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The ability of the immune system to eliminate and shape the immunogenicity of tumours defines the process of cancer immunoediting.sup.1. Immunotherapies such as those that target immune checkpoint molecules can be used to augment immune-mediated elimination of tumours and have resulted in durable responses in patients with cancer that did not respond to previous treatments. However, only a subset of patients benefit from immunotherapy and more knowledge about what is required for successful treatment is needed.sup.2-4. Although the role of tumour neoantigen-specific CD8.sup.+ T cells in tumour rejection is well established.sup.5-9, the roles of other subsets of T cells have received less attention. Here we show that spontaneous and immunotherapy-induced anti-tumour responses require the activity of both tumour-antigen-specific CD8.sup.+ and CD4.sup.+ T cells, even in tumours that do not express major histocompatibility complex (MHC) class II molecules. In addition, the expression of MHC class II-restricted antigens by tumour cells is required at the site of successful rejection, indicating that activation of CD4.sup.+ T cells must also occur in the tumour microenvironment. These findings suggest that MHC class II-restricted neoantigens have a key function in the anti-tumour response that is nonoverlapping with that of MHC class I-restricted neoantigens and therefore needs to be considered when identifying patients who will most benefit from immunotherapy. In a mouse tumour model, immunotherapy-induced rejection of tumour cells requires presentation of both MHC class I and MHC class II antigens, which activate CD4.sup.+ and CD8.sup.+ T cells, respectively., Author(s): Elise Alspach [sup.1] [sup.2] , Danielle M. Lussier [sup.1] [sup.2] , Alexander P. Miceli [sup.1] [sup.2] , Ilya Kizhvatov [sup.1] , Michel DuPage [sup.3] [sup.8] , Adrienne M. Luoma [...]

Author(s): Andreas Blees [1]; Dovile Januliene [2]; Tommy Hofmann [3]; Nicole Koller [1]; Carla Schmidt [3]; Simon Trowitzsch (corresponding author) [1]; Arne Moeller (corresponding author) [2]; Robert Tamp (corresponding author) [...]

The bottlenose dolphin (Tursiops truncatus) is the most common cetacean species worldwide and the only marine mammal species resident in the Croatian part of the Adriatic Sea. To gain insight into genetic diversity of bottlenose dolphins at adaptively important loci relevant to conservation, we analysed the polymorphism of major histocompatibility complex (MHC) genes, which play a key role in pathogen confrontation and clearance. Specifically, we examined the diversity of MHC class II DRA, DQA and DQB alleles in 50 bottlenose dolphins from the Adriatic Sea collected between 1997 and 2011 and in 12 animals from other Mediterranean locations. Notable variation in DQA, DQB and three-locus haplotypes was found, with all 10 DQA and 12 DQB alleles encoding unique protein products. Analysis of the ratio of non-synonymous to synonymous substitution rates suggests that positive selection acts at both highly variable loci. Phylogenetic analyses revealed trans-species polymorphism at the DQB locus, strongly indicating the influence of balancing selection in the long term. In fact, the balancing selection observed in bottlenose dolphins is higher than that reported for most other cetaceans and comparable to that seen in terrestrial mammals., Introduction The bottlenose dolphin (Tursiops truncatus) is the most abundant cetacean species; it inhabits primarily coastal and inshore regions of warm and temperate seas around the world (Jefferson et al. [...]

