even facilitated the in vitro expansion of UCB-g-NK cells. These findings underscore the phenotypic and functional heterogeneity of g-NK cells based on their origin and demonstrate that components within g-NK⁺ plasma may directly contribute to the acquisition of an adult phenotype by the “immature” UCB-g-NK cells.
... Therefore, the effect of MHC Ia on Ib-CD8 + T cells is distinct from allo-reaction or superantigen activation. On the contrary, ligation of killer-cell immunoglobulin-like receptors by MHC Ia prevents the cytotoxic effects of NK cells and CD8 + T cells (36,37). Further investiga tions to parse out the specific receptor(s) through which MHC Ia molecules affect Ib-CD8 + T cells will yield important insights into the immunoregulatory function of MHC Ia and the anti-tumor activity of Ib-CD8 + T cells. ...
The importance of classical CD8 ⁺ T cells in tumor eradication is well acknowledged. However, the anti-tumor activity of MHC (major histocompatibility complex) Ib-restricted CD8 ⁺ T (Ib-CD8 ⁺ T) cells remains obscure. Here, we show that CX3CR1-expressing Ib-CD8 ⁺ T cells (Ib-restricted CD8 ⁺ T cells) highly express cytotoxic factors, austerely resist exhaustion, and effectively eliminate various tumors. These Ib-CD8 ⁺ T cells can be primed by MHC Ia (MHC class Ia molecules) expressed on various cell types for optimal activation in a Tbet-dependent manner. Importantly, MHC Ia does not allogeneically activate Ib-CD8 ⁺ T cells, rather, sensitizes these cells for T cell receptor activation. Such effects were observed when MHC Ia ⁺ cells were administered to tumor-bearing K b−/− D b−/− mice. A similar population of tumoricidal CX3CR1 ⁺ CD8 ⁺ T cells was identified in wild-type mice and melanoma patients. Adoptive transfer of Ib-CD8 ⁺ T cells to wild-type mice inhibited tumor progression without damaging normal tissues. Taken together, we demonstrate that MHC class Ia can prime Ib-CD8 ⁺ T cells for robust tumoricidal activities.
... These inhibitory receptors bind to inhibitory KIRs, restrict NK cell activity, and prevent healthy self-cell death. Acting as antigen-presenting molecules for T cell or NK cell target receptors, they play a vital role in the adaptive immune response [14][15][16]. The human KIR gene family has been subdivided into four groups on chromosome 19q13.4 ...
Background
KIR/HLA mismatch in hematopoietic stem cell transplantation (HSCT), particularly in patients with acute myeloid leukemia (AML), was related to decreased recurrence rates, improved engraftment, and a reduction in graft-versus-host disease, according to recent research (GVHD). Uncertainty exists about the impact of KIR/HLA mismatch on haploidentical-HSCTs treated with post-transplant cyclophosphamide (PTCy). We attempted to analyze the effects of KIR/HLA mismatch on clinical outcomes on transplant outcomes using the cohort of 54 AML patients who received a haplo-HSCT with PTCy.
Results
In contrast to KIR/HLA match, our findings showed that donor KIR/HLA mismatch was substantially associated with superior OS (HR, 2.92; (P = 0.04)). Moreover, donor KIR/HLA mismatch (KIR2DS1D/C2⁺R and KIR2DS2D/C1⁺R mismatch versus KIR2DL1D/C2⁻R mm, KIR2DL2/3D/C1⁻R mm and KIR3DL1D/Bw4⁻ mm) was correlated with the improvements in OS (HR, 0.74; P = 0.085) and activating. KIR/HLA mismatch versus KIR/HLA match was significantly correlated with improvements in OS (HR, .46; P = 0.03) and inhibitory. KIR/HLA mismatch versus KIR/HLA match was enhancement in the OS (HR, .93; P = 0.06). Despite a higher rate of aGvHD (grade I-IV) in the patients with KIR/HLA mismatch compared to KIR/HLA matched (57% vs. 33% (p = 0.04). However, the KIR/HLA mismatch group saw a decreased relapse rate (3.2% vs. 23%, p = 0.04).
Conclusion
This analysis shows the significance of KIR/HLA Incompatibility, other clinical variables like CMV, the relationship between donor/recipient and donor age, and the relationship between donor/recipient and donor age in the haplo-donor selection process. It also suggests that KIR and HLA mismatching between donor and recipient could be routinely performed for haplo-donor selection and may improve clinical outcomes after haplo-HSCTs with PTCy.
... Multigene families involved in innate immunity include natural killer cell receptors (NKRs) and their homologues that primarily recognize MHC molecules but also pathogen-encoded peptides and decoys (Djaoud & Parham, 2020), as well as PRRs that recognize pathogen molecules (pathogen-associated molecular patterns, PAMPs) but also self-molecules located in the wrong context indicating stress (danger-or damage-associated molecular patterns, DAMPs; Gong et al., 2020). The NKRs and their homologues include the multigene families of KIRs, LILRs, FcRs, NKG2A/CD94, NKG2D, NKR-P1 and CD300 among others, while the PRRs include TLRs, NLRs, CLRs, scavenger receptors, AIMs, Siglecs and butyrophilins among others. ...
Driven by co-evolution with pathogens, host immunity continuously adapts to optimize defence against pathogens within a given environment. Recent advances in genetics, genomics and transcriptomics have enabled a more detailed investigation into how immunogenetic variation shapes the diversity of immune responses seen across domestic and wild animal species. However, a deeper understanding of the diverse molecular mechanisms that shape immunity within and among species is still needed to gain insight into—and generate evolutionary hypotheses on—the ultimate drivers of immunological differences. Here, we discuss current advances in our understanding of molecular evolution underpinning jawed vertebrate immunity. First, we introduce the immunome concept, a framework for characterizing genes involved in immune defence from a comparative perspective, then we outline how immune genes of interest can be identified. Second, we focus on how different selection modes are observed acting across groups of immune genes and propose hypotheses to explain these differences. We then provide an overview of the approaches used so far to study the evolutionary heterogeneity of immune genes on macro and microevolutionary scales. Finally, we discuss some of the current evidence as to how specific pathogens affect the evolution of different groups of immune genes. This review results from the collective discussion on the current key challenges in evolutionary immunology conducted at the ESEB 2021 Online Satellite Symposium: Molecular evolution of the vertebrate immune system, from the lab to natural populations.
Abstract
Reviewing current advances in our understanding of molecular evolution underpinning vertebrate immunity, we propose hypotheses to explain differences in selection modes across immune genes and discuss supporting evidence.
... This finding is consistent with previous tissue based reports 11 . Our study adds to the growing literature on the genetic basis of the immune dysregulation of AD and, specifically, that KIR allelic variation is associated with an increased risk of AD 20,27,29,40,41 . Further a paradigm shift has occurred in understanding of the immunology of AD suggesting that NK deficiency may be real, may interplay with barrier dysfunction and that treatment studies specifically examining NK induction and modulation are now warranted 11 . ...
Natural killer cells (NK) have been associated with the pathophysiology of atopic dermatitis (AD). NK function is regulated by killer cell Ig-like receptor family (KIR) receptors that interact with HLA ligands. The study goal was to focus on allelic variation in genes KIR2DL5, KIR2DS5, and KIR2DS1 with respect to AD. This was a case–control study of individuals with (n = 313) and without (n = 176) AD. Associations were estimated using logistic regression. The prevalence of KIR2DL5 was 52.5% (95% CI 48.0,57.0), KIR2DS5 was 33.0% (28.8,37.3), and KIR2DS1 was 33.6% (29.4,38.0). The presence of the KIR2DL5*001:01 increased the odds of having AD by about 86% (odds ratio (OR): 1.86(1.23,2.82) p = 0.003). The risk for individuals homozygous for KIR2DL5*001:01 was even greater (OR: 2.16 (95% CI 1.31,3.53) p = 0.0023). The odds of having AD with KIR2DL5*001:01 was similar in Whites and Blacks. Allelic variation in KIR2DS5 and KIR2DS1 was not associated with AD. There is no known HLA binding ligand for KIR2DL5. The effect of KIR2DL5*001:01 increased in the presence of HLA-B*-21TT leader sequence (2.46(1.37,4.41) p = 0.0025) and the HLA-C2 ligand (2.07 (1.37,4.41, p = 0.000002). Our study shows an independent association of the KIR2DL5*001:01 with AD and is the first study to associate AD with KIR allelic variation.
... Through this negative and positive selection, T cells provide specificity for pathogen-derived antigens while preventing autoimmune responses (5,6). Human leukocyte antigen (HLA), also known as human MHC, is highly polymorphic, with more than 18,000 class I and 7,000 class II allotypes, which leads to many allelespecific peptide binding repertoires that can generally be characterized by sequence motifs (4,7,8). ...
Advances in next-generation sequencing (NGS) have improved the resolution of T-cell receptor (TCR) repertoire analysis, and recent single-cell sequencing has made it possible to obtain information about TCR pairs. In our previous study, cytomegalovirus (CMV) pp65-specific T-cell response restricted by a single human leukocyte antigen (HLA) class I allotype was observed in an individual. Therefore, to effectively clone an antigen-specific TCR from these T cells, we developed a TCR cloning system that does not require a single cell level. First, we established the improved Jurkat reporter cell line, which was TCRαβ double knock-out and expressed CD8αβ molecules. Furthermore, functional TCRs were directly obtained by reverse TCR cloning using unique CDR3-specific PCR primers after bulk TCR sequencing of activation marker-positive CD8 T cells by NGS. A total of 15 TCRα and 14 TCRβ strands were successfully amplified by PCR from cDNA of 4-1BB-positive CD8 T cells restricted by HLA-A*02:01, HLA-A*02:06, HLA-B*07:02, and HLA-B*40:06. The panels with combinations of TCRα and TCRβ genes were investigated using Jurkat reporter cell line and artificial antigen-presenting cells (APCs). In two TCR pairs restricted by HLA-A*02:01, one TCR pair by HLA-A*02:06, four TCR pairs by HLA-B*07:02, and one TCR pair by HLA-B*40:06, their specificity and affinity were confirmed. The TCR pair of A*02:01/1-1 showed alloreactivity to HLA-A*02:06. The one TCR pair showed a higher response to the naturally processed antigen than that of the peptide pool. This reverse TCR cloning system will not only provide functional information to TCR repertoire analysis by NGS but also help in the development of TCR-T therapy.
... The activity of NK cells is modulated by a balance between the net input of signals derived from an array of inhibitory and activating receptors 14 . Among these germ-line encoded receptors, killer-cell immunoglobulin-like (Table 3). ...
Killer-cell immunoglobulin-like receptors (KIR) are essential for acquiring natural killer (NK) cell effector function, which is modulated by a balance between the net input of signals derived from inhibitory and activating receptors through engagement by human leukocyte antigen (HLA) class I ligands. KIR and HLA loci are polygenic and polymorphic and exhibit substantial variation between individuals and populations. We attempted to investigate the contribution of KIR complex and HLA class I ligands to the genetic predisposition to lung cancer in the native population of southern Iran. We genotyped 16 KIR genes for a total of 232 patients with lung cancer and 448 healthy controls (HC), among which 85 patients and 178 HCs were taken into account for evaluating combined KIR-HLA associations. KIR2DL2 and 2DS2 were increased significantly in patients than in controls, individually (OR 1.63, and OR 1.42, respectively) and in combination with HLA-C1 ligands (OR 1.99, and OR 1.93, respectively). KIR3DS1 (OR 0.67) and 2DS1 (OR 0.69) were more likely presented in controls in the absence of their relative ligands. The incidence of CxTx subset was increased in lung cancer patients (OR 1.83), and disease risk strikingly increased by more than fivefold among genotype ID19 carriers (a CxTx genotype that carries 2DL2 in the absence of 2DS2, OR 5.92). We found that genotypes with iKIRs > aKIRs (OR 1.67) were more frequently presented in lung cancer patients. Additionally, patients with lung cancer were more likely to carry the combination of CxTx/2DS2 compared to controls (OR 2.04), and iKIRs > aKIRs genotypes in the presence of 2DL2 (OR 2.05) increased the likelihood of lung cancer development. Here we report new susceptibility factors and the contribution of KIR and HLA-I encoding genes to lung cancer risk, highlighting an array of genetic effects and disease setting which regulates NK cell responsiveness. Our results suggest that inherited KIR genes and HLA-I ligands specifying the educational state of NK cells can modify lung cancer risk.
