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alphabeta T cell receptors (TCRs) are genetically restricted to corecognize peptide antigens bound to self-major histocompatibility complex (pMHC) molecules; however, the basis for this MHC specificity remains unclear. Despite the current dogma, evaluation of the TCR-pMHC-I structural database shows that the nongermline-encoded complementarity-determining region (CDR)-3 loops often contact the MHC-I, and the germline-encoded CDR1 and -2 loops frequently participate in peptide-mediated interactions. Nevertheless, different TCRs adopt a roughly conserved docking mode over the pMHC-I, in which three MHC-I residues (65, 69, and 155) are invariably contacted by the TCR in one way or another. Nonetheless, the impact of mutations at these three positions, either individually or together, was not uniformly detrimental to TCR recognition of pHLA-B*0801 or pHLA-B*3508. Moreover, when TCR-pMHC-I recognition was impaired, this could be partially restored by expression of the CD8 coreceptor. The structure of a TCR-pMHC-I complex in which these three (65, 69, and 155) MHC-I positions were all mutated resulted in shifting of the TCR footprint relative to the cognate complex and formation of compensatory interactions. Collectively, our findings reveal the inherent adaptability of the TCR in maintaining peptide recognition while accommodating changes to the central docking site on the pMHC-I.

The heterodimeric natural killer cells antigen CD94 (CD94)–NKG2‐A/NKG2‐B type II integral membrane protein (NKG2A) receptor family expressed on human and mouse natural killer (NK) cells monitors global major histocompatibility complex (MHC) class I cell surface expression levels through binding to MHC class Ia‐derived leader sequence peptides presented by HLA class I histocompatibility antigen, alpha chain E (HLA‐E; in humans) or H‐2 class I histocompatibility antigen, D‐37 (Qa‐1b; in mice). Although the molecular basis underpinning human CD94–NKG2A recognition of HLA‐E is known, the equivalent interaction in the murine setting is not. By determining the high‐resolution crystal structure of murine CD94–NKG2A in complex with Qa‐1b presenting the Qa‐1 determinant modifier peptide (QDM), we resolved the mode of binding. Compared to the human homologue, the murine CD94–NKG2A–Qa‐1b–QDM displayed alterations in the distribution of interactions across CD94 and NKG2A subunits that coincide with differences in electrostatic complementarity of the ternary complex and the lack of cross‐species reactivity. Nevertheless, we show that Qa‐1b could be modified through W65R + N73I mutations to mimic HLA‐E, facilitating binding with both human and murine CD94–NKG2A. These data underscore human and murine CD94–NKG2A cross‐species heterogeneity and provide a foundation for humanising Qa‐1b in immune system models. [ABSTRACT FROM AUTHOR]

Manipulation of the MHC-I presentation pathway, and thus limiting MHC-I cell surface expression, is used by many viruses to evade immune recognition. In particular, downregulation of MHC-I molecules at the cell surface can reduce the ability of CD8+ T cells to recognize viral peptides presented by MHC-I molecules and thereby delay viral clearance by CD8+ T cells. To date, MHC-I downregulation by influenza viruses has not been reported. Given that influenza virus infections are a global health concern and that CD8+ T cells play an important role in promoting influenza virus clearance and recovery from influenza disease, we investigated whether influenza A and B viruses (IAV, IBV) downregulated MHC-I as a novel mechanism to evade cellular immunity. Here, we showed that infection of several cell types, including epithelial A549 cells, with a panel of IAV and IBV viruses downregulated the surface MHC-I expression on IAV/IBV-infected cells during the late stages of influenza virus infection in vitro. This observation was consistent across a panel of class I-reduced (C1R) cell lines expressing 14 different HLA-A or -B alleles and a panel of 721.221 cell lines expressing 11 HLA-C alleles. Interestingly, IBV infection caused more pronounced reduction in surface MHC-I expression compared to IAV. Importantly, the two viruses utilized two distinct mechanisms for MHC-I downregulation. Our data demonstrated that while IAV caused a global loss of MHC-I within influenza-infected cells, IBV infection resulted in the preferential loss of MHC-I molecules from the cell surface, consequent of delayed MHC-I trafficking to the cell surface, resulting from retaining MHC-I intracellularly during IBV infection. Overall, our study suggests that influenza viruses across both IAV and IBV subtypes have the potential to downregulate MHC-I surface expression levels. Our findings provide new insights into the host-pathogen interaction of influenza A and B viruses and inform the design of novel vaccine strategies against influenza viruses. [ABSTRACT FROM AUTHOR]

