Recognition Of Glycolipid Antigens Research Articles

Antigen Presentation in an Artificial and Cell Membrane: Blood Group A Glycan Recognition.

Glycosphingolipids (GSL) are functionally important components of the cell membrane and recognition of their glycan "head" by the immune system is a key part of normal and pathological processes. Recognition of glycolipid antigens on a living cell, their structure, "context" (microenvironment and clustering), presentation including orientation and distance from the plasma membrane, as well as molecular dynamics are important. GSL antigens are targets for the development of anticancer vaccines and therapeutic antibodies, therefore, control of the presentation of their glycans by synthetic methods opens up new possibilities in medicine. In this work, we synthesized 13 GSL analogues as function-spacer-lipids (FSLs) with the same head (blood group A tetrasaccharide, ABO system) but each either differing in the structure of the lipid tail, or the length of the region between the head and the tail, or cluster organization of the head (from 2 to 4 tetrasaccharides in a cluster). The insertion of these FSLs into an artificial and erythrocyte membranes was compared, and their interaction with antibodies was studied, including erythrocyte agglutination. The results give further insight into knowledge of the membrane presentation of GSLs (some results can be extrapolated to glycoproteins) and factors involved in their interaction with antibodies.

Glycolipid antigen recognition by invariant natural killer T cells and its role in homeostasis and antimicrobial responses.

Due to the COVID-19 pandemic, the importance of developing effective vaccines has received more attention than ever before. To maximize the effects of vaccines, it is important to select adjuvants that induce strong and rapid innate and acquired immune responses. Invariant natural killer T (iNKT) cells, which constitute a small population among lymphocytes, bypass the innate and acquired immune systems through the rapid production of cytokines after glycolipid recognition; hence, their activation could be used as a vaccine strategy against emerging infectious diseases. Additionally, the diverse functions of iNKT cells, including enhancing antibody production, are becoming more understood in recent years. In this review, we briefly describe the functional subset of iNKT cells and introduce the glycolipid antigens recognized by them. Furthermore, we also introduce novel vaccine development taking advantages of iNKT cell activation against infectious diseases.

Anti-CD1d treatment suppresses immunogenic maturation of lung dendritic cells dependent on lung invariant natural killer T cells in asthmatic mice

Our previous findings show that invariant natural killer T (iNKT)cells can promote immunogenic maturation of lung dendritic cells (LDCs) to enhance Th2 cell responses in asthma. It has been accepted that recognition of glycolipid antigens presented by CD1d molecules by the T cell receptors of iNKT cells leads to iNKT cell activation. Therefore, we examine the immunoregulatory influences of anti-CD1d treatment on Th2 cell response and immunogenic maturation of LDCs and subsequently explored whether these influences were dependent on lung iNKT cells in asthmatic mice. We discoveredthat in wild-type mice sensitized and challenged with house dust mite or ovalbumin (OVA), anti-CD1d treatment inhibited Th2 cell response and immunogenic maturation of LDCs. LDCs from asthmatic mice with anti-CD1d treatment had a markedly decreased influence on Th2 cell responses in vivo and in vitro. Furthermore, anti-CD1d treatment reduced the abundance and activation of lung iNKT cells in asthmatic mice. Moreover, in asthmatic iNKT cell-deficient Jα18-/- mice, anti-CD1d treatment did not influence Th2 cell responses and immunogenic maturation of LDCs. Meanwhile, the quantity of CD40L+ iNKT cells in asthmatic mice was significant decreased by anti-CD1d treatment. Finally, the inhibition of anti-CD1d treatment on LDC immunogenic maturation and Th2 cell responses in asthmatic mice was reversed by anti-CD40 treatment. Our data suggest that anti-CD1d treatment can suppress Th2 cell responses through inhibiting immunogenic maturation of LDCs dependent on lung iNKT cells, which couldbe partially related to the downregulation of CD40L expression on lung iNKT cells in asthmatic mice.