The formation of precise connections between retina and lateral geniculate nucleus (LGN) involves the activity- dependent elimination of some synapses, with strengthening and retention of others. Here we show that the major histocompatibility complex (MHC) class I molecule H2-[D.sup.b] is necessary and sufficient for synapse elimination in the retinogeniculate system. In mice lacking both H2-[K.sup.b] and H2-[D.sup.b] ([K.sup.b][D.sup.b-/-]), despite intact retinal activity and basal synaptic transmission, the developmentally regulated decrease in functional convergence of retinal ganglion cell synaptic inputs to LGN neurons fails and eye-specific layers do not form. Neuronal expression of just H2-[D.sup.b] in [K.sup.b][D.sup.b-/-] mice rescues both synapse elimination and eye-specific segregation despite a compromised immune system. When patterns of stimulation mimicking endogenous retinal waves are used to probe synaptic learning rules at retinogeniculate synapses, long-term potentiation (LTP) is intact but long-term depression (LTD) is impaired in [K.sup.b][D.sup.b-/-] mice. This change is due to an increase in [Ca.sup.2+]-permeable AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) receptors. Restoring H2-[D.sup.b] to [K.sup.b][D.sup.b-/-] neurons renders AMPA receptors [Ca.sup.2+] impermeable and rescues LTD. These observations reveal an MHC-class-I-mediated link between developmental synapse pruning and balanced synaptic learning rules enabling both LTD and LTP, and demonstrate a direct requirement for H2-[D.sup.b] in functional and structural synapse pruning in CNS neurons., Early in development, before photoreceptors function, retinal ganglion cells (RGCs) spontaneously generate correlated bursts of action potentials called 'retinal waves' (1-3). Postsynaptic LGN neurons in turn are driven to fire [...]

[alpha][beta] 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. MHC restriction | T cell recognition | structural immunology | immunogenetics doi/ 10.1073/pnas.1004926107

MHC class I molecules function to display peptides generated from cellular and pathogen gene products for immune surveillance by [CD8.sup.+] T cells. Cells typically express ~100,000 class I molecules, or ~1 per 30,000 cellular proteins. Given none protein, one peptide' representation, immunosurveillance would be heavily biased toward the most abundant cell proteins. Cells use several mechanisms to prevent this, including the predominant use of defective ribosomal products (DRiPs) to generate peptides from nascent proteins and, as we show here, compartmentalization of DRiP peptide generation to prevent competition from abundant cytosolic peptides. This provides an explanation for the exquisite ability of T cells to recognize peptides generated from otherwise undetected gene products. MHC | peptide | translation | virus doi/ 10.1073/pnas.0910997107

Residues within processed protein fragments bound to major histocompatibility complex class I (MHC-I) glycoproteins have been considered to function as a series of 'independent pegs' that either anchor the peptide (p) to the MHC-I and/or interact with the spectrum of [alpha][beta]-T-cell receptors (TCRs) specific for the pMHC-I epitope in question. Mining of the extensive pMHC-I structural database established that many selfand viral peptides show extensive and direct interresidue interactions, an unexpected finding that has led us to the idea of 'constrained' peptides. Mutational analysis of two constrained peptides (the HLA B44 restricted self-peptide (B44DP[alpha]-EEFGRAFSF) and an H2-[D.sup.b] restricted influenza peptide (DbPA, SSLENFRAYV) demonstrated that the conformation of the prominently exposed arginine in both peptides was governed by interactions with MHC-I-orientated flanking residues from the peptide itself. Using reverse genetics in a murine influenza model, we revealed that mutation of an MHC-I-orientated residue (SSLENFRAYVSSLENARAYV) within the constrained PA peptide resulted in a diminished cytotoxic T lymphocyte (CTL) response and the recruitment of a limited pMHC-I specific TCR repertoire. Interactions between individual peptide positions can thus impose fine control on the conformation of pMHC-I epitopes, whereas the perturbation of such constraints can lead to a previously unappreciated mechanism of viral escape. crystal structure | cytotoxic T cell | MHC class I | viral immunity | epitope www.pnas.org/cgi/doi/10.1073/pnas.1000032107