The surveillance of target cells by natural killer ( NK) cells utilizes an ensemble of inhibitory and activating receptors, many of which interact with major histocompatibility complex ( MHC) class I molecules. NK cell recognition of MHC class I proteins is important developmentally for the acquisition of full NK cell effector capacity and during target cell recognition, where the engagement of inhibitory receptors and MHC class I molecules attenuates NK cell activation. Human NK cells have evolved two broad strategies for recognition of human leukocyte antigen ( HLA) class I molecules: (i) direct recognition of polymorphic classical HLA class I proteins by diverse receptor families such as the killer cell immunoglobulin-like receptors ( KIRs), and (ii) indirect recognition of conserved sets of HLA class I-derived peptides displayed on the non-classical HLA-E for recognition by CD94- NKG2 receptors. In this review, we assess the structural basis for the interaction between these NK receptors and their HLA class I ligands and, using the suite of published KIR and CD94- NKG2 ternary complexes, highlight the features that allow NK cells to orchestrate the recognition of a range of different HLA class I proteins. [ABSTRACT FROM AUTHOR]

The activation of murine and human natural killer (NK) cells is regulated by families of receptors including the Ly49 and Killer immunoglobulin-like receptors, respectively, both of which contain activating and inhibitory members. The archetypal role of inhibitory Ly49 receptors is to attenuate NK cell responses to normal cells that express major histocompatibility complex (MHC) class-I molecules, in essence allowing for more robust responses to infected or cancerous cells that lack MHC-I on their cell surface. However, it is now evident that Ly49 receptors have an appreciably more sophisticated array of functions. In particular, some activating Ly49 receptors can bind directly to MHC-I-like viral gene products such as m157, whereas others recognize self-MHC-I but only in the presence of viral chaperones. Although Ly49 receptor recognition is centred on the MHC-I-like fold, these NK cell receptors can also engage related ligands in unexpected ways. Herein we review the varied strategies employed by Ly49 receptors to recognize both self and viral ligands, with particular emphasis on the recently determined mode of Ly49-m157 ligation, and highlight the versatile nature of this family in the control of viral infections. [ABSTRACT FROM AUTHOR]

Murine cerebral malaria is a complex disease caused by Plasmodium berghei ANKA infection. Several cell types, including CD8+ T cells, are essential effectors of disease. Although the use of transgenic parasites expressing model antigens has revealed the induction of cytotoxic T lymphocytes (CTL) specific for these model antigens, there is no direct evidence for a response to authentic blood-stage parasite antigens, nor any knowledge of its magnitude. Our studies show that there is a dramatic primary parasite-specific CTL response, akin to viral immunity, reaching approximately 30% of splenic CD8+ T cells, with many producing interferon-&ggr; and tumor necrosis factor-&agr;. These cells express granzyme B and other markers of specific responders, are cytolytic, and respond to a broad array of major histocompatibility complex (MHC) I-restricted epitopes, 5 of which are identified here. Our studies indicate that vigorous CTL responses can be induced to pathogens even when they largely reside in red blood cells, which lack MHC I processing machinery. [ABSTRACT FROM AUTHOR]

αβ T cell receptors (TCRs) are genetically restricted to corecognize peptide antigens bound to self-major histocompatibility complex (pMHC) molecules; however, the basis for this MHC specificity remains unclear. Despite the current dogma, evaluation of the TCR-pMHC-l structural database shows that the nongermline-encoded complementaritydetermining region (CDR)-3 loops often contact the MHC-l, and the germline-encoded CDR1 and -2 loops frequently participate in peptide-mediated interactions. Nevertheless, different TCR5 adopt a roughly conserved docking mode over the pMHC-l, in which three MHC-I residues (65, 69, and 155) are invariably contacted by the TCR in one way or another. Nonetheless, the impact of mutations at these three positions, either individually or together, was not uniformly detrimental to TCR recognition of pHLA.B*0801 or pHLAB*35O8. Moreover, when TCR-pMHC-I recognition was impaired, this could be partially restored by expression of the CD8 coreceptor. The structure of a TCR-pMHC-I complex in which these three (65,69, and 155) MHC-l positions were all mutated resulted in shifting of the TCR footprint relative to the cognate complex and formation of compensatory interactions. Collectively, our findings reveal the inherent adaptability of the TCR in maintaining peptide recognition while accommodating changes to the central docking site on the pMHC-I. [ABSTRACT FROM AUTHOR]