Temporal Regulation of Natural Killer T Cell Interferon Gamma Responses by β-Catenin-Dependent and -Independent Wnt Signaling

Natural killer T (NKT) cells are prominent innate-like lymphocytes in the liver with critical roles in immune responses during infection, cancer, and autoimmunity. Interferon gamma (IFN-γ) and IL-4 are key cytokines rapidly produced by NKT cells upon recognition of glycolipid antigens presented by antigen-presenting cells (APCs). It has previously been reported that the transcriptional coactivator β-catenin regulates NKT cell differentiation and functionally biases NKT cell responses toward IL-4, at the expense of IFN-γ production. β-Catenin is not only a central effector of Wnt signaling but also contributes to other signaling networks. It is currently unknown whether Wnt ligands regulate NKT cell functions. We thus investigated how Wnt ligands and β-catenin activity shape liver NKT cell functions in vivo in response to the glycolipid antigen, α-galactosylceramide (α-GalCer) using a mouse model. Pharmacologic targeting of β-catenin activity with ICG001, as well as myeloid-specific genetic ablation of Wntless (Wls), to specifically target Wnt protein release by APCs, enhanced early IFN-γ responses. By contrast, within several hours of α-GalCer challenge, myeloid-specific Wls deficiency, as well as pharmacologic targeting of Wnt release using the small molecule inhibitor IWP-2 impaired α-GalCer-induced IFN-γ responses, independent of β-catenin activity. These data suggest that myeloid cell-derived Wnt ligands drive early Wnt/β-catenin signaling that curbs IFN-γ responses, but that, subsequently, Wnt ligands sustain IFN-γ expression independent of β-catenin activity. Our analyses in ICG001-treated mice confirmed a role for β-catenin activity in driving early IL-4 responses by liver NKT cells. However, neither pharmacologic nor genetic perturbation of Wnt production affected the IL-4 response, suggesting that IL-4 production by NKT cells in response to α-GalCer is not driven by released Wnt ligands. Collectively, these data reveal complex temporal roles of Wnt ligands and β-catenin signaling in the regulation of liver NKT cell activation, and highlight Wnt-dependent and -independent contributions of β-catenin to NKT cell functions.

Atypical natural killer T-cell receptor recognition of CD1d–lipid antigens

Crucial to Natural Killer T (NKT) cell function is the interaction between their T-cell receptor (TCR) and CD1d-antigen complex. However, the diversity of the NKT cell repertoire and the ensuing interactions with CD1d-antigen remain unclear. We describe an atypical population of CD1d–α-galactosylceramide (α-GalCer)-reactive human NKT cells that differ markedly from the prototypical TRAV10-TRAJ18-TRBV25-1+ type I NKT cell repertoire. These cells express a range of TCR α- and β-chains that show differential recognition of glycolipid antigens. Two atypical NKT TCRs (TRAV21-TRAJ8-TRBV7–8 and TRAV12-3-TRAJ27-TRBV6-5) bind orthogonally over the A′-pocket of CD1d, adopting distinct docking modes that contrast with the docking mode of all type I NKT TCR-CD1d-antigen complexes. Moreover, the interactions with α-GalCer differ between the type I and these atypical NKT TCRs. Accordingly, diverse NKT TCR repertoire usage manifests in varied docking strategies and specificities towards CD1d–α-GalCer and related antigens, thus providing far greater scope for diverse glycolipid antigen recognition.

The role of invariant natural killer T cells in microbial immunity.

Invariant natural killer T cells (iNKT cells) are unique lymphocytes with characteristic features, such as expression of an invariant T-cell antigen receptor (TCR) α-chain, recognition of glycolipid antigens presented by CD1d molecules, and ability to rapidly produce large amounts of cytokines, including interferon-γ (IFN-γ) and interleukin 4 (IL-4) upon TCR stimulation. Many studies have demonstrated that iNKT cells participate in immune response against diverse microbes, including bacteria, fungi, protozoan parasites, and viruses. Generally, these cells play protective roles in host defense against infections. However, in some contexts they play pathogenic roles, by inducing or augmenting inflammation. Recent reports show that iNKT cells recognize glycolipid antigens from pathogenic bacteria including Streptococcus pneumoniae, and they contribute to host defense against infection. iNKT cell responses to these microbial glycolipid antigens are highly conserved between rodents and humans, suggesting that iNKT cells are evolutionally conserved because their invariant TCR is useful in detecting certain pathogens. Furthermore, glycolipid-mediated iNKT cell activation during immunization has adjuvant activity, enhancing humoral and cell-mediated responses. Therefore, iNKT cell activation is an attractive target for developing new vaccines for infectious diseases.