Pathogen selection is postulated to drive MHC allelic diversity at loci for antigen presentation. However, readily apparent MHC infectious disease associations are rare in most species. The strong link between MHC-B haplotype and the occurrence of virally induced tumors in the chicken provides a means for defining the relationship between pathogen selection and MHC polymorphism. Here, we verified a significant difference in resistance to gallid herpesvirus-2 (GaHV-2)-induced lymphomas (Marek's disease) conferred by two closely-related recombinant MHC-B haplotypes. We mapped the crossover breakpoints that distinguish these haplotypes to the highly polymorphic BG1 locus. BG1 encodes an Ig-superfamily type I transmembrane receptor-like protein that contains an immunoreceptor tyrosine-based inhibition motif (ITIM), which undergoes phosphorylation and is recognized by Src homology 2 domain-containing protein tyrosine phosphatase (SHP-2). The recombinant haplotypes are identical, except for differences within the BG1 3'-untranslated region (3'-UTR). The 3'-UTR of the BG1 allele associated with increased lymphoma contains a 225-bp insert of retroviral origin and showed greater inhibition of luciferase reporter gene translation compared to the other allele. These findings suggest that BG1 could affect the outcome of GaHV-2 infection through modulation of the lymphoid cell responsiveness to infection, a condition that is critical for GaHV-2 replication and in which the MHC-B haplotype has been previously implicated. This work provides a mechanism by which MHC-B region genetics contributes to the incidence of GaHV-2-induced malignant lymphoma in the chicken and invites consideration of the possibility that similar mechanisms might affect the incidence of lymphomas associated with other oncogenic viral infections. disease resistance gene | GaHV-2 induced lymphoma | Gallus gallus | Marek's disease

Major histocompatibility complex (MHC) class I molecules present peptides, produced through cytosolic proteasomal degradation of cellular proteins, to cytotoxic T lymphocytes. In dendritic cells, the peptides can also be derived from internalized antigens through a process known as cross-presentation. The cellular compartments involved in cross-presentation remain poorly defined. We found a role for peptide trimming by insulin-regulated aminopeptidase (IRAP) in cross-presentation. In human dendritic cells, IRAP was localized to a [Rab14.sup.+] endosomal storage compartment in which it interacted with MHC class I molecules. IRAP deficiency compromised cross-presentation in vitro and in vivo but did not affect endogenous presentation. We propose the existence of two pathways for proteasome-dependent cross-presentation in which final peptide trimming involves IRAP in endosomes and involves the related aminopeptidases in the endoplasmic reticulum.

Multiple sclerosis (MS), a common central nervous system inflammatory disease, has a major heritable component. Susceptibility is associated with the MHC class II region, especially HLA-DRB5*0101-HLA-DRB1*1501-HLA-DQA1*0102-HLA-DQB1*0602 haplotypes (hereafter DR2), which dominate genetic contribution to MS risk. Marked linkage disequilibrium (LD) among these loci makes identification of a specific locus difficult. The once-leading candidate, HLA-DRB1*15, localizes to risk, neutral, and protective haplotypes. HLA-DRB1*15 and HLA-DQB1*0602, nearly always Iocated together on a small ancestral chromosome segment, are strongly MS-associated. One intervening allele on this haplotype, viz. HLA-DQA1*0102, shows no primary MS association. Two Canadian cohorts (n = 830 and n = 438 trios) genotyped for HLA-DRB1, HLA-DQA1 and HLA-DQB1 alleles were tested for association using TDT. To evaluate epistasis involving HLA-DRB1*15, transmissions from HLA-DRB1*15-negative parents were stratified by the presence/absence of HLA-DRB1*15 in affected offspring. All 3 alleles contribute to MS susceptibility through novel epistatic interactions. HLA-DQA 1*0102 increased disease risk when combined with HLA-DRB1*1501 in trans, thereby unambiguously implicating HLA-DQ in MS susceptibility. Three-locus haplotypes demonstrated that HLA-DRB1*1501 and HLA-DQB1*0602 each influence risk. Transmissions of rare morcellated DR2 haplotypes showed no interaction with HLA-DQA1*0102. Incomplete haplotypes bearing only HLA-DRB1*1501 or HLA-DQB1*0602 d id not predispose to MS. Balanced reciprocal transmission distortion can mask epistatic allelic association. These findings implicate epistasis among HLA class II alleles in human immune responses generally, provide partial explanation for intense linkage disequilibrium in the MHC, have relevance to animal models, and demonstrate key roles for DR2-specific interactions in MS susceptibility. MHC disease associations may be more generally haplotypic or diplotypic. genetics | MHC | linkage disequilibrium