... The major histocompatibility complex (MHC) is a genomic region originally discovered as the primary genetic locus responsible for graft rejection, but now is known to encode the highly polymorphic classical class I and class II molecules that present antigenic peptides to play crucial roles in innate and adaptive immune responses (1). Over three decades of research into human and mouse MHC molecules have provided very clear models of how MHC molecules acquire, bind and present peptides to thymus-derived (T) lymphocytes and natural killer (NK) cells (2,3). Much less is known about the MHC systems in other species, ranging from mammals outside of primates and rodents to cartilaginous fish (4). ...
The functions of a wide variety of molecules with structures similar to the classical class I and class II molecules encoded by the major histocompatibility complex (MHC) have been studied by biochemical and structural studies over decades, with many aspects for humans and mice now enshrined in textbooks as dogma. However, there is much variation of the MHC and MHC molecules among the other jawed vertebrates, understood in the most detail for the domestic chicken. Among the many unexpected features in chickens is the co-evolution between polymorphic TAP and tapasin genes with a dominantly-expressed class I gene based on a different genomic arrangement compared to typical mammals. Another important discovery was the hierarchy of class I alleles for a suite of properties including size of peptide repertoire, stability and cell surface expression level, which is also found in humans although not as extreme, and which led to the concept of generalists and specialists in response to infectious pathogens. Structural studies of chicken class I molecules have provided molecular explanations for the differences in peptide binding compared to typical mammals. These unexpected phenomena include the stringent binding with three anchor residues and acidic residues at the peptide C-terminus for fastidious alleles, and the remodelling binding sites, relaxed binding of anchor residues in broad hydrophobic pockets and extension at the peptide C-terminus for promiscuous alleles. The first few studies for chicken class II molecules have already uncovered unanticipated structural features, including an allele that binds peptides by a decamer core. It seems likely that the understanding of how MHC molecules bind and present peptides to lymphocytes will broaden considerably with further unexpected discoveries through biochemical and structural studies for chickens and other non-mammalian vertebrates.
... There is substantial diversity in the number of KIR genes that individuals have and these are encoded as two haplotypes (13). The KIR A haplotype consists of predominantly inhibitory KIR and one activating KIR (KIR2DS4), whilst the KIR B haplotype encodes a more variable gene content which includes more activating KIR (KIR2DS1, KIR2DS2, KIR2DS3, KIR2DS5 and KIR3DS1). ...
NK cells are promising cellular therapeutics against hematological and solid malignancies. Immunogenetic studies have identified that various activating killer cell Ig-like receptors (KIRs) are associated with cancer outcomes. Specifically, KIR2DS2 has been associated with reduced incidence of relapse following transplant in hematological malignancies and improved outcomes in solid tumors, but the mechanism remains obscure. Therefore, we investigated how KIR2DS2 expression impacts NK cell function. Using a novel flow cytometry panel, we show that human NK cells with high KIR2DS2 expression have enhanced spontaneous activation against malignant B cell lines, liver cancer cell lines, and primary chronic lymphocytic leukemia cells. Surface expression of CD16 was increased on KIR2DS2high NK cells, and, accordingly, KIR2DS2high NK cells had increased activation against lymphoma cells coated with the clinically relevant anti-CD20 Abs rituximab and obinutuzumab. Bulk RNA sequencing revealed that KIR2DS2high NK cells have upregulation of NK-mediated cytotoxicity, translation, and FCGR gene pathways. We developed a novel single-cell RNA-sequencing technique to identify KIR2DS2+ NK cells, and this confirmed that KIR2DS2 is associated with enhanced NK cell-mediated cytotoxicity. This study provides evidence that KIR2DS2 marks a population of NK cells primed for anticancer activity and indicates that KIR2DS2 is an attractive target for NK-based therapeutic strategies.
... Propargyl-peg8-CD22L (2) was synthesized using 2 equiv of NHS-peg8-propargyl with 1 equiv of CD22 ligand (CD22L) in dry DMSO at room temperature for 5 h. Next, propargyl-peg8-CD22L were conjugated to anti-IgD-N 3 (4) under Cu(I)-catalyzed click chemistry to afford anti-IgD with 5−6 CD22L per antibody (anti-IgD-CD22L, 5a), as assessed by MALDI-TOF mass spectroscopy. Anti-IgD-N 3 was similarly treated without propargyl-peg8-CD22L to afford 5b to serve as a control that was subjected to the same chemical treatments but without CD22L. ...
The sialic acid-binding immunoglobulin-type lectins (Siglecs) are expressed predominantly on white blood cells and participate in immune cell recognition of self. Most Siglecs contain cytoplasmic inhibitory immunoreceptor tyrosine-based inhibitory motifs characteristic of inhibitory checkpoint co-receptors that suppress cell signaling when they are recruited to the immunological synapse of an activating receptor. Antibodies to activatory receptors typically activate immune cells by ligating the receptors on the cell surface. Here, we report that the conjugation of high affinity ligands of Siglecs to antibodies targeting activatory immune receptors can suppress receptor-mediated activation of immune cells. Indeed, B-cell activation by antibodies to the B-cell receptor IgD is dramatically suppressed by conjugation of anti-IgD with high affinity ligands of a B-cell Siglec CD22/Siglec-2. Similarly, degranulation of mast cells induced by antibodies to IgE, which ligate the IgE/FcεR1 receptor complex, is suppressed by conjugation of anti-IgE to high affinity ligands of a mast cell Siglec, CD33/Siglec-3 (CD33L). Moreover, the anti-IgE-CD33L suppresses anti-IgE-mediated systemic anaphylaxis of sensitized humanized mice and prevents anaphylaxis upon subsequent challenge with anti-IgE. The results demonstrate that attachment of ligands of inhibitory Siglecs to anti-receptor antibodies can suppress the activation of immune cells and modulate unwanted immune responses.
... Since the α3 region of HLA class I antigens and β2m are relatively conserved, LILRB1 binds to a broad range of HLA class I. The LILRB1-HLA class I binding pattern is different from that of KIRs and T cell receptors (TCR), which recognize the polymorphic α1 and α2 domains of subsets of HLA class I (Djaoud and Parham 2020). Jones et al. found that LILRB1 bound to a broad range of HLA class I by testing the binding of one LILRB1 variant (unspecified, but most likely variant 1) to a panel of 97 HLA class I antigens in a single antigen bead assay (Jones et al. 2011). ...
Leukocyte immunoglobulin-like receptor B1 (LILRB1) is widely expressed on various immune cells and the engagement of LILRB1 to HLA class I and pathogen-derived proteins can modulate the immune response. In the current study, 108 LILRB1 alleles were identified by screening the LILRB1 locus from the 1000 Genomes Phase 3 database. Forty-six alleles that occurred in three or more individuals encode 28 LILRB1 allotypes, and the inferred LILRB1 allotypes were then grouped into 9 LILRB1 D1-D2 variants for further analysis. We found that variants 1, 2, and 3 represent the three most frequent LILRB1 D1-D2 variants and the nine variants show frequency differences in populations. The binding assay demonstrated that variant 1 bound to HLA class I with the highest avidity, and all tested LILRB1 D1-D2 variants bound to HLA-C with lower avidity than to HLA-A and -B. Locus-specific polymorphisms at positions 183, 189, and 268 in HLA class I and dimorphisms in HLA-A (positions 207 and 253) and in HLA-B (position 194) affect their binding to LILRB1. Notably, the electrostatic interaction plays a critical role in the binding of LILRB1 to HLA class I as revealed by electrostatic analysis and by comparison of different binding avidities caused by polymorphisms at positions 72 and 103 of LILRB1. In this paper, we present a comprehensive study of the population genetics and binding abilities of LILRB1. The data will help us better understand the LILRB1-related diversity of the immune system and lay a foundation for functional studies.
... The up-regulation of ARID5B and its inducing UQCRB with low H3K9Me2 in the promoter region alter metabolic reprogramming, prolong survival, and motivate abundant IFN-g secretion of adaptive NK cells. Abs, antibodies; CD, cluster of differentiation; TIGIT, T-cell immunoglobulin and ITIM domain; interleukin, IL; IFN-g, Interferon-gamma; TNF-a, Tumor necrosis factor alpha; and ARID5B, AT-rich interaction domain 5. 2DS4), which are correlated with resistance to viral infection (28,41). Whether KIRs play a role in adaptive NK cell expansion requires further study. ...
Natural killer (NK) cells are an important component of the innate immune system due to their strong ability to kill virally infected or transformed cells without prior exposure to the antigen (Ag). However, the biology of human NK (hNK) cells has largely remained elusive. Recent advances have characterized several novel hNK subsets. Among them, adaptive NK cells demonstrate an intriguing specialized antibody (Ab)-dependent response and several adaptive immune features. Most adaptive NK cells express a higher level of NKG2C but lack an intracellular signaling adaptor, FcϵRIγ (hereafter abbreviated as FcRγ). The specific expression pattern of these genes, with other signature genes, is the result of a specific epigenetic modification. The expansion of adaptive NK cells in vivo has been documented in various viral infections, while the frequency of adaptive NK cells among peripheral blood mononuclear cells correlates with improved prognosis of monoclonal Ab treatment against leukemia. This review summarizes the discovery and signature phenotype of adaptive NK cells. We also discuss the reported association between adaptive NK cells and pathological conditions. Finally, we briefly highlight the application of adaptive NK cells in adoptive cell therapy against cancer.
... In humans, killer immunoglobulin-like receptors (KIRs) recognize classical class I molecules, including some HLA-A alleles (A3 and A11), some HLA-B alleles (those with Bw4 epitopes) and all HLA-C alleles (divided into C1 and C2 allelic lineages). On most human cell types, HLA-A and HLA-B molecules are much better expressed than HLA-C molecules (Djaoud and Parham 2020). In chickens, the classical class I molecules encoded by the BF2 gene are far better expressed than those expressed by the BF1 gene (Shaw et al. 2007;Wallny et al. 2006). ...
Compared to the major histocompatibility complex (MHC) of typical mammals, the chicken BF/BL region is small and simple, with most of the genes playing central roles in the adaptive immune response. However, some genes of the chicken MHC are almost certainly involved in innate immunity, such as the complement component C4 and the lectin-like receptor/ligand gene pair BNK and Blec. The poorly expressed classical class I molecule BF1 is known to be recognised by natural killer (NK) cells and, analogous to mammalian immune responses, the classical class I molecules BF1 and BF2, the CD1 homologs and the butyrophilin homologs called BG may be recognised by adaptive immune lymphocytes with semi-invariant receptors in a so-called adaptate manner. Moreover, the TRIM and BG regions next to the chicken MHC, along with the genetically unlinked Y and olfactory/scavenger receptor regions on the same chromosome, have multigene families almost certainly involved in innate and adaptate responses. On this chicken microchromosome, the simplicity of the adaptive immune gene systems contrasts with the complexity of the gene systems potentially involved in innate immunity.
... Our understanding of the host-pathogen interplay has improved with the discovery of newer technologies, opening unprecedented avenues into the field of rational vaccine design (Khatoon et al. 2017). Such outstanding advances include major histocompatibility complex (MHC) molecule structure and restriction, the nature of antigen presentation, T cell receptor (TCR) elucidation as well as identification of cytokines (Djaoud and Parham 2020). An efficacious vaccine candidate would be able to strongly stimulate IFN-γ-releasing Th-1 cells, through antigen presenting cells (APCs), with subsequent activation of macrophages leading to upsurge in reactive oxygen species (ROS) and nitric oxide to combat intracellular amastigotes (Rodrigues et al. 2016). ...