Interplay between carbohydrate and lipid in recognition of glycolipid antigens by natural killer T cells

Natural killer T (NKT) cells are a T cell subpopulation that were named originally based on coexpression of receptors found on natural killer (NK) cells, cells of the innate immune system, and by T lymphocytes. The maturation and activation of NKT cells requires presentation of glycolipid antigens by CD1d, a cell surface protein distantly related to the major histocompatibility complex (MHC)-encoded antigen presenting molecules. This specificity distinguishes NKT cells from most CD4(+) and CD8(+) T cells that recognize peptides presented by MHC class I and class II molecules. The rapid secretion of a large amount of both Th1 and Th2 cytokines by activated NKT cells endows them with the ability to play a vital role in the host immune defense against various microbial infections. In this review, we summarize progress on identifying the sources of microbe-derived glycolipid antigens recognized by NKT cells and the biochemical basis for their recognition.

Microbial glycolipid antigen recognition by invariant natural killer T cells

Microbial glycolipid antigen recognition by invariant natural killer T cells

A semi-invariant Vα10+ T cell antigen receptor defines a population of natural killer T cells with distinct glycolipid antigen–recognition properties

Type I natural killer T cells (NKT cells) are characterized by an invariant variable region 14-joining region 18 (V(α)14-J(α)18) T cell antigen receptor (TCR) α-chain and recognition of the glycolipid α-galactosylceramide (α-GalCer) restricted to the antigen-presenting molecule CD1d. Here we describe a population of α-GalCer-reactive NKT cells that expressed a canonical V(α)10-J(α)50 TCR α-chain, which showed a preference for α-glucosylceramide (α-GlcCer) and bacterial α-glucuronic acid-containing glycolipid antigens. Structurally, despite very limited TCRα sequence identity, the V(α)10 TCR-CD1d-α-GlcCer complex had a docking mode similar to that of type I TCR-CD1d-α-GalCer complexes, although differences at the antigen-binding interface accounted for the altered antigen specificity. Our findings provide new insight into the structural basis and evolution of glycolipid antigen recognition and have notable implications for the scope and immunological role of glycolipid-specific T cell responses.

Antibody responses to glycolipid‐borne carbohydrates require CD4+ T cells but not CD1 or NKT cells

Naturally occurring anti-carbohydrate antibodies play a major role in both the innate and adaptive immune responses. To elicit an anti-carbohydrate immune response, glycoproteins can be processed to glycopeptides and presented by the classical antigen-presenting molecules, major histocompatibility complex (MHC) Class I and II. In contrast, much less is known about the mechanism(s) for anti-carbohydrate responses to glycolipids, although it is generally considered that the CD1 family of cell surface proteins presents glycolipids to T cells or natural killer T (NKT) cells. Using model carbohydrate systems (isogloboside 3 and B blood group antigen), we examined the anti-carbohydrate response on glycolipids using both antibody neutralisation and knockout mouse-based experiments. These studies showed that CD4 T cells were required to generate antibodies to the carbohydrates expressed on glycolipids, and unexpectedly, these antibody responses were CD1d and NKT cell independent. They also did not require peptide help. These data provide new insight into glycolipid antigen recognition by the immune system and indicate the existence of a previously unrecognised population of glycolipid antigen-specific, CD1-independent, CD4 T cells.