T cell recognition of minor histocompatibility antigens (mHags) underlies allogeneic immune responses that mediate graft-versus-host disease and the graft-versus-leukemia effect following stem cell transplantation. Many mHags derive from single amino acid polymorphisms in MHC-restricted epitopes, but our understanding of the molecular mechanisms governing mHag immunogenicity and recognition is incomplete. Here we examined antigenic presentation and T-cell recognition of HA-1, a prototypic autosomal mHag derived from single nucleotide dimorphism (HA-[1.sup.H] versus HA-[1.sup.R]) in the HMHA1 gene. The HA-[1.sup.H] peptide is restricted by HLA-A2 and is immunogenic in HA-[1.sup.R/R] into HA-[1.sup.H] transplants, while HA-[1.sup.R] has been suggested to be a 'null allele' in terms of T cell reactivity. We found that proteasomal cleavage and TAP transport of the 2 peptides is similar and that both variants can bind to MHC. However, the His>Arg change substantially decreases the stability and affinity of HLA-A2 association, consistent with the reduced immunogenicity of the HA-[1.sup.R] variant. To understand these findings, we determined the structure of an HLA-A2-HA-[1.sup.H] complex to 1.3[Angstrom] resolution. Whereas His-3 is accommodated comfortably in the D pocket, incorporation of the lengthy Arg-3 is predicted to require local conformational changes. Moreover, a soluble TCR generated from HA-[1.sup.H]-specific T-cells bound HA-[1.sup.H] peptide with moderate affinity but failed to bind HA-[1.sup.R], indicating complete discrimination of HA-1 variants at the level of TCR/MHC interaction. Our results define the molecular mechanisms governing immunogenicity of HA-1, and highlight how single amino acid polymorphisms in mHags can critically affect both MHC association and TCR recognition. graft-versus-leukemia | stem cell transplantation | major histocompatibility complex

Many viruses avoid immune surveillance during latent infection through reduction in the synthesis of virally encoded proteins. Although antigen presentation critically depends on the level of viral protein synthesis, the precise mechanism used to regulate the generation of antigenic peptide precursors remains elusive. Here, we demonstrate that a purine overloaded virally encoded mRNA lacking secondary structure significantly impacts the efficiency of protein translation and prevents endogenous antigen presentation. Reducing this purine bias through the generation of constructs expressing codon-modified sequences, while maintaining the encoded protein sequence, increased the stem-loop structure of the corresponding mRNA and dramatically enhanced self-synthesis of the viral protein. As a consequence, a higher number of HLA-peptide complexes were detected on the surface of cells expressing this viral protein. Furthermore, these cells were more efficiently recognized by virus-specific T cells compared with those expressing the same antigen expressed by a purine-biased mRNA. These findings delineate a mechanism by which viruses regulate self-synthesis of proteins and offer an effective strategy to evade [CD8.sup.+] T cell-mediated immune regulation. antigen processing | EBV-encoded nuclear antigen 1 | immune evasion | protein synthesis

This study investigated the dynamic regulation of IIx-IIb MHC genes in the fast white medial gastrocnemius (WMG) muscle in response to intermittent resistance exercise training (RE), a model associated with a rapid shift from IIb to IIx expression (11). We investigated the effect of 4 days of RE on the transcriptional activity across the skeletal MHC gene locus in the WMG in female Sprague-Dawley rats. Our results show that RE resulted in significant shifts from lib to IIx observed at both the pre-mRNA and mRNA levels. An antisense RNA (xII NAT) was detected in the intergenic (IG) region between IIx and IIb, extending across the entire IIx gene and into its promoter. The expression of the xII NAT was positively correlated with IIb pre-mRNA (R = +0.8), and negatively correlated with IIx pre-mRNA (R = -0.8). Transcription mapping of the IIx-IIb IG region revealed the generation of sense IIb and xII NATs from a single promoter region. This bidirectional promoter is highly conserved among species and contains several regulatory elements that may be implicated in its regulation. These results suggest that the IIx and the lib genes are physically and functionally linked via the bidirectional promoter. In order for the IIx MHC gene to be regulated, a feedback mechanism from the IG xII NAT is needed. In conclusion, the IG bidirectional promoter generating antisense RNA appears to be essential for the coordinated regulation of the skeletal muscle MHC genes during dynamic pheno-type shifts. gene transcription; pre-mRNA; natural antisense RNA; comparative genomics

