Major Histocompatibility Complex Specificity Research Articles

In silico prediction of B and T cell epitopes of infectious salmon anemia virus proteins and molecular modeling of T cell epitopes to salmon major histocompatibility complex (MHC) class I

Infectious salmon anemia (ISA) can be devastating in farmed Atlantic salmon (Salmo salar). The disease can evolve into epidemics if it is not contained and controlled. ISA epidemics were seen in Norway in the early 1990s and Chile in 2007–2009.Consequently, there is an urgent need to develop a vaccine to prevent or treat the infection. In this study, an immunoinformatic approach was employed to predict 32 lineal B-cell epitopes based on antigenicity and surface accessibility prediction for ISAV fusion (F), hemagglutinin-esterase (HE), and matrix (M) proteins. On the other hand, twelve conformational B-cell epitopes were also predicted.We further identified six antigenic cytotoxic T lymphocyte (CTL) epitopes and investigated the binding interactions with five salmon MHC-I proteins after docking the peptides to the binding groove of the MHC-I proteins. Our results showed that all the predicted epitopes could bind to salmon MHC-I with high negative ΔG values with medium to high binding affinities. Hence, the predicted epitopes have a high potential of being recognized by Atlantic salmon MHC-I to elicit a CD8+ T cell response in salmon. The predicted and analyzed B and T cell antigenic epitopes in this work might present an initial set of peptides for future vaccine development against ISAV.The ability to model and predict these interactions will ultimately lead to the ability to predict potential binding for MHCs and epitopes that were not studied previously. As current knowledge of salmon MHC specificity is limited, studying and modeling interactions in the peptide/MHC complex is a key to resolving unknown epitope specificity.

MhcVizPipe: A Quality Control Software for Rapid Assessment of Small- to Large-Scale Immunopeptidome Datasets

MS-based immunopeptidomics is maturing into an automatized and high-throughput technology, producing small- to large-scale datasets of clinically relevant major histocompatibility complex (MHC) class I-associated and class II-associated peptides. Consequently, the development of quality control (QC) and quality assurance systems capable of detecting sample and/or measurement issues is important for instrument operators and scientists in charge of downstream data interpretation. Here, we created MhcVizPipe (MVP), a semiautomated QC software tool that enables rapid and simultaneous assessment of multiple MHC class I and II immunopeptidomic datasets generated by MS, including datasets generated from large sample cohorts. In essence, MVP provides a rapid and consolidated view of sample quality, composition, and MHC specificity to greatly accelerate the “pass–fail” QC decision-making process toward data interpretation. MVP parallelizes the use of well-established immunopeptidomic algorithms (NetMHCpan, NetMHCIIpan, and GibbsCluster) and rapidly generates organized and easy-to-understand reports in HTML format. The reports are fully portable and can be viewed on any computer with a modern web browser. MVP is intuitive to use and will find utility in any specialized immunopeptidomic laboratory and proteomics core facility that provides immunopeptidomic services to the community.

Thymic Origins of T Cell Receptor Alloreactivity.

Major histocompatibility complex (MHC) restriction is a unique feature of T cell antigen recognition. Mature T cells respond to antigenic nonself peptides bound to self-MHC molecules, but a sizeable fraction of peripheral T cells can also respond to nonself peptide-MHC (pMHC) complexes in the context of transplantation. MHC specificity of the T cell receptor (TCR) repertoire is shaped during thymic development. Two hypotheses have been proposed to explain MHC specificity of T cells. It has been suggested that MHC specificity is an intrinsic feature of TCR structure, mediated by the germline-encoded regions of the TCR sequence. In support of this model, an estimated 15% to 30% of preselection TCR repertoire is estimated to be MHC-specific. Moreover, structural studies have shown some degree of conserved binding topology for TCR-peptide MHC complexes. However, there is also evidence that MHC restriction can be imposed on the TCR repertoire during thymic development, and it has been proposed that the interaction of the Lck kinase with CD4 or CD8 coreceptors is critical for generation of MHC specificity. This review will discuss recent work on assessment of the preselection of TCR repertoire, molecular evidence for the germline encoded TCR bias for MHC, and for the coreceptor sequestration model in the context of alloreactivity and transplantation.