Visceral leishmaniasis (VL) is a severe disease with particular endemicity in over 80 countries worldwide. There is no approved human vaccine against VL in the market. This study was aimed at designing and evaluation of a multimeric vaccine candidate against Leishmania infantum through utilization of helper T lymphocyte (HTL) and cytotoxic T lymphocyte (CTL) immunodominant proteins from histone H1, KMP11, LACK and LeIF antigens. Top-ranked mouse MHC-I, MHC-II binders and CTL epitopes were predicted and joined together via spacers. Also, a TLR-4 agonist (RS-09 synthetic protein) and His-tag were added to the N- and C-terminal of the vaccine sequence, respectively. The final chimeric vaccine had a length of 184 amino acids with a molecular weight of 18.99 kDa. Physico-chemical features showed a soluble, highly-antigenic and non-allergenic candidate. Secondary and tertiary structures were predicted, and subsequent analyses confirmed the construct stability that was capable to properly interact with TLR-4/MD2 receptor. Immunoinformatics simulation displayed potent stimulation of T cell immune responses, with particular rise in IFN-γ, upon vaccination with the proposed multi-epitope candidate. In conclusion, immunoinformatics data demonstrated a highly antigenic vaccine candidate in mouse, which could develop considerable levels clearance mechanisms and other components of cellular immune profile, and can be directed for VL prophylactic purposes.
Supplementary information:
The online version contains supplementary material available at 10.1007/s11756-021-00934-3.
MHC I proteins present epitopic peptides to CD8+ T cells to elicit multifaceted adaptive immune responses. The affinity and avidity of interactions between peptide/MHC molecules and T cell receptors (TCR) are fundamental parameters that contribute to the induction of activated or anergic T cell states. Here, we present a loadable system, VLP/Open HLA, featuring a virus-like particle (VLP) that can accommodate up to 60 loadable HLA (HLA, human leukocyte antigen) molecules. HLA nanoparticles, pre-loaded with a placeholder ligand, allow efficient peptide exchange upon incubation with target peptides. We show that fluorescently tagged VLP-Open HLA particles can be used to stain CD8+ T cells, providing a screening tool for novel TCRs. Finally, we demonstrate that our system can induce activation of T cells in an antigen specific manner. Our platform can be adapted to encompass multiple HLA allotypes and costimulatory molecules as mosaic nanoparticles, to enable a range of applications in experimental immunology.
The limited infiltration of CD8+ T cells in tumors hampers the effectiveness of T cell–based immunotherapy, yet the mechanisms that limit tumor infiltration by CD8+ T cells remain unclear. Through bulk RNA sequencing of human tumors, we identified a strong correlation between WNT7A expression and reduced CD8+ T-cell infiltration. Further investigation demonstrated that inhibiting WNT7A substantially enhanced MHC-I expression on tumor cells. Mechanistically, WNT7A inhibition inactivated the Wnt/β-catenin signaling pathway and thus resulted in reduced physical interaction between β-catenin and p65 in the cytoplasm, which increased the nuclear translocation of p65 and activated the NF-κB pathway, ultimately promoting the transcription of genes encoding MHC-I molecules. We found that our lead compound, 1365-0109, disrupted the protein–protein interaction between WNT7A and its receptor FZD5, resulting in the upregulation of MHC-I expression. In murine tumor models, both genetic and pharmaceutical suppression of WNT7A led to increased MHC-I levels on tumor cells, and consequently enhanced the infiltration and functionality of CD8+ T cells, which bolstered antitumor immunity and improved the effectiveness of immune checkpoint blockade therapy. These findings have elucidated the intrinsic mechanisms of WNT7A-induced immune suppression, suggesting that therapeutic interventions targeting WNT7A hold promise for enhancing the efficacy of immunotherapy.
Natural killer (NK) cells, initially identified for their rapid virus-infected and leukemia cell killing and tumor destruction, are pivotal in immunity. They exhibit multifaceted roles in cancer, viral infections, autoimmunity, pregnancy, wound healing, and more. Derived from a common lymphoid progenitor, they lack CD3, B-cell, or T-cell receptors but wield high cytotoxicity via perforin and granzymes. NK cells orchestrate immune responses, secreting inflammatory IFNγ or immunosuppressive TGFβ and IL-10. CD56 dim and CD56 bright NK cells execute cytotoxicity, while CD56 bright cells also regulate immunity. However, beyond the CD56 dichotomy, detailed phenotypic diversity reveals many functional subsets that may not be optimal for cancer immunotherapy. In this review, we provide comprehensive and detailed snapshots of NK cells’ functions and states of activation and inhibitions in cancer, autoimmunity, angiogenesis, wound healing, pregnancy and fertility, aging, and senescence mediated by complex signaling and ligand-receptor interactions, including the impact of the environment. As the use of engineered NK cells for cancer immunotherapy accelerates, often in the footsteps of T-cell-derived engineering, we examine the interactions of NK cells with other immune effectors and relevant signaling and the limitations in the tumor microenvironment, intending to understand how to enhance their cytolytic activities specifically for cancer immunotherapy.
The genetic diversity of killer cell immunoglobulin‐like receptors (KIRs) and human leukocyte antigen (HLA) genes influences the host's immune response to viral pathogens. This study aims to explore the impact of five single nucleotide polymorphisms (SNPs) in KIR3DL2 and HLA‐A genes on hepatitis C virus (HCV) infection. A total of 2251 individuals were included in the case‐control study. SNPs including KIR3DL2 rs11672983, rs3745902, rs1654644, and HLA‐A rs3869062, rs12202296 were genotyped. By controlling various confounding factors using a modified logistic regression model, as well as incorporating stratified analysis, joint effects analysis, and multidimensional bioinformatics analysis, we analyzed the relationship between SNPs and HCV infection. The logistic regression analysis showed a correlation between KIR3DL2 rs11672983 AA, KIR3DL2 rs3745902 TT, and increased HCV susceptibility ( p < 0.01). Stratified analysis indicated that KIR3DL2 rs1654644 and HLA‐A rs3869062 also heightened HCV susceptibility in certain subgroups. A linear trend of rising HCV infection rates was observed when combining KIR3DL2 rs11672983 AA and KIR3DL2 rs3745902 TT ( p trend = 0.007). Bioinformatics analysis suggested these SNPs' regulatory potential and their role in altering messenger RNA secondary structure, implying their functional relevance in HCV susceptibility. Our findings indicate that KIR3DL2 rs11672983 AA and KIR3DL2 rs3745902 TT are significantly associated with increased susceptibility to HCV infection.
Helicobacter pylori ( H. pylori ) seems to play causative roles in gastric cancers. H. pylori has also been detected in established gastric cancers. How the presence of H. pylori modulates immune response to the cancer is unclear. The cytotoxicity of natural killer ( NK ) cells, toward infected or malignant cells, is controlled by the repertoire of activating and inhibitory receptors expressed on their surface. Here, we studied H. pylori ‐induced changes in the expression of ligands, of activating and inhibitory receptors of NK cells, in the gastric adenocarcinoma AGS cells, and their impacts on NK cell responses. AGS cells lacked or had low surface expression of the class I major histocompatibility complex ( MHC‐I ) molecules HLA‐E and HLA‐C —ligands of the major NK cell inhibitory receptors NKG2A and killer‐cell Ig‐like receptor ( KIR ), respectively. However, AGS cells had high surface expression of ligands of activating receptors DNAM ‐1 and CD2 , and of the adhesion molecules LFA ‐1. Consistently, AGS cells were sensitive to killing by NK cells despite the expression of inhibitory KIR on NK cells. Furthermore, H. pylori enhanced HLA‐C surface expression on AGS cells. H. pylori infection enhanced HLA‐C protein synthesis, which could explain H. pylori ‐induced HLA‐C surface expression. H. pylori infection enhanced HLA‐C surface expression also in the hepatoma Huh7 and HepG2 cells. Furthermore, H. pylori ‐induced HLA‐C surface expression on AGS cells promoted inhibition of NK cells by KIR , and thereby protected AGS cells from NK cell cytotoxicity. These results suggest that H. pylori enhances HLA‐C expression in host cells and protects them from the cytotoxic attack of NK cells expressing HLA‐C‐specific inhibitory receptors.
The human leukocyte antigen (HLA) system is a major factor controlling cancer immunosurveillance and response to immunotherapy, yet its status in pediatric cancers remains fragmentary. We determined high-confidence HLA genotypes in 576 children, adolescents and young adults with recurrent/refractory solid tumors from the MOSCATO-01 and MAPPYACTS trials, using normal and tumor whole exome and RNA sequencing data and benchmarked algorithms. There was no evidence for narrowed HLA allelic diversity but discordant homozygosity and allele frequencies across tumor types and subtypes, such as in embryonal and alveolar rhabdomyosarcoma, neuroblastoma MYCN and 11q subtypes, and high-grade glioma, and several alleles may represent protective or susceptibility factors to specific pediatric solid cancers. There was a paucity of somatic mutations in HLA and antigen processing and presentation (APP) genes in most tumors, except in cases with mismatch repair deficiency or genetic instability. The prevalence of loss-of-heterozygosity (LOH) ranged from 5.9 to 7.7% in HLA class I and 8.0 to 16.7% in HLA class II genes, but was widely increased in osteosarcoma and glioblastoma (~15-25%), and for DRB1-DQA1-DQB1 in Ewing sarcoma (~23-28%) and low-grade glioma (~33-50%). HLA class I and HLA-DR antigen expression was assessed in 194 tumors and 44 patient-derived xenografts (PDXs) by immunochemistry, and class I and APP transcript levels quantified in PDXs by RT-qPCR. We confirmed that HLA class I antigen expression is heterogeneous in advanced pediatric solid tumors, with class I loss commonly associated with the transcriptional downregulation of HLA-B and transporter associated with antigen processing (TAP) genes, whereas class II antigen expression is scarce on tumor cells and occurs on immune infiltrating cells. Patients with tumors expressing sufficient HLA class I and TAP levels such as some glioma, osteosarcoma, Ewing sarcoma and non-rhabdomyosarcoma soft-tissue sarcoma cases may more likely benefit from T cell-based approaches, whereas strategies to upregulate HLA expression, to expand the immunopeptidome, and to target TAP-independent epitopes or possibly LOH might provide novel therapeutic opportunities in others. The consequences of HLA class II expression by immune cells remain to be established. Immunogenetic profiling should be implemented in routine to inform immunotherapy trials for precision medicine of pediatric cancers.
Rejection remains the main cause of premature graft loss after kidney transplantation, despite the use of potent immunosuppression. This highlights the need to better understand the composition and the cell-to-cell interactions of the alloreactive inflammatory infiltrate. Here, we performed droplet-based single-cell RNA sequencing of 35,152 transcriptomes from 16 kidney transplant biopsies with varying phenotypes and severities of rejection and without rejection, and identified cell-type specific gene expression signatures for deconvolution of bulk tissue. A specific association was identified between recipient-derived FCGR3A+ monocytes, FCGR3A+ NK cells and the severity of intragraft inflammation. Activated FCGR3A+ monocytes overexpressed CD47 and LILR genes and increased paracrine signaling pathways promoting T cell infiltration. FCGR3A+ NK cells overexpressed FCRL3, suggesting that antibody-dependent cytotoxicity is a central mechanism of NK-cell mediated graft injury. Multiplexed immunofluorescence using 38 markers on 18 independent biopsy slides confirmed this role of FcγRIII+ NK and FcγRIII+ nonclassical monocytes in antibody-mediated rejection, with specificity to the glomerular area. These results highlight the central involvement of innate immune cells in the pathogenesis of allograft rejection and identify several potential therapeutic targets that might improve allograft longevity.
The function of natural killer (NK) cells has previously been implicated in hematopoietic-related diseases. Killer immunoglobulin-like receptors (KIR) play an important role in NK cells after hematopoietic stem cell transplantation. To explore the immunogenetic predisposition of hematological-related diseases, herein, a multi-center retrospective study in China was conducted, analyzing and comparing 2519 patients with hematopathy (mainly, acute lymphoblastic leukemia, acute myeloid leukemia, aplastic anemia, and myelodysplastic syndrome) to 18,108 individuals without known pathology. Genotyping was performed by polymerase chain reaction with specific sequence primers (PCR-SSP). As a result, we discovered four genes including KIR2DL5 (OR: 0.74, 95% CI 0.59–0.93; Pc = 0.0405), 2DS1 (OR: 0.74, 95% CI 0.59–0.93; Pc = 0.0405), 2DS3 (OR: 0.58, 95% CI 0.41–0.81; Pc = 0.0180), and 3DS1 (OR: 0.74, 95% CI 0.58–0.94; Pc = 0.0405) to be protective factors that significantly reduce the risk of aplastic anemia. Our findings offer new approaches to immunotherapy for hematological-related diseases. As these therapies mature, they are promising to be used alone or in combination with current treatments to help to make blood disorders a manageable disease.