Pigtail macaques ( Macaca nemestrina) are an increasingly common primate model for the study of human AIDS. Major Histocompatibility complex (MHC) class I-restricted CD8+ T cell responses are a critical part of the adaptive immune response to HIV-1 in humans and simian immunodeficiency virus (SIV) in macaques; however, MHC class I alleles have not yet been comprehensively characterized in pigtail macaques. The frequencies of ten previously defined alleles (four Mane-A and six Mane-B) were investigated in detail in 109 pigtail macaques using reference strand-mediated conformational analysis (RSCA). The macaques were derived from three separate breeding colonies in the USA, Indonesia and Australia, and allele frequencies were analysed within and between these groups. Mane-A*10, an allele that restricts the immunodominant SIV Gag epitope KP9, was the most common allele, present in 32.1% of the animals overall, with similar frequencies across the three cohorts. Additionally, RSCA identified a new allele ( Mane-A*17) common to three Indonesian pigtail macaques responding to the same Gag CD8+ T cell epitope. This broad characterization of common MHC class I alleles in more than 100 pigtail macaques further develops this animal model for the study of virus-specific CD8+ T cell responses. [ABSTRACT FROM AUTHOR]

In contrast to antigen-specific immunity orchestrated by major histocompatibility complex (MHC) class Ia molecules, the ancestrally related nonclassical MHC class Ib molecules generally mediate innate immune responses. Here we have demonstrated the structural basis by which the MHC class Ib molecule HLA-E mediates an adaptive MHC-restricted cytotoxic T lymphocyte response to human cytomegalovirus. Highly constrained by host genetics, the response showed notable fine specificity for position 8 of the viral peptide, which is the sole discriminator of self versus nonself. Despite the evolutionary divergence of MHC class Ia and class Ib molecules, the structure of the T cell receptor–MHC class Ib complex was very similar to that of conventional T cell receptor–MHC class Ia complexes. These results emphasize the evolutionary 'ambiguity' of HLA-E, which not only interacts with innate immune receptors but also has the functional capacity to mediate virus-specific cytotoxic T lymphocyte responses during adaptive immunity. [ABSTRACT FROM AUTHOR]

The pigtail macaque ( Macaca nemestrina) is a common model for the study of AIDS. The pigtail major histocompatibility complex class I allele Mane-A* 10 restricts an immunodominant simian immunodeficiency virus (SIV) Gag epitope (KP9) which rapidly mutates to escape T cell recognition following acute simian/human immunodeficiency virus infection. Two technologies for the detection of Mane-A* 10 in outbred pigtail macaques were developed: reference strand-mediated conformational analysis and sequence-specific primer polymerase chain reaction. A Mane-A*10/KP9 tetramer was then developed to quantify CD8+ T lymphocytes primed by multigenic DNA vaccination, which have previously been difficult to detect using standard interferon- γ-based T cell assays. We also demonstrate mutational escape at KP9 following acute SIV infection. Mane-A* 10+ animals have lower set point SIV levels than Mane-A* 10− animals, suggesting a significant fitness cost of escape. These studies pave the way for a more robust understanding of HIV vaccines in pigtail macaques. [ABSTRACT FROM AUTHOR]

HLA-G is a nonclassical major histocompatibility complex class I (MHC-l) molecule that is primarily expressed at the fetal-maternal interface, where it is thought to play a role in protecting the fetus from the maternal immune response. HLA-G binds a limited repertoire of peptides and interacts with the inhibitory leukocyte Ig-like receptors LIR-1 and LIR-2 and possibly with certain natural killer cell receptors. To gain further insights into HLA-G function, we determined the is-A structure of a monomeric HLA-G complexed to a natural endogenous peptide ligand from histone H2A (RIIPRHLQL). An extensive network of contacts between the peptide and the antigen-binding cleft reveal a constrained mode of binding reminiscent of the nonclassical HLA-E molecule, thereby providing a structural basis for the limited peptide repertoire of HLA-G. The α3 domain of HLA-G, a candidate binding site for the LIR-1 and -2 inhibitory receptors, is structurally distinct from the α3 domains of classical MHC-l molecules, providing a rationale for the observed affinity differences for these ligands. The structural data suggest a head-to-tail mode of dimerization, mediated by an intermolecular disulfide bond, that is consistent with the observation of HLA-G dimers on the cell surface. [ABSTRACT FROM AUTHOR]