How an alloreactive T-cell receptor achieves peptide and MHC specificity

T-cell receptor (TCR) allorecognition is often presumed to be relatively nonspecific, attributable to either a TCR focus on exposed major histocompatibility complex (MHC) polymorphisms or the degenerate recognition of allopeptides. However, paradoxically, alloreactivity can proceed with high peptide and MHC specificity. Although the underlying mechanisms remain unclear, the existence of highly specific alloreactive TCRs has led to their use as immunotherapeutics that can circumvent central tolerance and limit graft-versus-host disease. Here, we show how an alloreactive TCR achieves peptide and MHC specificity. The HCV1406 TCR was cloned from T cells that expanded when a hepatitis C virus (HCV)-infected HLA-A2- individual received an HLA-A2+ liver allograft. HCV1406 was subsequently shown to recognize the HCV nonstructural protein 3 (NS3):1406-1415 epitope with high specificity when presented by HLA-A2. We show that NS3/HLA-A2 recognition by the HCV1406 TCR is critically dependent on features unique to both the allo-MHC and the NS3 epitope. We also find cooperativity between structural mimicry and a crucial peptide "hot spot" and demonstrate its role, along with the MHC, in directing the specificity of allorecognition. Our results help explain the paradox of specificity in alloreactive TCRs and have implications for their use in immunotherapy and related efforts to manipulate TCR recognition, as well as alloreactivity in general.

Establishment of six homozygous MHC-B haplotype populations associated with susceptibility to Marek’s disease in Chinese specific pathogen-free BWEL chickens

The highly polymorphic chicken major histocompatibility complex (MHC) is associated with different levels of immunologic responses to certain avian pathogens. MHC-B haplotype chickens are an important genetic resource for studying the genetic determination of pathogen resistance and susceptibility. The BWEL chicken population is the only specific pathogen-free (SPF) chickens bred and developed by the State Center of Poultry Genetic Resources of Laboratory Animals in China. In this study, we successfully established six homozygous MHC-B haplotype populations from the BWEL chickens using microsatellite marker technology, named as BW/G(1, 2, 3, 5, 6, 7) lines, and their molecular genotypes were matched to six serologically defined MHC-B haplotypes, B13, B15, B2, B5, B21 and B19, respectively. The sequences of BF genes exons 2 and 3 from four successive generations (F1–F4) of the BW/G(n) lines were completely consistent with those of serologically defined MHC-B haplotypes. Subsequently, six BW/G(n) line specific allo-antisera were prepared by immunization with red blood cells (RBCs) and hemagglutination tests results showed the BW/G(n) SPF chickens could be serologically differentiated. Additionally, susceptibility to Marek’s disease (MD) in the BW/G3 (B2 haplotype) and BW/G7 (B19 haplotype) lines were determined by comparing mortality, macroscopic and histopathological lesions, and viral loads in feather pulp. The BW/G7 line showed greater genetic susceptibility to the very virulent MD virus (MDV) strain than the BW/G3 line. The establishment of MHC-B haplotype chicken populations associated with susceptibility to MD will be helpful for studying host immune responses and further developing the more effective vaccines in the context of MHC specificities, and they are also very useful for an understanding of MHC genes architecture and function.

Characterization of binding specificities of bovine leucocyte class I molecules: impacts for rational epitope discovery.

The binding of peptides to classical major histocompatibility complex (MHC) class I proteins is the single most selective step in antigen presentation. However, the peptide-binding specificity of cattle MHC (bovine leucocyte antigen, BoLA) class I (BoLA-I) molecules remains poorly characterized. Here, we demonstrate how a combination of high-throughput assays using positional scanning combinatorial peptide libraries, peptide dissociation, and peptide-binding affinity binding measurements can be combined with bioinformatics to effectively characterize the functionality of BoLA-I molecules. Using this strategy, we characterized eight BoLA-I molecules, and found the peptide specificity to resemble that of human MHC-I molecules with primary anchors most often at P2 and P9, and occasional auxiliary P1/P3/P5/P6 anchors. We analyzed nine reported CTL epitopes from Theileria parva, and in eight cases, stable and high affinity binding was confirmed. A set of peptides were tested for binding affinity to the eight BoLA proteins and used to refine the predictors of peptide-MHC binding NetMHC and NetMHCpan. The inclusion of BoLA-specific peptide-binding data led to a significant improvement in prediction accuracy for reported T. parva CTL epitopes. For reported CTL epitopes with weak or no predicted binding, these refined prediction methods suggested presence of nested minimal epitopes with high-predicted binding affinity. The enhanced affinity of the alternative peptides was in all cases confirmed experimentally. This study demonstrates how biochemical high-throughput assays combined with immunoinformatics can be used to characterize the peptide-binding motifs of BoLA-I molecules, boosting performance of MHC peptide-binding prediction methods, and empowering rational epitope discovery in cattle.