Naturally occurring [Foxp3.sup.+][CD4.sup.+][CD25.sup.+] T cells (nTregs) isolated from lungs of naive mice regulate allergic airway hyperresponsiveness (AHR) and inflammation. Here, we demonstrate the critical requirement for engagement of MHC class I on [CD4.sup.+][CD25.sup.+] T cells by CD8 for the functional activation of these nTregs. Suppression of allergen-induced AHR and inflammation by nTregs was abolished in mice treated with anti-CD8. Correspondingly, decreased levels of IL-10 and TGF-[beta] and increased levels of Th2 cytokines in bronchoalveolar lavage were detected in these treated mice. Similarly, nTregs isolated from [beta]2[m.sup.-/-] mice or from mice treated with anti-MHC I antibody in vitro before intratracheal transfer failed to modulate AHR or inflammation. Coculture of nTregs with [CD8.sup.+] T cells increased IL-10 and TGF-[beta]. Addition of anti-MHC I or anti-CD8 reduced IL-10 and TGF-[beta]. These results demonstrate that functional activation of nTregs requires the interaction between MHC I on CD4+[CD25.sup.+] T cells and CD8. IL-10 | TGF-[beta] | airway reactivity

Natural killer (NK) cells are known to reject MHC-mismatched targets within blood organs, yet their role in peripheral lymphoid tissue remains unresolved. Here we address the capacity of NK cells to migrate within lymph nodes (LN) using two-photon microscopy to characterize cell velocities and interaction dynamics within the native lymphoid-tissue environment. Adoptively transferred unmanipulated NK cells were highly motile (6-7 [micro]m/min) and capable of forming transient contacts with both syngeneic and allogeneic B cells. Stable conjugate interactions (lasting >5 min) formed preferentially with allogeneic cells, resulting in diminished motility and subsequent elimination of the target cell. In marked contrast to unmanipulated cells, NK cells purified by CD49b-positive selection exhibited only limited motility (2-3 [micro]m/min). This velocity impairment arose largely because CD49b cross-linking enhanced NK cell adhesion to collagen fibers within the node. Moreover, CD49b cross-linking prevented NK cells from reconstituting effector cytolytic function in vivo, inhibited target cell lysis in vitro, and augmented IFN-[gamma] responses to IL-2 activation in vitro. Taken together our data demonstrate that NK cells are a functionally important component of the LN microenvironment, and that cell motility and effector function are strongly modulated via CD49b manipulation. motility | two-photon microscopy

This article delineates the cellular basis for MHC-1 downregulation of HIV-1 immunoevasion. Results demonstrate that three Nef sorting motifs cooperate, in an orderly fashion, to bring about Nef downregulation of MHC-1 and occur by P13K-mediated ARF6 endosomal pathway.

Particular major histocompatibility complex (MHC) class II alleles clearly contribute to T cell-mediated autoimmune type 1 diabetes (T1D) in both humans and nonobese diabetic (NOD) mice. However, studies in NOD mice indicate MHC class I-restricted T cell responses are also essential to T1D development. In humans, epidemiological studies have suggested that some common class I alleles, including HLA-A2.1 (A*02011), may confer increased susceptibility to T1D when expressed in conjunction with certain class II alleles. We show here that when HLA-A2.1 molecules are transgenically expressed in NOD mice, A2-restricted T cell responses arise against pancreatic [beta] cells, leading to an earlier onset of T1D. The accelerated onset of T1D in the NOD.HLA-A2.1 transgenic mice is not due to nonspecific effects of expressing a third class I molecule, because a stock of NOD mice transgenically expressing HLA-B27 class I molecules showed no such acceleration of T1D, but rather were significantly protected from disease. These findings provide the first functional evidence that certain human MHC class I molecules can contribute to the development of T1D.