Killer immunoglobulin-like receptors (KIRs) are polymorphic receptors for human leukocyte antigens (HLAs) that provide positive or negative signals controlling lymphocyte activation. Expression of inhibitory KIRs by CD8+ T cells affects their survival and function, which is linked to improved antiviral immunity and prevention of autoimmunity. In this issue of the JCI, Zhang, Yan, and co-authors demonstrate that increased numbers of functional inhibitory KIR-HLA pairs equating to greater negative regulation promoted longer lifespans of human T cells. This effect was independent of direct signals provided to KIR-expressing T cells and was instead driven by indirect mechanisms. Since the long-term maintenance of CD8+ T cells is critical for immune readiness against cancer and infection, this discovery has implications for immunotherapy and the preservation of immune function during aging.
Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, is considered a Neglected Tropical Disease. Limited investment is assigned to its study and control, even though it is one of the most prevalent parasitic infections worldwide. An innovative vaccination strategy involving an epitope-based vaccine that displays multiple immune determinants originating from different antigens could counteract the high biological complexity of the parasite and lead to a wide and protective immune response. In this chapter, we describe a computational reverse vaccinology pipeline applied to identify the most promising peptide sequences from T. cruzi proteins, prioritizing evolutionary conserved sequences, to finally select a list of T and B cell epitope candidates to be further tested in an experimental setting.Key wordsChagas disease
Trypanosoma cruzi
EpitopesReverse vaccinology
Major histocompatibility complexes (MHC) play a key role in the immune surveillance system in all jawed vertebrates. MHC class I molecules randomly sample cytosolic peptides from inside the cell, while MHC class II sample exogenous peptides. Both types of peptide:MHC complex are then presented on the cell surface for recognition by αβ T cells (CD8+ and CD4+, respectively). The three-dimensional structure of such complexes can give crucial insights in the presentation and recognition mechanisms. For this reason, softwares like PANDORA have been developed to rapidly and accurately generate peptide:MHC (pMHC) 3D structures. In this chapter, we describe the protocol of PANDORA. PANDORA exploits the structural knowledge on anchor pockets that MHC molecules use to dock peptides. PANDORA provides anchor positions as restraints to guide the modeling process. This allows PANDORA to generate twenty 3D models in just about 5 min. PANDORA is highly customizable, easy to install, supports parallel processing, and is suitable to provide large datasets for deep learning algorithms.
As a nontraditional T-cell subgroup, γδT cells have gained popularity in the field of immunotherapy in recent years. They have extraordinary antitumor potential and prospects for clinical application. Immune checkpoint inhibitors (ICIs), which are efficacious in tumor patients, have become pioneer drugs in the field of tumor immunotherapy since they were incorporated into clinical practice. In addition, γδT cells that have infiltrated into tumor tissues are found to be in a state of exhaustion or anergy, and there is upregulation of many immune checkpoints (ICs) on their surface, suggesting that γδT cells have a similar ability to respond to ICIs as traditional effector T cells. Studies have shown that targeting ICs can reverse the dysfunctional state of γδT cells in the tumor microenvironment (TME) and exert antitumor effects by improving γδT-cell proliferation and activation and enhancing cytotoxicity. Clarification of the functional state of γδT cells in the TME and the mechanisms underlying their interaction with ICs will solidify ICIs combined with γδT cells as a good treatment option.
The extreme polymorphisms of HLA-I proteins enable the presentation of diverse peptides to cytotoxic T lymphocytes (CTL). The canonical endoplasmic reticulum (ER) HLA-I assembly pathway enables presentation of cytosolic peptides, but effective intracellular surveillance requires multi-compartmental antigen sampling. Endo-lysosomes are generally sites of HLA class II assembly, but human monocytes and monocyte-derived dendritic cells (moDCs) also contain significant reserves of endo-lysosomal HLA-I molecules. We hypothesized variable influences of HLA-I polymorphisms upon outcomes of endo-lysosomal trafficking, as the stabilities and peptide occupancies of cell surface HLA-I are variable. Consistent with this model, when the endo-lysosomal pH of moDCs is disrupted, HLA-B allotypes display varying propensities for reductions in surface expression, with HLA-B*08:01 or HLA-B*35:01 being among the most resistant or sensitive respectively, among eight tested HLA-B allotypes. Perturbations of moDC endo-lysosomal pH result in redistribution of HLA-B*35:01, but not HLA-B*08:01, to LAMP1+ compartments and increase HLA-B*35:01 peptide receptivity. These findings reveal the intersection of the vacuolar cross-presentation pathway with a constitutive assembly pathway for some HLA-B allotypes. Notably, cross-presentation of epitopes derived from two soluble antigens was also more efficient for B*35:01 compared to B*08:01, even when matched for T cell response sensitivity, and more affected by cathepsin inhibition. Thus, HLA-I polymorphisms dictate the degree of endo-lysosomal assembly, which can supplement ER assembly for constitutive HLA-I expression and increase the efficiency of cross-presentation.
Atopic dermatitis (AD) is a common chronic skin disease. Although generally thought to be a disease of T-cell dysregulation, recent studies have suggested that immune dysregulation of NK cells is also important. Killer cell Ig-like receptors (KIRs) are involved with NK cell regulation. The Pediatric Eczema Elective Registry is a U.S. nationwide longitudinal cohort with up to 10 y of follow-up in which 655 children had DNA available for full allelic KIR sequencing. Every 6 mo, AD activity was reported by Pediatric Eczema Elective Registry children. Using generalized estimating equations, we evaluated the association of KIR allelic variation in concert with known HLA binding ligands and whether the child reported AD in “remission” (no skin lesions and not using AD medication). KIR2DS4*001:01 (odds ratio 0.53, 95% CI [0.32, 0.88]) and KIR2DL4*001:02 (0.54, [0.33, 0.89]) in the presence of C*04:01 had the largest effect on decreasing the likelihood of AD remission. The haplotype KIR 2DL4*001:02 ∼ 2DS4*001:01 ∼ 3DL2*002:01 (0.77, [0.60, 0.99]) was also associated with a decreased likelihood of AD remission. Our findings add to the general body of evidence of a growing literature on the importance of NK cells with respect to the immunopathogenesis and natural history of AD.
The worldwide coronavirus disease 2019 pandemic was sparked by the severe acute respiratory syndrome caused by coronavirus 2 (SARS-CoV-2) that first surfaced in December 2019 (COVID-19). The effects of COVID-19 differ substantially not just between patients individually but also between populations with different ancestries. In humans, the human leukocyte antigen (HLA) system coordinates immune regulation. Since HLA molecules are a major component of antigen-presenting pathway, they play an important role in determining susceptibility to infectious disease. It is likely that differential susceptibility to SARS-CoV-2 infection and/or disease course in COVID-19 in different individuals could be influenced by the variations in the HLA genes which are associated with various immune responses to SARS-CoV-2. A growing number of studies have identified a connection between HLA variation and diverse COVID-19 outcomes. Here, we review research investigating the impact of HLA on individual responses to SARS-CoV-2 infection and/or progression, also discussing the significance of MHC-related immunological patterns and its use in vaccine design.
Proper embryo implantation depends on the tolerance of the maternal immune system to the fetus and its foreign paternal antigens. During implantation and early pregnancy, the dominant leukocytes in the uterus are uterine NK cells, expressing killer immunoglobulin-like receptors (KIR). KIRs recognize human leukocyte antigens (HLA-C) on the human trophoblast inherited from the father and mother. The antigenic peptides presented by the HLA are formed via their cleavage by endoplasmic reticulum aminopeptidases ERAP1 and ERAP2. The aim of this study was to assess the association of combined KIR genes and their HLA-C ligands, as well as ERAP1 and ERAP2 polymorphisms with recurrent implantation failure after in vitro fertilization (RIF). We tested 491 couples who underwent in vitro fertilization (IVF) and 322 fertile couples. Genotype CC rs27044 ERAP1 in female with a male’s HLA-C1C1 or HLA-C1C2 protected from RIF (p/pcorr. = 0.005/0.044, OR = 0.343; p/pcorr. = 0.003/0.027, OR = 0.442, respectively). Genotype TT rs30187 ERAP1 in female with a male’s HLA-C1C2 genotype increased the risk of RIF. Summarizing, in the combination of female ERAP1 and an HLA-C partner, the rs30187 C>T and rs27044 C>G polymorphisms play an important role in implantation failure.
The role of the maternal immune system in reproductive success in humans remains controversial. Here we focus on the events that occur in the maternal decidua during the first few weeks of human pregnancy, because this is the site at which maternal leukocytes initially interact with and can recognize fetal trophoblast cells, potentially involving allorecognition by both T cells and natural killer (NK) cells. NK cells are the dominant leukocyte population in first-trimester decidua, and genetic studies point to a role of allorecognition by uterine NK cells in establishing a boundary between the mother and the fetus. By contrast, definitive evidence that allorecognition by decidual T cells occurs during the first trimester is lacking. Thus, our view is that during the crucial period when the placenta is established, damaging T cell-mediated adaptive immune responses towards placental trophoblast are minimized, whereas NK cell allorecognition contributes to successful implantation and healthy pregnancy. Major human pregnancy disorders arise from the failure of placental trophoblast to access sufficient supplies of maternal oxygen and nutrients. Key to understanding this process are the interactions that occur early in pregnancy between trophoblast cells that invade the decidua and maternal uterine immune cells.
The major histocompatibility complex (MHC) is a large genetic region with hundreds of genes, among them the most polymorphic loci known in humans and other vertebrates. MHC genes are intensively studied because they influence so many important traits, including compatibility of transplants, autoimmunity, resistance to infectious diseases and reproduction.
Key Concepts
• The human MHC (HLA) is a large genetic region with hundreds of genes, including the classical class I and class II MHC molecules that are responsible for tissue graft rejection.
• Human classical class I and class II MHC molecules have hundreds of alleles, are well-expressed across many tissues and present peptides to T lymphocytes and natural killer (NK) cells, while nonclassical MHC molecules (some of which are encoded outside of the MHC) lack one or more of these properties.
• Human classical class I and class II MHC molecules are polymorphic due to a molecular arms race with pathogens, with MHC molecules selecting pathogens that no longer express the previously presented peptides, which then drives the appearance of new alleles of MHC molecules that bind the newly evolved pathogen peptides.
• Human classical class I and class II MHC molecules play key roles in resistance to infectious disease, susceptibility to autoimmune disease, allergy and asthma, and in reproduction.
• Human classical class I MHC molecules present peptides primarily derived from the cytoplasm and nucleus where viruses replicate, and are recognized by cytotoxic CD8-bearing T lymphocytes (CTLs) and natural killer (NK) cells.
• Human classical class II MHC molecules present peptides primarily derived from intracellular vesicles and the extracellular space where many different kinds of pathogens are found, and are recognized by a variety of CD4-bearing T cells with nuanced responses depending on the nature of the pathogen.
• Human nonclassical MHC molecules (particularly class I molecules encoded outside of the MHC) have a wide variety of functions, only some of which are involved with immunity.
• Not all genetic associations with the human MHC involve classical and nonclassical MHC genes, since other genes in the MHC have alleles which affect many biological functions.
Significance
Natural killer (NK) cells are known for their role in defense against viruses and cancer. Their activity is regulated, in part, by killer cell immunoglobulin-like receptors (KIRs) that bind to polymorphic human leukocyte antigen (HLA) class I molecules. The KIR family includes an activation receptor of unknown function, KIR2DS4. Here, we show that KIR2DS4 binding to HLA-C*05:01 is dependent on specific peptides that include a Trp at position 8 of 9-mer peptides associated with HLA-C*05:01. Through sequence homology, we identified a highly conserved peptide sequence in bacterial recombinase A that binds to HLA-C*05:01 and stimulates KIR2DS4 ⁺ NK cells. We predict that over 1,000 bacterial species contain this epitope and propose that NK cells contribute also to immune defense against bacteria.