HLA molecules and nasal carriage of Staphylococcus aureus isolated from dialysis and kidney transplant patients at a hospital in Southern Brazil

BackgroundHealthy individuals can host Staphylococcus aureus in the nasopharynx, body surface and vagina. Most invasive infections by this bacterium are endogenous, caused by strains spread from the nasopharynx of carriers. S. aureus is a pathogen involved in the etiology of hospital- and community-acquired infections. Transplant and dialysis patients are at risk of colonization or infection by multi-resistant S. aureus. Infection is directly linked to individual immunity, and the major histocompatibility complex (MHC) plays a crucial role in determining susceptibility to diseases. Different MHC specificities have been shown to be more frequent in individuals suffering from certain diseases. This study aimed to investigate the association between HLA class I (HLA-A and -B) and class II (HLA-DRB1) molecules and nasal carriage of S. aureus in dialysis and kidney transplant patients at a hospital in Southern Brazil.ResultsThe sample consisted of 70 dialysis and 46 kidney transplant patients, totaling 116 patients. All subjects were typed for HLA molecules using LABType® SSO (One Lambda). Nasal swab samples of S. aureus were isolated from the nasal cavity (both nostrils) of patients undergoing dialysis or kidney transplantation.In renal dialysis patients, HLA-A*02 was the most frequent allele in both carriers (25.5%) and non-carriers (21.2%) of S. aureus. Allele A*68 was not observed in the carrier group, but the allele was observed six times in the non-carrier group (p = 0.0097). Regarding HLA-B and HLA-DRB1, no allele was shown to be involved in protection against or susceptibility to carriage of S. aureus. In kidney transplant patients, allele A*03 was more frequent in the non-carrier (20.83%) than in the carrier (5.88%) group (p = 0.0486). HLA-B*15 was present in carriers (5.88%) and non-carriers (25%) (p = 0.0179). Regarding class II alleles, DRB1*03 appeared to be related to susceptibility to carriage of S. aureus (p = 0.0319).ConclusionsOur findings suggest that HLA-DRB1*03 may be involved in susceptibility to nasal carriage of S. aureus in transplant patients. In addition, HLA-A*68 (dialysis patients) and HLA-A*03 and HLA-B*15 (transplant patients) appear to be associated with increased resistance to S. aureus nasal carriage.

αβ T Cell Receptors that Do Not Undergo Major Histocompatibility Complex-Specific Thymic Selection Possess Antibody-like Recognition Specificities

Major histocompatibility complex (MHC) restriction is the cardinal feature of T cell antigen recognition and is thought to be intrinsic to αβ T cell receptor (TCR) structure because of germline-encoded residues that impose MHC specificity. Here, we analyzed αβTCRs from T cells that had not undergone MHC-specific thymic selection. Instead of recognizing peptide-MHC complexes, the two αβTCRs studied here resembled antibodies in recognizing glycosylation-dependent conformational epitopes on a native self-protein, CD155, and they did so with high affinity independently of MHC molecules. Ligand recognition was via the αβTCR combining site and involved the identical germline-encoded residues that have been thought to uniquely impose MHC specificity, demonstrating that these residues do not only promote MHC binding. This study demonstrates that, without MHC-specific thymic selection, αβTCRs can resemble antibodies in recognizing conformational epitopes on MHC-independent ligands.