T cell receptor (TCR)-mediated activation of [CD4.sup.+] T cells is known to require multivalent engagement of the TCR by, for example, oligomeric peptide--MHC complexes. In contrast, for [CD8.sup.+] T cells, there is evidence for TCR-mediated activation by univalent engagement of the TCR. We have here compared oligomeric and monomeric [L.sup.d] and [K.sup.b] peptide--MHC complexes and free peptide as stimulators of [CD8.sup.+] T cells expressing the 2C TCR. We found that the monomers are indeed effective in activating naive and effector [CD8.sup.+] T cells, but through an unexpected mechanism that involves transfer of peptide from soluble monomers to T cell endogenous MHC ([K.sup.b]) molecules. The result is that T cells, acting as antigen-presenting cells, are able to activate other naive T cells. [CD8.sup.+] T lymphocytes | T cell receptor | major histocompatibility complex | antigen presentation | lymphocyte activation

Some members of the major histocompatibility complex (MHC) class I gene family are encoded outside the MHC. Here we describe a family of mouse class I-like genes mapping to the vicinity of the leukocyte receptor complex (LRC) on chromosome 7. This family, which we call Mill (MHC class I-like located near the LRC), has two members designated Mill1 and Mill2. Both genes are predicted to encode membrane glycoproteins with domain organization essentially similar to that of MHC class I heavy chains. The following features of Mill are noteworthy. (i) The deduced MILL proteins lack most of the residues known to be involved in the docking of peptides in classical MHC class I molecules. (ii) Among the known members of the class I gene family, MILL1 and MILL2 are related most closely to MICA/MICB encoded in the human MHC. (iii) Unlike all other known members of the class I gene family, Mill1 and Mill2 have an exon between those coding for the signal peptide and the [alpha]1 domain. (iv) Mill1 has a more restricted expression profile than Mill2. (v) The gene orthologous to Mill1 or Mill2 apparently is absent in the human. (vi) Mill1 and Mill2 show a limited degree of polymorphism in laboratory mice. The observation that the Mill family is related most closely to the MIC family, together with its apparent absence in the human, suggests its involvement in innate immunity.

Human embryonic stem (ES) cells are pluripotent cells that may be used in transplantation medicine. These cells can be induced to differentiate into cells from the three embryonic germ layers both in vivo and in vitro. To determine whether human ES cells might be rejected after transplantation, we examined cell surface expression of the MHC proteins in these cells. Our results show very low expression levels of MHC class I (MHC-I) proteins on the surface of human ES cells that moderately increase on in vitro or in vivo differentiation. A dramatic induction of MHC-I proteins was observed when the cells were treated with IFN-[gamma] but not with IFN-[alpha] or -[beta]. However, all three IFNs induced expression of MHC-I proteins in differentiated human ES cells. MHC-II proteins and HLA-G were not expressed on the surface of undifferentiated or differentiated cells. Ligands for natural killer cell receptors were either absent or expressed in very low levels in human ES cells and in their differentiated derivatives. In accordance, natural killer cytotoxic assays demonstrated only limited lysis of both undifferentiated and differentiated cells. To initiate a histocompatibility databank of human ES cells, we have isotyped several of the published ES cell lines for their human leukocyte antigens. In conclusion, our results demonstrate that human ES cells can express high levels of MHC-I proteins and thus may be rejected on transplantation.

We examine here the nature of the differential recognition by CD[4.sup.+] T cells of a single peptide from hen-egg white lysozyme (HEL) presented by I-[A.sup.k] class II MHC molecules. Two subsets of T cells (called A and B) interact with the same peptide, each in unique ways that reflect the nature of the complex of peptide and MHC. We show that the A and B set of T cells can be distinguished by their functional interaction with the three T cell receptor (TCR) contact residues of the bound peptide. The dominant peptide of HEL selected from processing is bound in a single register where a critical TCR contact residue is situated about the middle of the core segment of the peptide: all T cells establish functional contact with it. Three sets of T cells, however, can be distinguished by their differential recognition of two TCR contacts situated at the amino and carboxyl sides of the central TCR contact residue. Type A T cells, the conventional cells that see the peptide after processing of HEL, need to recognize all three TCR contact residues. In contrast, the type B T cells recognize the peptide given exogenously, but not when processed: these T cells recognize either one of the peripheral TCR contact residues, indicating a much more flexible interaction of peptide with I-[A.sup.k] molecules. We discuss the mode of generation of the various T cells and their biological relevance. histocompatibility molecules | antigen presentation | T cell receptors | antigen-presenting cells