During early human pregnancy the uterine mucosa transforms into the decidua, into which the fetal placenta implants and where placental trophoblast cells intermingle and communicate with maternal cells. Trophoblast–decidual interactions underlie common diseases of pregnancy, including pre-eclampsia and stillbirth. Here we profile the transcriptomes of about 70,000 single cells from first-trimester placentas with matched maternal blood and decidual cells. The cellular composition of human decidua reveals subsets of perivascular and stromal cells that are located in distinct decidual layers. There are three major subsets of decidual natural killer cells that have distinctive immunomodulatory and chemokine profiles. We develop a repository of ligand–receptor complexes and a statistical tool to predict the cell-type specificity of cell–cell communication via these molecular interactions. Our data identify many regulatory interactions that prevent harmful innate or adaptive immune responses in this environment. Our single-cell atlas of the maternal–fetal interface reveals the cellular organization of the decidua and placenta, and the interactions that are critical for placentation and reproductive success.
Killer cell immunoglobulin-like receptors (KIR) are rapidly evolving species-specific natural killer cell receptors associated with protection against multiple different human viral infections. We report that the activating receptor KIR2DS2 directly recognizes viral peptides derived from conserved regions of flaviviral superfamily 2 RNA helicases in the context of MHC class I. The peptide LNPSVAATL, from the HCV helicase, binds HLA-C*0102 leading to NK cell activation through engagement of KIR2DS2. Similarly, HLA-C*0102 presents
highly conserved peptides from the helicase motif 1b region of related flaviviruses, including dengue, Zika, yellow fever and Japanese encephalitis viruses, to KIR2DS2. These flaviviral peptides all contain an “MCHAT” motif, which is present in 61 out of 63 flaviviruses. LNPSVAATL is also highly conserved across HCV genotypes and mutation of this epitope is poorly tolerated by HCV. KIR2DS2 recognizes endogenously presented helicase peptides and KIR2DS2 is sufficient to inhibit HCV and dengue virus replication in the context of HLAC*
0102. Targeting short, but highly conserved, viral peptides provide non-rearranging innate immune receptors with an efficient mechanism to recognize multiple, highly variable pathogenic RNA viruses.
Introduction:
KIR2DS5 is an activating human NK cell receptor of lineage III KIR. These include both inhibitory KIR2DL1, 2 and 3 and activating KIR2DS1 that recognize either the C1 or C2 epitope of HLA-C. In Europeans KIR2DS5 is essentially monomorphic, with KIR2DS5*002 being predominant. Pioneering investigations showed that KIR2DS5*002 has activating potential, but cannot recognize HLA-A, -B, or -C. Subsequent studies have shown that KIR2DS5 is highly polymorphic in Africans, and that KIR2DS5*006 protects pregnant Ugandan women from preeclampsia. Because inhibitory C2-specific KIR2DL1 correlates with preeclampsia, whereas activating C2-specific KIR2DS1 protects, this association pointed to KIR2DS5*006 being an activating C2-specific receptor. To test this hypothesis we made KIR-Fc fusion proteins from all ten KIR2DS5 allotypes and tested their binding to a representative set of HLA-A, -B and -C allotypes.
Results:
Six African-specific KIR2DS5 bound to C2(+) HLA-C but not to other HLA class I. Their avidity for C2 is ∼20% that of C2-specific KIR2DL1 and ∼40% that of C2-specific KIR2DS1. Among the African C2 receptors is KIR2DS5*006, which protected a cohort of pregnant Ugandans from pre-eclampsia. Three African KIR2DS5 allotypes and KIR2DS5*002, bound no HLA-A, -B or -C. As a group the C2-binding KIR2DS5 allotypes protect against pre-eclampsia compared to the non-binding KIR2DS5 allotypes. Natural substitutions that contribute to loss or reduction of C2 receptor function are at positions 127, 158, and 176 in the D2 domain.
Conclusions:
KIR2DS5*005 has the KIR2DS5 consensus sequence, is the only allele found at both centromeric and telomeric locations of KIR2DS5, and is likely the common ancestor of all KIR2DS5 alleles. That KIR2DS5*005 has C2 receptor activity, points to KIR2DS5*002, and other allotypes lacking C2 receptor function, being products of attenuation, a characteristic feature of most KIR B haplotype genes. Alleles encoding attenuated and active KIR2DS5 are present in both centromeric and telomeric locations.
Most humans become infected with Epstein-Barr virus (EBV), which then persists for life. Infrequently, EBV infection causes infectious mononucleosis (IM) or Burkitt lymphoma (BL). Type I EBV infection, particularly type I BL, stimulates strong responses of innate immune cells. Humans respond to EBV in two alternative ways. Of 24 individuals studied, 13 made strong NK and γδ T cell responses, whereas 11 made feeble γδ T cell responses but stronger NK cell responses. The difference does not correlate with sex, HLA type, or previous exposure to EBV or cytomegalovirus. Cohorts of EBV(+) children and pediatric IM patients include both group 1 individuals, with high numbers of γδ T cells, and group 2 individuals, with low numbers. The even balance of groups 1 and 2 in the human population points to both forms of innate immune response to EBV having benefit for human survival. Correlating these distinctive responses with the progress of EBV infection might facilitate the management of EBV-mediated disease.
γδ T lymphocytes are unconventional immune cells, which have both innate- and adaptive-like features allowing them to respond to a wide spectrum of pathogens. For many years, we and others have reported on the role of these cells in the immune response to human cytomegalovirus in transplant patients, pregnant women, neonates, immunodeficient children, and healthy people. Indeed, and as described for CD8⁺ T cells, CMV infection leaves a specific imprint on the γδ T cell compartment: (i) driving a long-lasting expansion of oligoclonal γδ T cells in the blood of seropositive individuals, (ii) inducing their differentiation into effector/memory cells expressing a TEMRA phenotype, and (iii) enhancing their antiviral effector functions (i.e., cytotoxicity and IFNγ production). Recently, two studies using murine CMV (MCMV) have corroborated and extended these observations. In particular, they have illustrated the ability of adoptively transferred MCMV-induced γδ T cells to protect immune-deficient mice against virus-induced death. In vivo, expansion of γδ T cells is associated with the clearance of CMV infection as well as with reduced cancer occurrence or leukemia relapse risk in kidney transplant patients and allogeneic stem cell recipients, respectively. Taken together, all these studies show that γδ T cells are important immune effectors against CMV and cancer, which are life-threatening diseases affecting transplant recipients. The ability of CMV-induced γδ T cells to act independently of other immune cells opens the door to the development of novel cellular immunotherapies that could be particularly beneficial for immunocompromised transplant recipients.
Natural killer (NK) cells are lymphocytes that have vital functions in innate and adaptive immunity, as well as placental reproduction. Polymorphic human leukocyte antigen (HLA) class I educates NK cells through interactions with killer cell immunoglobulin-like receptors (KIRs) and by supplying peptides that bind HLA-E to form ligands for CD94/NKG2A receptors on NK cells. HLA-B dimorphism in the leader peptide modulates this latter function: −21methionine (−21M) delivers functional peptides, but −21threonine (−21T) does not. Genetic analysis of human populations worldwide showed that haplotypes with −21M HLA-B rarely encoded the KIR ligands Bw4⁺HLA-B and C2⁺HLA-C KIR. Thus, there are two fundamental forms of HLA haplotype: one preferentially supplying CD94/NKG2A ligands and the other preferentially supplying KIR ligands. This −21 HLA-B dimorphism divides the human population into three groups: M/M, M/T, and T/T. Mass cytometry and assays of immune function demonstrated that M/M and M/T individuals have CD94/NKG2A⁺ NK cells that are better educated, phenotypically more diverse, and functionally more potent than those in T/T individuals. The KIR school of NK cell education evolved in the context of the much older CD94/NKG2A school, and this complementary coevolution may have facilitated the specialization of HLA haplotypes toward one school or the other.
The activating natural killer (NK)-cell receptor KIR3DS1 has been linked to the outcome of various human diseases, including delayed progression of disease caused by human immunodeficiency virus type 1 (HIV-1), yet a ligand that would account for its biological effects has remained unknown. We screened 100 HLA class I proteins and found that KIR3DS1 bound to HLA-F, a result we confirmed biochemically and functionally. Primary human KIR3DS1(+) NK cells degranulated and produced antiviral cytokines after encountering HLA-F and inhibited HIV-1 replication in vitro. Activation of CD4(+) T cells triggered the transcription and surface expression of HLA-F mRNA and HLA-F protein, respectively, and induced binding of KIR3DS1. HIV-1 infection further increased the transcription of HLA-F mRNA but decreased the binding of KIR3DS1, indicative of a mechanism for evading recognition by KIR3DS1(+) NK cells. Thus, we have established HLA-F as a ligand of KIR3DS1 and have demonstrated cell-context-dependent expression of HLA-F that might explain the widespread influence of KIR3DS1 in human disease.
Natural killer (NK) cells play a key role in immunity, but how HLA class I (HLA-I) and killer cell immunoglobulin-like receptor 3DL1 (KIR3DL1) polymorphism impacts disease outcome remains unclear. KIR3DL1 (*001/*005/*015) tetramers were screened for reactivity against a panel of HLA-I molecules. This revealed different and distinct hierarchies of specificity for each KIR3DL1 allotype, with KIR3DL1*005 recognizing the widest array of HLA-I ligands. These differences were further reflected in functional studies using NK clones expressing these specific KIR3DL1 allotypes. Unexpectedly, the Ile/Thr80 dimorphism in the Bw4-motif did not categorically define strong/weak KIR3DL1 recognition. Although the KIR3DL1*001, *005, and *015 polymorphisms are remote from the KIR3DL1–HLA-I interface, the structures of these three KIR3DL1–HLA-I complexes showed that the broader HLA-I specificity of KIR3DL1*005 correlated with an altered KIR3DL1*005 interdomain positioning and increased mobility within its ligand-binding site. Collectively, we provide a generic framework for understanding the impact of KIR3DL1 polymorphism on the recognition of HLA-I allomorphs.
KIR2DL2 and KIR2DL3 segregate as alleles of a single locus in the centromeric motif of the killer cell immunoglobulin-like receptor (KIR) gene family. Although KIR2DL2/L3 polymorphism is known to be associated with many human diseases and is an important factor for donor selection in allogeneic hematopoietic stem cell transplantation, the molecular determinant of functional diversity among various alleles is unclear. In this study we found that KIR2DL2/L3 with glutamic acid at position 35 (E35) are functionally stronger than those with glutamine at the same position (Q35). Cytotoxicity assay showed that NK cells from HLA-C1 positive donors with KIR2DL2/L3-E35 could kill more target cells lacking their ligands than NK cells with the weaker -Q35 alleles, indicating better licensing of KIR2DL2/L3+ NK cells with the stronger alleles. Molecular modeling analysis reveals that the glutamic acid, which is negatively charged, interacts with positively charged histidine located at position 55, thereby stabilizing KIR2DL2/L3 dimer and reducing entropy loss when KIR2DL2/3 binds to HLA-C ligand. The results of this study will be important for future studies of KIR2DL2/L3-associated diseases as well as for donor selection in allogeneic stem cell transplantation.
Most of the polymorphic amino acids of the class I histocompatibility antigen, HLA-A2, are clustered on top of the molecule in a large groove identified as the recognition site for processed foreign antigens. Many residues critical for T-cell recognition of HLA are located in this site, in positions allowing them to serve as ligands to processed antigens. These findings have implications for how the products of the major histocompatibility complex (MHC) recognize foreign antigens.