A Comprehensive Analysis of Shared Loci between Systemic Lupus Erythematosus (SLE) and Sixteen Autoimmune Diseases Reveals Limited Genetic Overlap

In spite of the well-known clustering of multiple autoimmune disorders in families, analyses of specific shared genes and polymorphisms between systemic lupus erythematosus (SLE) and other autoimmune diseases (ADs) have been limited. Therefore, we comprehensively tested autoimmune variants for association with SLE, aiming to identify pleiotropic genetic associations between these diseases. We compiled a list of 446 non–Major Histocompatibility Complex (MHC) variants identified in genome-wide association studies (GWAS) of populations of European ancestry across 17 ADs. We then tested these variants in our combined Caucasian SLE cohorts of 1,500 cases and 5,706 controls. We tested a subset of these polymorphisms in an independent Caucasian replication cohort of 2,085 SLE cases and 2,854 controls, allowing the computation of a meta-analysis between all cohorts. We have uncovered novel shared SLE loci that passed multiple comparisons adjustment, including the VTCN1 (rs12046117, P = 2.02×10−06) region. We observed that the loci shared among the most ADs include IL23R, OLIG3/TNFAIP3, and IL2RA. Given the lack of a universal autoimmune risk locus outside of the MHC and variable specificities for different diseases, our data suggests partial pleiotropy among ADs. Hierarchical clustering of ADs suggested that the most genetically related ADs appear to be type 1 diabetes with rheumatoid arthritis and Crohn's disease with ulcerative colitis. These findings support a relatively distinct genetic susceptibility for SLE. For many of the shared GWAS autoimmune loci, we found no evidence for association with SLE, including IL23R. Also, several established SLE loci are apparently not associated with other ADs, including the ITGAM-ITGAX and TNFSF4 regions. This study represents the most comprehensive evaluation of shared autoimmune loci to date, supports a relatively distinct non–MHC genetic susceptibility for SLE, provides further evidence for previously and newly identified shared genes in SLE, and highlights the value of studies of potentially pleiotropic genes in autoimmune diseases.

OETMAP: a new feature encoding scheme for MHC class I binding prediction

Deciphering the understanding of T cell epitopes is critical for vaccine development. As recognition of specific peptides bound to Major histocompatibility complex (MHC) class I molecules, cytotoxic T cells are activated. This is the major step to initiate of immune system response. Knowledge of the MHC specificity will enlighten the way of diagnosis, treatment of pathogens as well as peptide vaccine development. So far, a number of methods have been developed to predict MHC/peptide binding. In this article, a novel feature amino acid encoding scheme is proposed to predict MHC/peptide complexes. In the proposed method, we have combined orthonormal encoding (OE) and Taylor's Venn-diagram, and have used Linear support vector machines as the classifier in the tests. We also have compared our method to current feature encoding scheme techniques. The tests have been carried out on comparatively large Human leukocyte antigen (HLA)-A and HLA-B allele peptide three binding datasets extracted from the Immune epitope database and analysis resource. On three datasets experimented, the IC50 cutoff a criteria is used to select the binders and non-binders peptides. Experimental results show that our amino acid encoding scheme leads to better classification performance than other amino acid encoding schemes on a standalone classifier.

Targeting B cell leukemia with highly specific allogeneic T cells with a public recognition motif

The possibility that allogeneic T cells may be targeted to leukemia has important therapeutic implications. As most tumor antigens represent self-proteins, high-avidity tumor-specific T cells are largely deleted from the repertoire of the patient. In contrast, T cells from major histocompatibility complex (MHC)-mismatched donors provide naïve repertoires wherein such cells have not been systematically eliminated. Yet, evidence for peptide degeneracy or poly-specificity warrants caution in the use of foreign human leukocyte antigen (HLA) or peptide complexes as therapeutic targets. Here, we cocultured HLA-A(*)0201-negative T cells with autologous dendritic cells engineered to present HLA-A(*)0201 complexed with a peptide from the B cell antigen CD20 (CD20p). HLA-A(*)0201/CD20p pentamer-reactive CD8(+) T cells were readily obtained from all donors. The polyclonal cells showed exquisite peptide and MHC specificity, and efficiently killed HLA-A(*)0201-positive B cells, including primary chronic lymphocytic leukemia cells. The T cell receptor (TCR) sequences displayed a novel type of conservation, with extensive homology in the TCR β chain complementarity-determining region 3 and in J, but not V, region. This is surprising, as the donors were HLA disparate and their TCR repertoires are expected to show little overlap. The results demonstrate the first public recognition motif for an allogeneic HLA/peptide complex. The allo-restricted T cells or TCRs could provide graft-versus-leukemia in the absence of graft-versus-host disease.