CD94/NKG2 is a recently described receptor present on natural killer (NK) cells and certain T cells that is composed of the CD94 chain covalently associated with a member of the NKG2 family of molecules. Both chains are glycosylated members of the C-type lectin superfamily. The CD94/NKG2 receptors are functionally heterogenous depending on which NKG2 family member is associated with CD94. Initially, it was thought that CD94/NKG2 receptors recognized a broad array of HLA-A, -B and -C (classical), as well as the nonclassical HLA-G, MHC class I molecules. Instead, recent data have suggested that this receptor is specific for HLA-E complexed with a peptide derived from the signal sequence (residues 3–11) of certain classical MHC class I molecules. Position 2 (residue 4) in the signal sequence derived peptides appears pivotal in determining whether the HLA-E/peptide complex confers resistance to NK-mediated lysis. The potential roles that the CD94/NKG2-HLA-E receptor ligand interaction might play in infection and tumor development are discussed. [ABSTRACT FROM AUTHOR]

Human leukocyte antigen (HLA)-C*06:02 is identified as the allele associated with the highest risk for the development of the autoimmune skin disease psoriasis. However, the diversity and mode of peptide presentation by the HLA-C*06:02 molecule remains unclear. Here, we describe the endogenous peptide repertoire of ~3,000 sequences for HLA-C*06:02 that defines the peptide-binding motif for this HLA allomorph. We found that HLA-C*06:02 predominantly presents nonamer peptides with dominant arginine anchors at the P2 and P7 positions and a preference for small hydrophobic residues at the C terminus (PΩ). To determine the structural basis of this selectivity, we determined crystal structures of HLA-C*06:02 in complex with two self-peptides (ARTELYRSL and ARFNDLRFV) and an analogue of a melanocyte autoantigen (ADAMTSL5, VRSRR-abu-LRL) implicated in psoriasis. These structures revealed that HLA-C*06:02 possesses a deep peptide-binding groove comprising two electronegative Band E-pockets that coincide with the preference for P2 and P7 arginine anchors. The ADAMTSL5 autoantigen possessed a P7-Leu instead of the P7-Arg residue, but nevertheless was accommodated within the HLA-C*06:02 antigen-binding cleft. Collectively, our results provide the structural basis for understanding peptide repertoire selection in HLA-C*06:02. [ABSTRACT FROM AUTHOR]

Murine natural killer (NK) cells are regulated by the interaction of Ly49 receptors with major histocompatibility complex class I molecules (MHC-I). Although the ligands for inhibitory Ly49 were considered to be restricted to classical MHC (MHCIa), we have shown that the non-classical MHC molecule (MHCIb) H2-M3 was a ligand for the inhibitory Ly49A. Here we establish that another MHC-Ib, H2-Q10, is a bona fide ligand for the inhibitory Ly49C receptor. H2-Q10 bound to Ly49C with a marginally lower affinity (~5 μM) than that observed between Ly49C and MHC-Ia (H-2Kb/H-2Dd, both ~1 μM), and this recognition could be prevented by cis interactions with H-2K in situ. To understand the molecular details underpinning Ly49· MHC-Ib recognition, we determined the crystal structures of H2-Q10 and Ly49C bound H2-Q10. Unliganded H2-Q10 adopted a classical MHC-I fold and possessed a peptide-binding groove that exhibited features similar to those found in MHC-Ia, explaining the diverse peptide binding repertoire of H2-Q10. Ly49C bound to H2-Q10 underneath the peptide binding platform to a region that encompassed residues from theα1,α2, and α3 domains, as well as the associated β2-microglobulin subunit. This docking mode was conserved with that previously observed for Ly49C·H-2Kb. Indeed, structure-guided mutation of Ly49C indicated that Ly49C·H2-Q10 and Ly49C·H-2Kb possess similar energetic footprints focused around residues located within the Ly49C β4-stand and L5 loop, which contact the underside of the peptide-binding platform floor. Our data provide a structural basis for Ly49·MHC-Ib recognition and demonstrate that MHC-Ib represent an extended family of ligands for Ly49 molecules. [ABSTRACT FROM AUTHOR]