The human killer cell Ig-like receptor (KIR) locus comprises two groups of KIR haplotypes, termed A and B. These are present in all human populations but with different relative frequencies, suggesting they have different functional properties that underlie their balancing selection. We studied the genomic organization and functional properties of the alleles of the inhibitory and activating HLA-C receptors encoded by KIR haplotypes. Because every HLA-C allotype functions as a ligand for KIR, the interactions between KIR and HLA-C dominate the HLA class I-mediated regulation of human NK cells. The C2 epitope is recognized by inhibitory KIR2DL1 and activating KIR2DS1, whereas the C1 epitope is recognized by inhibitory KIR2DL2 and KIR2DL3. This study shows that the KIR2DL1, KIR2DS1, and KIR2DL2/3 alleles form distinctive phylogenetic clades that associate with specific KIR haplotypes. KIR A haplotypes are characterized by KIR2DL1 alleles that encode strong inhibitory C2 receptors and KIR2DL3 alleles encoding weak inhibitory C1 receptors. In striking contrast, KIR B haplotypes are characterized by KIR2DL1 alleles that encode weak inhibitory C2 receptors and KIR2DL2 alleles encoding strong inhibitory C1 receptors. The wide-ranging properties of KIR allotypes arise from substitutions throughout the KIR molecule. Such substitutions can influence cell surface expression, as well as the avidity and specificity for HLA-C ligands. Consistent with the crucial role of inhibitory HLA-C receptors in self-recognition, as well as NK cell education and response, most KIR haplotypes have both a functional C1 and C2 receptor, despite the considerable variation that occurs in ligand recognition and surface expression.
Copyright © 2015 by The American Association of Immunologists, Inc.
Modulating natural killer cell functions in human immunity and reproduction are diverse interactions between the killer cell immunoglobulin-like receptors (KIR) of Natural Killer (NK) cells and HLA class I ligands on the surface of tissue cells. Dominant interactions are between KIR2DL1 and the C2 epitope of HLA-C and between KIR2DL2/3 and the C1 epitope of HLA-C. KhoeSan hunter-gatherers of Southern Africa represent the earliest population divergence known and are the most genetically diverse indigenous people, qualities reflected in their KIR and HLA genes. Of the ten KhoeSan KIR2DL1 alleles, KIR2DL1*022 and KIR2DL1*026 likely originated in the KhoeSan, and later were transmitted at low frequency to the neighboring Zulus through gene flow. These alleles arose by point mutation from other KhoeSan KIR2DL1 alleles that are more widespread globally. Mutation of KIR2DL1*001 gave rise to KIR2DL1*022, causing loss of C2 recognition and gain of C1 recognition. This makes KIR2DL1*022 a more avid and specific C1 receptor than any KIR2DL2/3 allotype. Mutation of KIR2DL1*012 gave rise to KIR2DL1*026, causing premature termination of translation at the end of the transmembrane domain. This makes KIR2DL1*026 a membrane-associated receptor that lacks both a cytoplasmic tail and signaling function. At higher frequencies than their parental allotypes, the combined effect of the KhoeSan-specific KIR2DL1*022 and KIR2DL1*026 is to reduce the frequency of strong inhibitory C2 receptors and increase the frequency of strong inhibitory C1 receptors. Because interaction of KIR2DL1 with C2 is associated with risk of pregnancy disorder, these functional changes are potentially advantageous. Whereas all other KhoeSan KIR2DL1 alleles are present on a wide diversity of centromeric KIR haplotypes, KIR2DL1*026 is present on a single KIR haplotype and KIR2DL1*022 is present on two very similar haplotypes. The high linkage disequilibrium across their haplotypes is consistent with a recent emergence for these KIR2DL1 alleles that have distinctive functions
Natural kiUer (NK) cell dones have been previously described which are inhibited by HLA-C aUdes with Asn77-LysS0 (NKl-specific cdls) or by HLA-C alldes with Ser77-Asn80 (NK2-specific ceils). In the present work, the generation of NK cells with HLA-B-rdated spedficities was attempted by stimulation of a Bw4 homozygous responder by a Bw6 homozygous donor. Two NK clones were found, which were inhibited by HLA-Bw4 (but not by HLA-Bw6) allotypes and by some HLA-A allotypes that share the Bw4 public epitope. Inhibition of NK cell-mediated lysis strongly correlated with the presence of an Ile residue at position 80 of the protective allele. These NK cdl dones define a new specificity termed NK3.
Significance
Pre-eclampsia is especially common in women of African ancestry and a major cause of maternal death. The killer-cell immunoglobulin-like receptor ( KIR ) genes that we analyzed are expressed by natural killer cells—immune cells that populate the uterus and are essential for successful pregnancy. KIR proteins bind HLA ligands on the implanting placental trophoblast cells. African and European women share similar risk associations for pre-eclampsia, but protection is associated with different KIR genes. African women are protected by a combination of KIR B haplotype genes that is present almost exclusively in Africans. This study emphasizes the importance of studying diseases in Africans, where the KIR / HLA genetic system is at its most diverse and maternal mortality rates are the highest in the world.
αβ and γδ T cells are disparate T cell lineages that can respond to distinct antigens (Ags) via the use of the αβ and γδ T cell Ag receptors (TCRs), respectively. Here we characterize a population of human T cells, which we term δ/αβ T cells, expressing TCRs comprised of a TCR-δ variable gene (Vδ1) fused to joining α and constant α domains, paired with an array of TCR-β chains. We demonstrate that these cells, which represent ∼50% of all Vδ1(+) human T cells, can recognize peptide- and lipid-based Ags presented by human leukocyte antigen (HLA) and CD1d, respectively. Similar to type I natural killer T (NKT) cells, CD1d-lipid Ag-reactive δ/αβ T cells recognized α-galactosylceramide (α-GalCer); however, their fine specificity for other lipid Ags presented by CD1d, such as α-glucosylceramide, was distinct from type I NKT cells. Thus, δ/αβTCRs contribute new patterns of Ag specificity to the human immune system. Furthermore, we provide the molecular bases of how δ/αβTCRs bind to their targets, with the Vδ1-encoded region providing a major contribution to δ/αβTCR binding. Our findings highlight how components from αβ and γδTCR gene loci can recombine to confer Ag specificity, thus expanding our understanding of T cell biology and TCR diversity.
© 2014 Pellicci et al.
The Immuno Polymorphism Database (IPD) was developed to provide a centralized system for the study of polymorphism in genes of the immune system. Through the IPD project we have established a central platform for the curation and publication of locus-specific databases involved either directly or related to the function of the Major Histocompatibility Complex in a number of different species. We have collaborated with specialist groups or nomenclature committees that curate the individual sections before they are submitted to IPD for online publication. IPD consists of five core databases, with the IMGT/HLA Database as the primary database. Through the work of the various nomenclature committees, the HLA Informatics Group and in collaboration with the European Bioinformatics Institute we are able to provide public access to this data through the website http://www.ebi.ac.uk/ipd/. The IPD project continues to develop with new tools being added to address scientific developments, such as Next Generation Sequencing, and to address user feedback and requests. Regular updates to the website ensure that new and confirmatory sequences are dispersed to the immunogenetics community, and the wider research and clinical communities.
© The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.
Epstein-Barr virus-induced lymphoproliferative disease (EBV-LPD) after transplantation remains a serious and life-threatening complication. Herein we showed that the aminobisphosphonate pamidronate-expanded human Vγ9Vδ2-T cells efficiently killed EBV-transformed autologous lymphoblastoid B cell lines (EBV-LCL) through γ/δ-TCR and NKG2D receptor triggering and Fas and TRAIL engagement. By inoculation of EBV-LCL in Rag2(-/-)γc(-/-) mice and humanized mice, we established lethal EBV-LPD with characteristics close to those of the human disease. Adoptive transfer of pamidronate-expanded Vγ9Vδ2-T cells alone effectively prevented EBV-LPD in Rag2(-/-)γc(-/-) mice and induced EBV-LPD regression in EBV(+) tumor-bearing Rag2(-/-)γc(-/-) mice. Pamidronate treatment inhibited EBV-LPD development in humanized mice through selective activation and expansion of Vγ9Vδ2-T cells. This study provides proof-of-principle for a therapeutic approach using pamidronate to control EBV-LPD through Vγ9Vδ2-T cell targeting.
HLA class I molecules and killer cell immunoglobulin-like receptors (KIR) form a diverse system of ligands and receptors that individualize human immune systems in ways that improve the survival of individuals and populations. Human settlement of Oceania by island-hopping East and Southeast Asian migrants started ~3,500 years ago. Subsequently, New Zealand was reached ~750 years ago by ancestral Māori. To examine how this history impacted KIR and HLA diversity, and their functional interaction, we defined at high resolution the allelic and haplotype diversity of the 13 expressed KIR genes in 49 Māori and 34 Polynesians. Eighty KIR variants, including four 'new' alleles, were defined, as were 35 centromeric and 22 telomeric KIR region haplotypes, which combine to give >50 full-length KIR haplotypes. Two new and divergent variant KIR form part of a telomeric KIR haplotype, which appears derived from Papua New Guinea and was probably obtained by the Asian migrants en route to Polynesia. Māori and Polynesian KIR are very similar, but differ significantly from African, European, Japanese, and Amerindian KIR. Māori and Polynesians have high KIR haplotype diversity with corresponding allotype diversity being maintained throughout the KIR locus. Within the population, each individual has a unique combination of HLA class I and KIR. Characterizing Māori and Polynesians is a paucity of HLA-B allotypes recognized by KIR. Compensating for this deficiency are high frequencies (>50 %) of HLA-A allotypes recognized by KIR. These HLA-A allotypes are ones that modern humans likely acquired from archaic humans at a much earlier time.
Human birth weight is subject to stabilizing selection; babies born too small or too large are less likely to survive. Particular combinations of maternal/fetal immune system genes are associated with pregnancies where the babies are ≤5th birth weight centile, specifically an inhibitory maternal KIR AA genotype with a paternally derived fetal HLA-C2 ligand. We have now analyzed maternal KIR and fetal HLA-C combinations at the opposite end of the birth weight spectrum. Mother/baby pairs (n = 1316) were genotyped for maternal KIR as well as fetal and maternal HLA-C. Presence of a maternal-activating KIR2DS1 gene was associated with increased birth weight in linear or logistic regression analyses of all pregnancies >5th centile (p = 0.005, n = 1316). Effect of KIR2DS1 was most significant in pregnancies where its ligand, HLA-C2, was paternally but not maternally inherited by a fetus (p = 0.005, odds ratio = 2.65). Thus, maternal KIR are more frequently inhibitory with small babies but activating with big babies. At both extremes of birth weight, the KIR associations occur when their HLA-C2 ligand is paternally inherited by a fetus. We conclude that the two polymorphic immune gene systems, KIR and HLA-C, contribute to successful reproduction by maintaining birth weight between two extremes with a clear role for paternal HLA.
Significance
Killer cell immunoglobulin-like receptors (KIRs) function as key recognition elements in innate immunity. Structural information for inhibitory KIRs 2DL2, 2DL1, and 3DL1 in complex with their respective HLA ligands is available, but such data for activating KIRs are lacking. We report here the successful crystallization and solved structure of the activating KIR2DS2 in complex with HLA-A*11:01. The structure clearly explains the role of Tyr45, which has long puzzled KIR researchers because it differentiates KIR2DS2 from all inhibitory KIRs, and is now shown to bind Thr80 of HLA-A*11:01. Using KIR2DS2 tetramers to bind HLA on live cells, we also provide evidence that peptide sequence can affect KIR–HLA binding. Our data thus resolve a long-standing problem in KIR biology.
Interactions between HLA class I molecules and killer-cell immunoglobulin-like receptors (KIR) control natural killer cell (NK) functions in immunity and reproduction. Encoded by genes on different chromosomes, these polymorphic ligands and receptors correlate highly with disease resistance and susceptibility. Although studied at low-resolution in many populations, high-resolution analysis of combinatorial diversity of HLA class I and KIR is limited to Asian and Amerindian populations with low genetic diversity. At the other end of the spectrum is the West African population investigated here: we studied 235 individuals, including 104 mother-child pairs, from the Ga-Adangbe of Ghana. This population has a rich diversity of 175 KIR variants forming 208 KIR haplotypes, and 81 HLA-A, -B and -C variants forming 190 HLA class I haplotypes. Each individual we studied has a unique compound genotype of HLA class I and KIR, forming 1-14 functional ligand-receptor interactions. Maintaining this exceptionally high polymorphism is balancing selection. The centromeric region of the KIR locus, encoding HLA-C receptors, is highly diverse whereas the telomeric region encoding Bw4-specific KIR3DL1, lacks diversity in Africans. Present in the Ga-Adangbe are high frequencies of Bw4-bearing HLA-B*53:01 and Bw4-lacking HLA-B*35:01, which otherwise are identical. Balancing selection at key residues maintains numerous HLA-B allotypes having and lacking Bw4, and also those of stronger and weaker interaction with LILRB1, a KIR-related receptor. Correspondingly, there is a balance at key residues of KIR3DL1 that modulate its level of cell-surface expression. Thus, capacity to interact with NK cells synergizes with peptide binding diversity to drive HLA-B allele frequency distribution. These features of KIR and HLA are consistent with ongoing co-evolution and selection imposed by a pathogen endemic to West Africa. Because of the prevalence of malaria in the Ga-Adangbe and previous associations of cerebral malaria with HLA-B*53:01 and KIR, Plasmodium falciparum is a candidate pathogen.