Absence of Epstein-Barr virus-specific, HLA class II-restricted CD4+ cytotoxic T lymphocytes in infectious mononucleosis

Cytotoxic T lymphocytes (CTL) with the CD4+ phenotype that recognize major histocompatibility complex (MHC) class II antigens are detectable very frequently in cultures of human alloreactive or virus-specific T cells. The significance of these CD4+ CTL for an immune reaction in vivo is not clear. Since Epstein-Barr virus (EBV) transformed B cells express HLA-class I and class II antigens equally well both CD8+ and CD4+ CTL should be stimulated during an acute EBV infection. We analysed the MHC specificity and the phenotype of EBV-specific CTL from patients with infectious mononucleosis (IM). When tested directly without any previous culture, T cells from patients in the acute phase of IM showed specific MHC-restricted cytotoxicity against the autologous B cell line. Addition of a HLA class I specific monoclonal antibody (MoAb) but not of a HLA class II specific MoAb resulted in a complete blocking of the lytic activity. Cell sorting revealed that the entire cytotoxic activity was present in the CD8+ fraction whereas no specific CTL were detectable in the CD4+ fraction. The absence of cytotoxicity in CD4+ cells was not due to a lack of activation of these cells since both CD8+ and CD4+ cells were activated in situ, showing spontaneous growth in interleukin-2 (IL-2) and expressing the activation marker TP103. Frequency estimation revealed that 1/300-1/600 CD8+ but only 1/2000-1/4000 CD4+ T cells gave rise to a specific CTL colony after 10 days. If CD4+ colonies were tested repeatedly for cytotoxicity we found that CD4+ CTL acquired their cytotoxicity during in vitro culture. In addition, we isolated EBV-specific CD4+ T cell clones able to lyse their stimulator cells in the presence but not in the absence of lectin, even after a long period of culture. Taken together our results show that cytotoxicity mediated by CD4+ T cells does not play a role in an anti-viral immune response.

Deletion of CD4 and CD8 Coreceptors Permits Generation of αβT Cells that Recognize Antigens Independently of the MHC

The thymus generates major histocompatibility complex (MHC)-restricted alphabetaT cells that only recognize antigenic ligands in association with MHC or MHC-like molecules. We hypothesized that MHC specificity might be imposed on a broader alphabetaTCR repertoire during thymic selection by CD4 and CD8 coreceptors that bind and effectively sequester the tyrosine kinase Lck, thereby preventing T cell receptor (TCR) signaling by non-MHC ligands that do not engage either coreceptor. This hypothesis predicts that, in coreceptor-deficient mice, alphabeta thymocytes would be signaled by non-MHC ligands to differentiate into alphabetaT cells lacking MHC specificity. We now report that MHC-independent alphabetaT cells were indeed generated in mice deficient in both coreceptors as well as MHC ("quad-deficient" mice) and that such mice contained a diverse alphabetaT cell repertoire whose MHC independence was confirmed at the clonal level. We conclude that CD4 and CD8 coreceptors impose MHC specificity on a broader alphabetaTCR repertoire during thymic selection by preventing thymocytes from being signaled by non-MHC ligands.