Atypical T cell receptor usage defines a population of Natural Killer T cells with distinct glycolipid antigen recognition properties (160.13)

Abstract Lipid Ag reactive T cells represent an important component of the T cell repertoire. The most well studied example is the classical (or Type-1) NKT cells that respond to the glycolipid Ag α-galactosylceramide (α-GalCer) presented by CD1d. While often referred to as 'invariant' due to the expression of an invariant TCR-α chain (Vα14-Jα18), in fact, Type-1 NKT cells carry a diverse TCR-β repertoire that influences Ag recognition. Furthermore, we have identified a completely novel population of NKT cells (Type-1A cells), that expresses a previously unidentified, canonical, Vα10-Jα50 TCR α-chain. These cells exhibit a different pattern of glycolipid Ag reactivity, including a preference for α-glucosylceramide (α-GlcCer) over α-GalCer and a bias towards Th2 cytokine production. Two populations of Type-1A NKT cells (TCRhi and TCRlo) are present in the thymus and they also have diverse TCR-β chains. We also provide the first structural analysis of a non-classical NKT TCR-CD1d-glycolipid complex, showing that the Type-1A TCR-CD1d-α-GlcCer complex displayed a similar docking mode to that of Type-1 NKT TCRs, although differences at the Ag-binding interfaces accounted for the altered specificity. The discovery of a second invariant TCR α-chain expressed by CD1d-restricted NKT cells provides new insights into the repertoire diversity and its influence on glycolipid recognition by NKT cells, and has significant implications for the immunologic role of glycolipid-specific T cell responses.

Antibody responses to glycolipid‐borne carbohydrates require CD4+ T cells but not CD1 or NKT cells

Naturally occurring anti-carbohydrate antibodies play a major role in both the innate and adaptive immune responses. To elicit an anti-carbohydrate immune response, glycoproteins can be processed to glycopeptides and presented by the classical antigen-presenting molecules, major histocompatibility complex (MHC) Class I and II. In contrast, much less is known about the mechanism(s) for anti-carbohydrate responses to glycolipids, although it is generally considered that the CD1 family of cell surface proteins presents glycolipids to T cells or natural killer T (NKT) cells. Using model carbohydrate systems (isogloboside 3 and B blood group antigen), we examined the anti-carbohydrate response on glycolipids using both antibody neutralisation and knockout mouse-based experiments. These studies showed that CD4(+) T cells were required to generate antibodies to the carbohydrates expressed on glycolipids, and unexpectedly, these antibody responses were CD1d and NKT cell independent. They also did not require peptide help. These data provide new insight into glycolipid antigen recognition by the immune system and indicate the existence of a previously unrecognised population of glycolipid antigen-specific, CD1-independent, CD4(+) T cells.

Antigen‐specific recognition of bacterial antigens by NKT cells

The objective of these studies is to understand the biochemistry and biological significance of microbial glycolipid antigen recognition by natural killer T (NKT) cells, a natural memory lymphocyte population that expresses an invariant T cell antigen receptor (TCR) alpha chain. We have identified two types of glycolipids presented by CD1d and recognized by the NKT cell TCR. Sphingomonas, which are environmental bacteria, have antigenic glycosphingolipids (GSLs), containing a ceramide lipid moiety. The second type of lipid is a glycosylated diacyl glycerol found in Borrelia burgdorferi, a spirochete that causes Lyme disease. Interestingly, mouse and human NKT cells selectively respond to diacylglycerols with different acyl chains, as mouse CD1d prefers to present compounds with shorter and less saturated fatty acids. Therefore, the recognition of these antigens is highly sensitive to the nature of the lipid, which is buried in the CD1d groove. To establish the significance of this response, we analyzed NKT cell deficient mice and controls infected with B. burgdorferi. The NKT cell deficient mice had increased joint inflammation at five weeks after infection and reduced bacterial clearance. Our data therefore strongly suggest that the NKT cells participate in host defense against bacterial pathogens through the recognition of microbial glycolipids by their invariant TCR.

T cell receptors get back to basics

For both mice and humans, an invariant T cell receptor mediates the recognition of glycolipid antigens presented in the context of CD1d molecules. Crystal structure and mutagenesis-based analyses now identify a minimalist binding mode well suited to an 'innate-style' pattern-recognition function.