Reduced trophoblast invasion and vascular conversion in decidua are thought to be the primary defect of common pregnancy disorders including preeclampsia and fetal growth restriction. Genetic studies suggest these conditions are linked to combinations of polymorphic killer cell Ig-like receptor (KIR) genes expressed by maternal decidual NK cells (dNK) and HLA-C genes expressed by fetal trophoblast. Inhibitory KIR2DL1 and activating KIR2DS1 both bind HLA-C2, but confer increased risk or protection from pregnancy disorders, respectively. The mechanisms underlying these genetic associations with opposing outcomes are unknown. We show that KIR2DS1 is highly expressed in dNK, stimulating strong activation of KIR2DS1+ dNK. We used microarrays to identify additional responses triggered by binding of KIR2DS1 or KIR2DL1 to HLA-C2 and found different responses in dNK coexpressing KIR2DS1 with KIR2DL1 compared with dNK only expressing KIR2DL1. Activation of KIR2DS1+ dNK by HLA-C2 stimulated production of soluble products including GM-CSF, detected by intracellular FACS and ELISA. We demonstrated that GM-CSF enhanced migration of primary trophoblast and JEG-3 trophoblast cells in vitro. These findings provide a molecular mechanism explaining how recognition of HLA class I molecules on fetal trophoblast by an activating KIR on maternal dNK may be beneficial for placentation.
The interactions of killer Ig-like receptor 2D (KIR2D) with HLA-C ligands contribute to functional NK cell education and regulate NK cell functions. Although simple alloreactive rules have been established for inhibitory KIR2DL, those governing activating KIR2DS function are still undefined, and those governing the formation of the KIR2D repertoire are still debated. In this study, we investigated the specificity of KIR2DL1/2/3 and KIR2DS1/2, dissected each KIR2D function, and assessed the impact of revisited specificities on the KIR2D NK cell repertoire formation from a large cohort of 159 KIR and HLA genotyped individuals. We report that KIR2DL2(+) and KIR2DL3(+) NK cells reacted similarly against HLA-C(+) target cells, irrespective of C1 or C2 allele expression. In contrast, KIR2DL1(+) NK cells specifically reacted against C2 alleles, suggesting a larger spectrum of HLA-C recognition by KIR2DL2 and KIR2DL3 than KIR2DL1. KIR2DS2(+) KIR2DL2(-) NK cell clones were C1-reactive irrespective of their HLA-C environment. However, when KIR2DS2 and KIR2DL2 were coexpressed, NK cell inhibition via KIR2DL2 overrode NK cell activation via KIR2DS2. In contrast, KIR2DL1 and KIR2DS2 had an additive enhancing effect on NK cell responses against C1C1 target cells. KIR2DL2/3/S2 NK cells predominated within the KIR repertoire in KIR2DL2/S2(+) individuals. In contrast, the KIR2DL1/S1 NK cell compartment is dominant in C2C2 KIR2DL2/S2(-) individuals. Moreover, our results suggest that together with KIR2DL2, activating KIR2DS1 and KIR2DS2 expression limits KIR2DL1 acquisition on NK cells. Altogether, our results suggest that the NK cell repertoire is remolded by the activating and inhibitory KIR2D and their cognate ligands.
Human T cells that express a T cell antigen receptor (TCR) containing γ-chain variable region 9 and δ-chain variable region 2 (Vγ9Vδ2) recognize phosphorylated prenyl metabolites as antigens in the presence of antigen-presenting cells but independently of major histocompatibility complex (MHC), the MHC class I-related molecule MR1 and antigen-presenting CD1 molecules. Here we used genetic approaches to identify the molecule that binds and presents phosphorylated antigens. We found that the butyrophilin BTN3A1 bound phosphorylated antigens with low affinity, at a stoichiometry of 1:1, and stimulated mouse T cells with transgenic expression of a human Vγ9Vδ2 TCR. The structures of the BTN3A1 distal domain in complex with host- or microbe-derived phosphorylated antigens had an immunoglobulin-like fold in which the antigens bound in a shallow pocket. Soluble Vγ9Vδ2 TCR interacted specifically with BTN3A1-antigen complexes. Accordingly, BTN3A1 represents an antigen-presenting molecule required for the activation of Vγ9Vδ2 T cells.
Human Vγ9Vδ2 T cells respond to microbial infections and malignancy by sensing diphosphate-containing metabolites called phosphoantigens, which bind to the intracellular domain of butyrophilin 3A1, triggering extracellular interactions with the Vγ9Vδ2 T cell receptor (TCR). Here, we examined the molecular basis of this “inside-out” triggering mechanism. Crystal structures of intracellular butyrophilin 3A proteins alone or in complex with the potent microbial phosphoantigen HMBPP or a synthetic analog revealed key features of phosphoantigens and butyrophilins required for γδ T cell activation. Analyses with chemical probes and molecular dynamic simulations demonstrated that dimerized intracellular proteins cooperate in sensing HMBPP to enhance the efficiency of γδ T cell activation. HMBPP binding to butyrophilin doubled the binding force between a γδ T cell and a target cell during “outside” signaling, as measured by single-cell force microscopy. Our findings provide insight into the “inside-out” triggering of Vγ9Vδ2 T cell activation by phosphoantigen-bound butyrophilin, facilitating immunotherapeutic drug design.
γδ T cells have been retained as a lineage over the majority of vertebrate evolution, are able to respond to immune challenges in unique ways, and are of increasing therapeutic interest. However, one central mystery has endured: the identity of the ligands recognized by the γδ T cell antigen receptor. Here we discuss the inherent challenges in answering this question, the new opportunities provided by recent studies, and the criteria by which the field might judge success. In this Perspective, Willcox and Willcox discuss the inherent challenges and future possibilities in identifying the ligands recognized by the γδ TCR.
Immunoprecipitates of the T3 antigen prepared from HPB‐ALL cells by using the monoclonal antibody UCH‐T1 were analysed by SDS‐polyacrylamide gel electrophoresis. Cells which had been biosynthetically labelled for up to 4 h gave a major polypeptide of mol. wt. 19 000 plus two weaker, more diffuse bands of mol. wts. 21 000 and 23 000, whereas surface labelled cells gave a prominent band of mol. wt. 19 000, a major band of 21 000 and a weaker diffuse band of approximately 26 000. As judged from their sensitivity to proteinase‐K digestion, all the above polypeptides possess a transmembrane orientation. Digestion with endoglycosidases H and F (endo‐H and endo‐F), and tunicamycin treatment indicate that all the polypeptides, except that of 19 000 mol. wt. are N‐glycosylated. The 21 000 and 23 000 mol. wt. chains possess both immature and mature oligosaccharide units, whereas the 26 000 mol. wt. band apparently has mature units only. Pulse chase experiments combined with digestion by endo‐F and endo‐H suggest that the N‐glycosylated polypeptides are derived from two polypeptides of mol. wts. 14 000 and 16 000. It is concluded that the T3 antigen is derived from three different non‐glycosylated polypeptides two of which are subsequently N‐glycosylated to give the 21 000, 23 000 and 26 000 forms. The cell surface T3 antigen most probably comprises at least two distinct, non‐covalently associated polypeptides, but the number and types of polypeptides giving rise to the whole molecule and whether different complexes exist is at present unclear.
The remarkable T cell receptor (TCR) performs essential functions in the initiation of intracellular signals required for T cell development, repertoire selection and effector responses to foreign antigens. How TCR signals elicit such diverse cellular responses and outcomes remains a major question for investigation. Recent years have witnessed important advances in our understanding of the regulatory processes that control and modulate the TCR signalling response. Here, we review newly identified mechanisms for the regulation of TCR signalling and then discuss how the TCR signalling response is regulated to control two critical cellular processes - namely, positive selection and T cell homeostasis.
Epithelial surfaces line the body and provide a crucial interface between the body and the external environment. Tissue-resident epithelial γδ T cells represent a major T cell population in the epithelial tissues and are ideally positioned to carry out barrier surveillance and aid in tissue homeostasis and repair. In this Review, we focus on the intraepithelial γδ T cell compartment of the two largest epithelial tissues in the body — namely, the epidermis and the intestine — and provide a comprehensive overview of the crucial contributions of intraepithelial γδ T cells to tissue integrity and repair, host homeostasis and protection in the context of the symbiotic relationship with the microbiome and during pathogen clearance. Finally, we describe epithelium-specific butyrophilin-like molecules and briefly review their emerging role in selectively shaping and regulating epidermal and intestinal γδ T cell repertoires.
Preeclampsia is a serious complication of pregnancy in which the fetus receives an inadequate supply of blood due to failure of trophoblast invasion. There is evidence that the condition has an immunological basis. The only known polymorphic histocompatibility antigens on the fetal trophoblast are HLA-C molecules. We tested the idea that recognition of these molecules by killer immunoglobulin receptors (KIRs) on maternal decidual NK cells is a key factor in the development of preeclampsia. Striking differences were observed when these polymorphic ligand: receptor pairs were considered in combination. Mothers lacking most or all activating KIR (AA genotype) when the fetus possessed HLA-C belonging to the HLA-C2 group were at a greatly increased risk of preeclampsia. This was true even if the mother herself also had HLA-C2, indicating that neither nonself nor missing-self discrimination was operative. Thus, this interaction between maternal KIR and trophoblast appears not to have an immune function, but instead plays a physiological role related to placental development. Different human populations have a reciprocal relationship between AA frequency and HLA-C2 frequency, suggesting selection against this combination. In light of our findings, reproductive success may have been a factor in the evolution and maintenance of human HLA-C and KIR polymorphisms.
Problem:
The extravillous trophoblasts (EVT) express HLA-C and HLA-G, but HLA-E and HLA-F are the subject of conflicting reports. In this study, we define the HLA expression profile during active EVT placental implantation, pregnancy development, and parturition.
Method of study:
Immunohistochemistry, q-PCR, and Western blot were used to investigate HLA-C, HLA-E, and HLA-F placental expression across gestation from the early first trimester, late first trimester, second trimester (n=10 in each), preterm gestation (n=6) to elective term cesarean section and term vaginal deliveries (n=12, 38-41 weeks). EVT explants and Swan71 cells were used to assess HLA-C and HLA-F during active EVT migration.
Results:
HLA-G, HLA-C, and HLA-F were expressed by 1st-trimester EVT and became intracellular and weaker as gestation progressed. HLA-E was only expressed in 1st-trimester placenta. HLA-F and HLA-C mRNA and protein expression levels showed a significant increase in the fetal villous mesenchyme across gestation. HLA-C levels increased with labor. We detected a 100-kDa HLA-F band in early pregnancy suggesting dimer formation on the EVT surface. These results were confirmed in EVT outgrowths and Swan71 trophoblast which showed that HLA-F and HLA-G are increased on the cell surface of migrating EVT, while HLA-C was internalized.
Conclusion:
Expression of HLA-F and HLA-G on the cell surface of actively migrating EVT supports their specific role in early EVT invasion and interactions with uterine natural killer cells. HLA-C's limited expression to the proliferative EVT suggests a protective role in the earliest events of implantation but not in active EVT invasion. We also show for the first time that HLA-C may be involved in parturition.
The overrepresentation of particular HLA alleles in patients with celiac disease was first noted two decades ago. Several lines of evidence obtained during the last years strongly suggest that a particular HLA-DQ heterodimer, encoded by the DQA1*0501 and DQB1*0201 genes in cis or trans configuration, confers the primary disease susceptibility. This paper reviews the evidence behind this concept and discusses how this particular DQ molecule may be involved in the pathogenesis.