NetMHCpan, a Method for Quantitative Predictions of Peptide Binding to Any HLA-A and -B Locus Protein of Known Sequence

BackgroundBinding of peptides to Major Histocompatibility Complex (MHC) molecules is the single most selective step in the recognition of pathogens by the cellular immune system. The human MHC class I system (HLA-I) is extremely polymorphic. The number of registered HLA-I molecules has now surpassed 1500. Characterizing the specificity of each separately would be a major undertaking.Principal FindingsHere, we have drawn on a large database of known peptide-HLA-I interactions to develop a bioinformatics method, which takes both peptide and HLA sequence information into account, and generates quantitative predictions of the affinity of any peptide-HLA-I interaction. Prospective experimental validation of peptides predicted to bind to previously untested HLA-I molecules, cross-validation, and retrospective prediction of known HIV immune epitopes and endogenous presented peptides, all successfully validate this method. We further demonstrate that the method can be applied to perform a clustering analysis of MHC specificities and suggest using this clustering to select particularly informative novel MHC molecules for future biochemical and functional analysis.ConclusionsEncompassing all HLA molecules, this high-throughput computational method lends itself to epitope searches that are not only genome- and pathogen-wide, but also HLA-wide. Thus, it offers a truly global analysis of immune responses supporting rational development of vaccines and immunotherapy. It also promises to provide new basic insights into HLA structure-function relationships. The method is available at http://www.cbs.dtu.dk/services/NetMHCpan.

Characterization of Genetic Polymorphism of Novel MHC B-LB II Alleles in Chinese Indigenous Chickens

Genetic polymorphism of the major histocompatibility complex (MHC) B-LB II gene was studied by amplification of exon 2 using PCR, followed by cloning and DNA sequencing in eight indigenous Chinese chicken populations. To reveal the genetic variation of the B-LB II gene, 37 types of patterns detected by PCR-SSCP were investigated first, which would be used to screen novel B-LB II sequences within the breeds. The types of PCR-SSCP patterns and final sequencing allowed for the identification of 31 novel MHC B-LB II alleles from 30 unrelated individuals of Chinese chickens that were sampled. These are the first designators for the alleles of chicken MHC B-LB II gene based on the rule of assignment for novel mammalian alleles. Sequence alignment of the 31 B-LB II alleles revealed a total of 68 variable sites in the fragment of exon 2, of which 51 parsimony informative and 17 singleton variable sites were observed. Among the polymorphic sites, the nucleotide substitutions in the first and second positions of the codons accounted for 36.76% and 35.29%, respectively. The sequence similarities between the alleles were estimated to be 90.6%–99.5%. The relative frequencies of synonymous and nonsynonymous nucleotide substitutions within the region were 2.92%±0.94% and 14.64%±2.67%, respectively. These results indicated that the genetic variation within exon 2 appeared to have largely arisen by gene recombination and balancing selection. Alignment of the deduced amino acid sequences of the β1 domain coded by exon 2 revealed 6 synonymous mutations and 27 nonsynonymous substitutions at the 33 disparate sites. In particular, the nonsynonymous substitutions at the putative peptide-binding sites are considered to be associated with immunological specificity of MHC B-LB II molecule in Chinese native chickens. These results can provide a molecular biological basis for the study of disease resistance in chicken breeding.

Antagonist peptide selects thymocytes expressing a class II major histocompatibility complex-restricted T cell receptor into the CD8 lineage.

CD4/CD8 lineage decision is an important event during T cell maturation in the thymus. CD8 T cell differentiation usually requires corecognition of major histocompatibility complex (MHC) class I by the T cell receptor (TCR) and CD8, whereas CD4 T cells differentiate as a consequence of MHC class II recognition by the TCR and CD4. The involvement of specific peptides in the selection of T cells expressing a particular TCR could be demonstrated so far for the CD8 lineage only. We used mice transgenic for an MHC class II-restricted TCR to investigate the role of antagonistic peptides in CD4 T cell differentiation. Interestingly, antagonists blocked the development of CD4(+) cells that normally differentiate in thymus organ culture from those mice, and they induced the generation of CD8(+) cells in thymus organ culture from mice impaired in CD4(+) cell development (invariant chain-deficient mice). These results are in line with recent observations that antagonistic signals direct differentiation into the CD8 lineage, regardless of MHC specificity.