Balanced on One Foot

Cd1 proteins bind lipid antigens and present them to T cells to initiate immune responses. Cd1d presents glycolipids with specific α-linked sugars to the natural killer T cells (NKT) for recognition by their T cell receptor (TCR) complex. Borg et al . report the crystal structure of Cd1 bound to α-galactosylceramide (α-GalCer) with the NKT TCR. They find that the TCR interaction is unlike that of TCRs with the peptide-presenting major histocompatibility proteins (pMHCs). In the case of the pMHC-TCR interaction, the TCR α and β chains both contact the pMHC (see Moody), revealing two binding site "footprints" on the surface of the pMHC. The crystal structures reported by Borg et al . show that NKT TCR only binds to the Cd1d-α-GalCer by the α subunit, with the β subunit showing limited interaction and mostly hanging over the side of Cd1d. Moody describes the two structures as the TCR standing with both feet contacting pMHC, whereas NKT TCR appears as a pirouette balanced on one foot at the Cd1d surface. The structure also shows that the α-linked galactose lies flat, optimizing the interaction surface between Cd1d and the NKT TCR, which may provide a basis for the selective recognition of α-linked sugars. This study reinforces the notion that glycolipid-antigen recognition is achieved through molecular mechanisms different from those for peptide-antigen recognition and cautions against casting the Cd1 lipid-presenting proteins as MHC-like. N. A. Borg, K. S. Wun, L. Kjer-Nielsen, M. C. J. Wilce, D. G. Pellicci, R. Koh, G. S. Besra, M. Bharadwaj, D. I. Godfrey, J. McCluskey, J. Rossjohn, CD1d-lipid-antigen recognition by the semi-invariant NKT T-cell receptor. Nature 448 , 44-49 (2007). [PubMed] D. B. Moody, How a T cell sees sugar. Nature 448 , 36-37 (2007). [PubMed]

Glycolipids for Natural Killer T Cells

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Glycolipids for natural killer T cells

Natural killer T cells (NKT cells) play a central role in regulating immune responses influencing conditions ranging from autoimmune to infectious diseases. NKT cell responses are induced by recognition of glycolipid antigens presented by CD1d, an antigen presentation protein. In the last 10 years great strides have been made in understanding the types of glycolipids recognized by NKT cells. These advances have included determination of the lipid and carbohydrate recognition requirements for stimulation and identification of "natural" antigens for these cells.

Intrathymic NKT cell development is blocked by the presence of alpha-galactosylceramide.

NKT cell development takes place in the thymus, beginning when these cells branch away from CD4+CD8+ mainstream thymocytes upon expression of the Valpha14Jalpha18 T cell receptor (TCR) and recognition of the CD1d molecule. Although NKT cells express an invariant TCR alpha chain, the diverse TCR beta expression leaves open the possibility that the development of these cells is shaped by glycolipid antigen recognition in the context of CD1d. Here, we show that the presence of an agonist glycolipid ligand, alpha-galactosylceramide, while NKT cells are developing in vitro or in vivo, specifically ablates their development. In contrast, the delayed introduction of this compound in vitro or in vivo, after NKT cells have developed, does not deplete these cells. These data indicate that NKT cells pass through a developmental window where they are susceptible to TCR-mediated negative selection, and suggest that NKT cells with a potentially high level of self reactivity can be removed from the NKT cell repertoire before they exit the thymus.

Structural requirements for glycolipid antigen recognition by CD1b-restricted T cells.

The human CD1b protein presents lipid antigens to T cells, but the molecular mechanism is unknown. Identification of mycobacterial glucose monomycolate (GMM) as a CD1b-presented glycolipid allowed determination of the structural requirements for its recognition by T cells. Presentation of GMM to CD1b-restricted T cells was not affected by substantial variations in its lipid tails, but was extremely sensitive to chemical alterations in its carbohydrate or other polar substituents. These findings support the view that the recently demonstrated hydrophobic CD1 groove binds the acyl chains of lipid antigens relatively nonspecifically, thereby positioning the hydrophilic components for highly specific interactions with T cell antigen receptors.