The killer cell Ig-like receptors (KIRs) expressed on the surface of NK cells recognize specific MHC class I (MHC-I) molecules and regulate NK cell activities against pathogen-infected cells and neoplasia. In HIV infection, survival is linked to host KIR and MHC-I genotypes. In the SIV macaque model, however, the role of NK cells is unclear due to the lack of information on KIR-MHC interactions. In this study, we describe, to our knowledge, the first in-depth characterization of KIR-MHC inter-actions in pigtailed macaques (Macaca nemestrina). Initially, we identified three distinct subsets of macaque NK cells that stained ex vivo with macaque MHC-I tetramers loaded with SIV peptides. We then cloned cDNAs corresponding to 15 distinct KIR3D alleles. One of these, KIR049-4, was an inhibitory KIR3DL that bound MHC-I tetramers and prevented activation, degranulation, and cytokine production by macaque NK cells after engagement with specific MHC-I molecules on the surface of target cells. Furthermore, KIR049-4 recognized a broad range of MHC-I molecules carrying not only the Bw4 motif, but also Bw6 and non-Bw4/Bw6 motifs. This degenerate, yet peptide-dependent, MHC reactivity differs markedly from the fine specificity of human KIRs. [ABSTRACT FROM AUTHOR]

We recently identified a protective MHC class lb-restricted CD8 T cell response to infection with mouse Polyomavirus. These CD8 T cells recognize a peptide from aa 139-147 of the VP2 viral capsid protein bound to the nonpolymorphic H-2Q9 molecule, a member of the Qa-2 family of &bgr;2m-associated MHC class Ib molecules. Q9:VP2.139-specific CD8 T cells exhibit an unusual inflationary response characterized by a gradual expansion over 3 mo followed by a stable maintenance phase. We previously demonstrated that Q9:VP2.139-specific CD8 T cells are dependent on Ag for expansion, but not for long-term maintenance. In this study, we tested the hypothesis that the expansion and maintenance components of the Q9:VP2.139-specific T cell response are differentially dependent on CD4 T cell help and CD28 costimulation. Depletion of CD4+ cells and CD28/CD40L blockade impaired expansion of Q9:VP2.139-specific CD8 T cells, and intrinsic CD28 signaling was sufficient for expansion. In contrast, CD4 T cell insufficiency, but not CD28/CD40L blockade, resulted in a decline in frequency of Q9:VP2.139-specific CD8 T cells during the maintenance phase. These results indicate that the Q9:VP2.139-specific CD8 T cell response to mouse Polyomavirus infection depends on CD4 T cell help and CD28 costimulation for inflationary expansion, but only on CD4 T cell help for maintenance. [ABSTRACT FROM AUTHOR]

While the prime function of classical MHC class I molecules (MHC-I) is to present peptide antigens to pathogen-specific cytotoxic T cells, non-classical MHC-I antigens perform a diverse array of functions in both innate and adaptive immunity. In this review we summarize recent evidence that non classical MHC-I molecules are not only recognized by pathogen-specific T cells but that they also serve as immunoregulatory molecules by stimulating a number of distinct non-conventional T cell subsets. [Copyright &y& Elsevier]

Abstract: The highly polymorphic major histocompatibility complex class Ia (MHC-Ia) molecules present a broad array of peptides to the clonotypically diverse αβ T-cell receptors. In contrast, MHC-Ib molecules exhibit limited polymorphism and bind a more restricted peptide repertoire, in keeping with their major role in innate immunity. Nevertheless, some MHC-Ib molecules do play a role in adaptive immunity. While human leukocyte antigen E (HLA-E), the MHC-Ib molecule, binds a very restricted repertoire of peptides, the peptide binding preferences of HLA-G, the class Ib molecule, are less stringent, although the basis by which HLA-G can bind various peptides is unclear. To investigate how HLA-G can accommodate different peptides, we compared the structure of HLA-G bound to three naturally abundant self-peptides (RIIPRHLQL, KGPPAALTL and KLPQAFYIL) and their thermal stabilities. The conformation of HLA-GKGPPAALTL was very similar to that of the HLA-GRIIPRHLQL structure. However, the structure of HLA-GKLPQAFYIL not only differed in the conformation of the bound peptide but also caused a small shift in the α2 helix of HLA-G. Furthermore, the relative stability of HLA-G was observed to be dependent on the nature of the bound peptide. These peptide-dependent effects on the substructure of the monomorphic HLA-G are likely to impact on its recognition by receptors of both innate and adaptive immune systems. [Copyright &y& Elsevier]