Thymic skewing of the CD4/CD8 ratio maps with the T-cell receptor α-chain locus

The thymic preference for CD4+ T cells over CD8+ T cells is often attributed to a default pathway favouring CD4+ T cells or to homeostatic mechanisms. It is also clear, however, that T-cell receptor (TCR) preferences for major histocompatibility complex (MHC) class I versus class II binding will strongly influence an individual clone's skewing to the CD4 or CD8 subset. The variable region of each TCR alpha chain (V alpha) studied to date is found to be overrepresented in either CD4+ or CD8+ cells, suggesting that each V alpha element can interact more favourably with either MHC class I or class II molecules. Indeed, TCRs appear to have an intrinsic ability to interact with MHC molecules, and single amino acid residues present in germline-encoded complementarity determining region 1 (CDR1) and CDR2 of the V alpha element can be responsible for determining MHC specificity. Interestingly, the degree of CD4/CD8 skewing is variable among different mouse strains and in human populations. Here, we have shown that polymorphism in CD4/CD8 skewing between B6 and BALB/c mice is determined by the stem cell genotype and not by environmental effects, and that it maps in or near the TCR alpha-chain complex, Tcra. This was confirmed by comparing Tcra(b) with Tcra(a) or Tcra(c) haplotypes in congenic mice. We propose that the array of V alpha genes in various Tcra haplotypes exerts influence over the proportion of CD4 and CD8 subsets generated and may account in part for the observed thymic skewing. Thus, while it has been suggested that the TCR genes have been selected by evolution for MHC binding, our results further indicate selection for class II MHC preference.

Major histocompatibility complex class I-dependent skewing of the natural killer cell Ly49 receptor repertoire.

Subsets of mouse natural killer (NK) cells express receptors encoded by the Ly49 gene family that recognize allelic determinants on major histocompatibility complex (MHC) class I molecules. Recognition of self class I molecules typically inhibits NK cell lytic function. The presence of NK cell subsets expressing receptors which are able to discriminate class I alleles raises the possibility that there exist mechanisms to coordinate the NK cell receptor repertoire with the class I molecules of the host. In the present study, we determined the effects of class I gene expression on the frequencies of NK cells expressing three different Ly49 receptors defined by monoclonal antibodies. We show here an MHC-dependent skewing of NK cell subsets expressing multiple Ly49 receptors with specificity for self MHC. The results provide the first evidence that the frequencies of NK cells expressing different Ly49 receptors are determined by the host's MHC molecules. The results also extend previous findings that MHC class I expression influences the cell surface levels of each Ly49 receptor, suggesting an additional mechanism by which MHC molecules may influence the effective specificity of NK cells. Models to account for self tolerance and MHC-controlled repertoire differences are discussed.

Peptide binding specificity of major histocompatibility complex class I resolved into an array of apparently independent subspecificities: quantitation by peptide libraries and improved prediction of binding.

Considerable interest has focused on understanding how major histocompatibility complex (MHC) specificity is generated and characterizing the specificity of MHC molecules with the ultimate goal being to predict peptide binding. We have used a strategy where all possible peptides of a particular size are distributed into positional scanning combinatorial peptide libraries (PSCPL) to develop a highly efficient, universal and unbiased approach to address MHC specificity. The PSCPL approach appeared qualitatively and quantitatively superior to other currently used strategies. The average effect of any amino acid in each position was quantitated, allowing a detailed description of extended peptide binding motifs including primary and secondary anchor residues. It also identified disfavored residues which were found to be surprisingly important in shaping MHC class I specificity. Assuming that MHC class I specificity is the result of largely independently acting subsites, the binding of unknown peptides could be predicted. Conversely, this argues that MHC class I specificities consist of an array of subspecificities acting in a combinatorial mode.

Cloning of immunoglobulin-superfamily members associated with HLA-C and HLA-B recognition by human natural killer cells.

Cytotoxicity by natural killer (NK) cells is inhibited by major histocompatibility complex (MHC) class I molecules on target cells. This inhibition may be mediated by NK receptors with different MHC specificities. A family of four NK-specific complementary DNAs (cDNAs), designated NKATs (NK-associated transcripts), was identified that encoded related transmembrane proteins, characterized by an extracellular region with two or three immunoglobulin-superfamily domains and by a cytoplasmic domain with an unusual antigen receptor activation motif (ARAM). The distribution of these cDNAs was clonotypic and correlated with NK cell inhibition by particular class I alleles. Thus, NKAT cDNAs may encode receptors for class I molecules on NK cells.