The interaction of NK inhibitory killer Ig-like receptors (KIRs) with self-MHC class I molecules mediates NK tolerance to self while conferring functional competence. Through single-cell analysis of intracellular IFN-γ production and NK clone cytotoxicity we evaluated the resting NK repertoire, analyzing the responsiveness of NK subgroups expressing discrete combinations of non-KIR and KIR class I-specific receptors. CD94:NKG2A and Ig-like transcript 2 (ILT2)-expressing cells have a modest response to class I-negative target cells, but NK cells expressing inhibitory KIRs to self-MHC class I ligands, both HLA-B and HLA-C ligands, achieve significantly higher effector capacity. There is a dose effect of KIR for self-MHC on effector capacity, but even in the most highly responsive NK cells expressing more than one inhibitory KIR for self-MHC the presentation of only one cognate MHC ligand is sufficient to abolish response. Among KIR⁺ cells there is preferential expression for inhibitory KIR for self-MHC. The likelihood of KIR expression is influenced by whether other KIRs are already expressed on the same cell, supporting a model of serial acquisition of KIR expression. These findings define how inhibitory receptor and autologous HLA interactions impact single-cell function and demonstrate that the resting human NK repertoire is highly attuned but variegated in response. These findings have important implications for the resting NK response to viral pathogens and malignancy, for donor selection in allogeneic hemopoietic cell transplantation, and for models of NK tolerance.
Allogeneic individuals co-exist during pregnancy in eutherian mammals. Maternal and fetal cells intermingle at the site of placental attachment in the uterus, where the arteries are remodeled to supply the fetus with oxygen and nutrients. This access by placental cells to the maternal supply line determines the growth and birth weight of the baby and is subject to stabilizing selection. Invading placental trophoblast cells express human leukocyte antigen class I ligands (HLA-E, HLA-G, and HLA-C) for receptors on maternal uterine natural killer (NK) and myelomonocytic cells, CD94/NKG2, leukocyte immunoglobulin-like receptor (LILR), and killer immunoglobulin receptor (KIR). Of these, only the KIR/HLA-C system is highly polymorphic. Different combinations of maternal KIR and fetal HLA-C variants are correlated with low birth weight and pre-eclampsia or high birth weight and obstructed labor, the two extremes of the obstetric dilemma. This situation has arisen because of the evolution of bipedalism and subsequently, in the last million years, larger brains. At this point, the human system began to reach a balance between KIR A and KIR B haplotypes and C1 and C2 epitopes of HLA-C alleles that reflects a functional compromise between the competing demands of immunity and reproduction.
© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
KIR3DL1 is a polymorphic inhibitory receptor that modulates NK cell activity through interacting with HLA-A and HLA-B alleles that carry the Bw4 epitope. Amino acid polymorphisms throughout KIR3DL1 impact receptor surface expression and affinity for HLA. KIR3DL1/S1 encodes inhibitory and activating alleles, but despite high homology with KIR3DL1, the activating receptor KIR3DS1 does not bind the same ligand. Allele KIR3DL1*009 resulted from a gene recombination event between the inhibitory receptor allele KIR3DL1*001 and the activating receptor allele KIR3DS1*013. This study analyzed the functional impact of KIR3DS1-specific polymorphisms on KIR3DL1*009 surface expression, binding to HLA, and functional capacity. Flow-cytometric analysis of primary human NK cells as well as transfected HEK293T cells shows that KIR3DL1*009 is expressed at a significantly lower surface density compared with KIR3DL1*001. Using recombinant proteins of KIR3DL1*001, KIR3DL1*009, and KIR3DS1*013 to analyze binding to HLA, we found that although KIR3DL1*009 displayed some evidence of binding to HLA compared with KIR3DS1*013, the binding was minimal compared with KIR3DL1*001 and KIR3DL1*005. Mutagenesis of polymorphic sites revealed that the surface phenotype and reduced binding of KIR3DL1*009 are caused by the combined amino acid polymorphisms at positions 58 and 92 within the D0 extracellular domain. Resulting from these effects, KIR3DL1*009(+) NK cells exhibited significantly less inhibition by HLA-Bw4(+) target cells compared with KIR3DL1*001(+) NK cells. The data from this study contribute novel insight into how KIR3DS1-specific polymorphisms in the extracellular region impact KIR3DL1 surface expression, ligand binding, and inhibitory function.
Copyright © 2015 by The American Association of Immunologists, Inc.
Natural killer (NK) cells have been shown to express a clonally distributed ability to recognize HLA class I alleles. The previously defined NK clones belonging to "group 1" recognize HLA-C*0401 (Cw4) and other HLA-C alleles sharing Asn at position 77 and Lys at position 80. Conversely, the "group 2" NK clones recognize HLA-Cw*0302 (Cw3) and other HLA-C alleles characterized by Ser at position 77 and Asn at position 80. We assessed directly the involvement of these two residues in the capacity of NK cell clones to discriminate between the two groups of HLA-C alleles. To this end, Cw3 and Cw4 alleles were subjected to site-directed mutagenesis. Substitution of the amino acids typical of the Cw3 allele (Ser-77 and Asn-80) with those present in Cw4 (Asn-77 and Lys-80) resulted in a Cw3 mutant that was no longer recognized by group 2 NK cell clones, but that was recognized by group 1 clones. Analysis of Cw3 or Cw4 molecules containing single amino acid substitutions indicates roles for Lys-80 in recognition mediated by group 1 clones and for Ser-77 in recognition mediated by group 2 clones. These results demonstrate that NK-mediated specific recognition of HLA-C allotypes is affected by single natural amino acid substitutions at positions 77 and 80 of the heavy chain.
A repertoire of antigen recognition specificities in mature T cell pool is formed by the selection during T cell development in the thymus. Positive selection is an essential process for the development of functionally competent T cells and is dependent on the interaction between T cell antigen receptors (TCRs) that newly generated thymocytes express and self-peptide-associated major histocompatibility complex (pMHC) molecules that cortical thymic epithelial cells (cTECs) express. Characterization of positive-selection-inducing peptides has revealed that the low-affinity TCR engagement by the positive-selection-inducing pMHC complexes initiates intracellular signals that induce the survival of immature thymocytes and their differentiation into mature T cells. Recent studies suggest unique mechanisms of antigen processing in cTECs for the production of positively selecting MHC-bound self-peptides.
© 2015 Elsevier Inc. All rights reserved.
IN a previous paper (Gorer, 1936) it was shown that antigenic differences in mouse erythrocytes could be detected by means of sera from immunized rabbits. In order to obtain sufficient quantities of antigenically -uniform material for immunization and the performance of absorption tests, it is necessary to use pooled cells from individuals drawn from strains that have been genetically purified by continuous brother-sister matings. Three such strains were used, described as the agoutis, albinos and blacks. Anti-agouti sera absorbed once with the cells of blacks still agglutinated the cells of agoutis and albinos. Two absorptions with the cells of blacks or a single absorption with cells from the other two groups exhausted the sera completely. The antibody (or antibodies) remaining after one absorption with the cells of blacks was referred to as antibody I, the cells capable of removing the antibody easily, viz. those of agoutis and albinos, were said to possess the antigen 1(3), while the blacks, whose cells appeared to have less affinity, were said to possess the antigen I(2). Anti-albino sera contained an antibody specific for the albino cells. The cells of agoutis, however, appeared to have slight affinity for the antibody concerned since repeated absorptions lowered the titre of the serum. The cells of blacks appeared to have no affinity for the antibody. On the other hand, animals selected from a heterogeneous stock (the ZS stock) could exhaust the serum completely with two successive absorptions. The albinos were said to possess the antigen 11(3), the agoutis 11(1), the blacks were negative, whilst the ZS had the antigen 11(2). Landsteiner and Levene (1928) found that the tendency of anti-M and anti-N sera to be absorbed non-specifically (i. e. by cells negative in the agglu-tination test) depended on the serum used and not upon the individuality of the cells. Since we had found no cells that could not absorb antibody I, it was impossible to say whether the effect was similar to that described by Landsteiner and Levene. In the case of anti-albino sera it could be shown that the effect of absorption depended on the genetic constitution of the cells and could not therefore be described as non-specific-at least as regards a comparison of the agoutis and blacks. The ZS stock was very heterogeneous, and the effects obtained might have been due to the presence of antigen II(3) in a heterozygous state. Unfortunately the strain is now extinct, but the effects of hybridization between albinos and blacks are reported in this paper.
γδ T cells are a prominent epithelial-resident lymphocyte population, possessing multi-functional capacities in the repair of host tissue, pathogen clearance, and tumor surveillance. Although three decades have now passed since their discovery, the nature of γδ T cell receptor (TCR)-mediated ligand recognition remains poorly defined. Recent studies have provided structural insight into this recognition, demonstrating that γδ T cells survey both CD1 and the presented lipid, and in some cases are exquisitely lipid specific. We review these findings here, examining the molecular basis for and the functional relevance of this interaction. We discuss potential implications on the notion that non-classical major histocompatibility complex (MHC) molecules may function as important restricting elements of γδ TCR specificity, and on our understanding of γδ T cell activation and function.
In humans, Vγ9Vδ2 T cells detect tumor cells and microbial infections, including Mycobacterium tuberculosis, through recognition of small pyrophosphate containing organic molecules known as phosphoantigens (pAgs). Key to pAg-mediated activation of Vγ9Vδ2 T cells is the butyrophilin 3A1 (BTN3A1) protein that contains an intracellular B30.2 domain critical to pAg reactivity. Here, we have demonstrated through structural, biophysical, and functional approaches that the intracellular B30.2 domain of BTN3A1 directly binds pAg through a positively charged surface pocket. Charge reversal of pocket residues abrogates binding and Vγ9Vδ2 T cell activation. We have also identified a gain-of-function mutation within this pocket that, when introduced into the B30.2 domain of the nonstimulatory BTN3A3 isoform, transfers pAg binding ability and Vγ9Vδ2 T cell activation. These studies demonstrate that internal sensing of changes in pAg metabolite concentrations by BTN3A1 molecules is a critical step in Vγ9Vδ2 T cell detection of infection and tumorigenesis.
Human Vγ9Vδ2 T cells recognize phosphorylated products of isoprenoid metabolism (phosphoantigens) PAg with TCR comprising Vγ9JP γ-chains and Vδ2 δ-chains dependent on butyrophilin 3 (BTN3) expressed by antigen-presenting cells. They are massively activated in many infections and show anti-tumor activity and so far, they have been considered to exist only in higher primates. We performed a comprehensive analysis of databases and identified the three genes in species of both placental magnorders, but not in rodents. The common occurrence or loss of in silico translatable Vγ9, Vδ2, and BTN3 genes suggested their co-evolution based on a functional relationship. In the peripheral lymphocytes of alpaca (Vicugna pacos), characteristic Vγ9JP rearrangements and in-frame Vδ2 rearrangements were found and could be co-expressed in a TCR-negative mouse T cell hybridoma where they rescued CD3 expression and function. Finally, database sequence analysis of the extracellular domain of alpaca BTN3 revealed complete conservation of proposed PAg binding residues of human BTN3A1. In summary, we show emergence and preservation of Vγ9 and Vδ2 TCR genes with the gene of the putative antigen-presenting molecule BTN3 in placental mammals and lay the ground for analysis of alpaca as candidate for a first non-primate species to possess Vγ9Vδ2 T cells.
The gcpE and lytB gene products control the terminal steps of isoprenoid biosynthesis via the 2-C-methyl-D-erythritol 4-phosphate pathway in Escherichia coli. In lytB-deficient mutants, a highly immunogenic compound accumulates significantly, compared to wild-type E. coli, but is apparently absent in gcpE-deficient mutants. Here, this compound was purified from E. coli ΔlytB mutants by preparative anion exchange chromatography, and identified by mass spectrometry, 1H, 13C and 31P NMR spectroscopy, and NOESY analysis as (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP). HMB-PP is 104 times more potent in activating human Vγ9/Vδ2 T cells than isopentenyl pyrophosphate.