Abstract: Human leukocyte antigen (HLA)-E is a non-classical major histocompatibility complex class I molecule that binds peptides derived from the leader sequences of other HLA class I molecules. Natural killer cell recognition of these HLA-E molecules, via the CD94–NKG2 natural killer family, represents a central innate mechanism for monitoring major histocompatibility complex expression levels within a cell. The leader sequence-derived peptides bound to HLA-E exhibit very limited polymorphism, yet subtle differences affect the recognition of HLA-E by the CD94–NKG2 receptors. To better understand the basis for this peptide-specific recognition, we determined the structure of HLA-E in complex with two leader peptides, namely, HLA-Cw*07 (VMAPRALLL), which is poorly recognised by CD94–NKG2 receptors, and HLA-G*01 (VMAPRTLFL), a high-affinity ligand of CD94–NKG2 receptors. A comparison of these structures, both of which were determined to 2.5-Å resolution, revealed that allotypic variations in the bound leader sequences do not result in conformational changes in the HLA-E heavy chain, although subtle changes in the conformation of the peptide within the binding groove of HLA-E were evident. Accordingly, our data indicate that the CD94–NKG2 receptors interact with HLA-E in a manner that maximises the ability of the receptors to discriminate between subtle changes in both the sequence and conformation of peptides bound to HLA-E. [Copyright &y& Elsevier]

The highly polymorphic MHC class Ia molecules have a central role in adaptive immunity. By contrast, the closely related MHC class Ib molecules, which show limited polymorphism, are best known for regulating innate immune responses. Nevertheless, a recent area of interest is the emerging role of class Ib molecules in adaptive immunity, particularly in response to tumours and pathogens such as Mycobacteria, Listeria and Salmonella. Here, we review recent findings in this area, highlighting the structure of a T-cell receptor complexed with a cytomegalovirus peptide bound to the class Ib molecule, HLA-E. Collectively, these findings have implications for immunity, transplantation and autoimmunity, and our understanding of the evolution and plasticity of the molecular interactions mediating adaptive immunity. [Copyright &y& Elsevier]

Successful human immunodeficiency virus (HIV) vaccines will need to induce effective T-cell immunity. We studied immunodominant simian immunodeficiency virus (SIV) Gag-specific T-cell responses and their restricting major histocompatibility complex (MHC) class I alleles in pigtail macaques (Macaca nemestrina), an increasingly common primate model for the study of HIV infection of humans. CD8+ T-cell responses to an SIV epitope, Gag164-172KP9, were present in at least 15 of 36 outbred pigtail macaques. The immunodominant KP9-specific response accounted for the majority (mean, 63%) of the SIV Gag response. Sequencing from six macaques identified 7 new Mane-A and 13 new Mane-B MHC class I alleles. One new allele, Mane-A*10, was common to four macaques that responded to the KP9 epitope. We adapted reference strand-mediated conformational analysis (RSCA) to MHC class I genotype M. nemestrina. Mane-A*10 was detected in macaques presenting KP9 studied by RSCA but was absent from non-KP9-presenting macaques. Expressed on class I-deficient cells, Mane-A*10, but not other pigtail macaque MHC class I molecules, efficiently presented KP9 to responder T cells, confirming that Mane-A*10 restricts the KP9 epitope. Importantly, naïve pigtail macaques infected with SIVmac251 that respond to KP9 had significantly reduced plasma SIV viral levels (log10 0.87 copies/ml; P = 0.025) compared to those of macaques not responding to KP9. The identification of this common M. nemestrina MHC class I allele restricting a functionally important immunodominant SIV Gag epitope establishes a basis for studying CD8+ T-cell responses against AIDS in an important, widely available nonhuman primate species. [ABSTRACT FROM AUTHOR]