We have solved the crystal structure of the MHCII molecule, [IA.sup.b], containing an antigenic variant of the major [IA.sup.b]-binding peptide derived from the MHCII IE[alpha] chain. The four MHC pockets at p1, p4, p6, and p9 that usually bind peptide side chains are largely empty because of alanines in the peptide at these positions. The complex is nevertheless very stable, apparently because of unique alternate interactions between the [IA.sup.b] and peptide. In particular, there are multiple additional hydrogen bonds between the N-terminal end of the peptide and the [IA.sup.b] [alpha] chain and an extensive hydrogen bond network involving an asparagine at p7 position of the peptide and the [IA.sup.b] [beta] chain. By using knowledge of the shape and size of the traditional side chain binding pockets and the additional possible interactions, an [IA.sup.b] peptide-binding motif can be deduced that agrees well with the sequences of known [IA.sup.b]-binding peptides.

Cytotoxic T lymphocytes (CTLs) with a C[D8.sup.+] phenotype have the potential to recognize and attack major histocompatibility complex (MHC) class I-expressing brain cells. Most brain cells, including neurons, can be stimulated to present peptides to C[D8.sup.+] CTLs by MHC class I molecules, and are susceptible to CTL-mediated cytotoxicity in culture. In disease-affected brain parenchyma, C[D8.sup.+] CTLs outnumber other T-cell subtypes. They show clonal expansion in several inflammatory and degenerative CNS diseases, such as multiple sclerosis (MS), virus-induced inflammatory brain diseases and paraneoplastic neurological disorders. In MS, damage of axons is closely linked to the C[D8.sup.+] CTLs, and protection against CTL-mediated damage should be considered as a new therapeutic approach in MS and other neuroinflammatory diseases.

If T cells require specific interactions with MHC-bound peptides during positive selection, then the specificities of T cells selected by one peptide should be distinct from those selected by another. We have examined positive selection of CD4 T cells in four strains of mice, each overexpressing a different peptide--1-[A.sup.b]([A.sup.b]) complex. We show that a subset of CD4 T cells is selected by the overexpressed peptide and that the specificities of the CD4 T cells, as measured by reactivity to wild-type antigen-presenting cells, vary greatly depending on which peptide is overexpressed. These differences in specificity are mediated through positive selection not negative selection. Each of the four peptide--[A.sup.b] complexes appears to adopt a different conformation, and these differences correlate with the differences in reactivity. Our results suggest that individual peptide--MHC complexes positively select different subsets of self-MHC-reactive T cells and that the conformation of the peptide--MHC complex may contribute to this process. thymus | CD4 T cells | development | T cell specificity | mice

We have analyzed the patterns of positive and negative selection of thymocytes expressing the T cell antigen receptor (TCR) from the D10.G4.1 T cell clone. This TCR confers reactivity to several non-self MHC class II alleles with a remarkably broad range of avidities. Therefore, negative selection can be studied when induced by high-, intermediate-, or low-avidity interactions with endogenous peptide-MHC complexes, all within the same TCR transgenic system. These data directly demonstrate that MHC class II-peptide ligands that fail to activate mature T cells can promote negative selection of immature thymocytes. Additionally, we show that negative selection of thymocytes can occur at two distinct 'time points' during development depending on the avidity of the TCR for the MHC-peptide complex. Finally, we show that the self-peptide repertoire plays a significant role in selection because alteration of the self-peptide repertoire by disruption of the H2-Ma gene drastically alters selection of D10 TCR-expressing thymocytes.

Adult naive T cells, which are at rest in normal conditions, proliferate strongly when transferred to lymphopenic hosts. In neonates, the first mature thymocytes to migrate to the periphery reach a compartment devoid of preexisting T cells. We have extensively analyzed the proliferation rate and phenotype of peripheral T cells from normal C57BL/6 and T cell antigen receptor transgenic mice as a function of age. We show that, like adult naive T cells transferred to lymphopenic mice, neonatal naive T cells proliferate strongly. By using bone-marrow transfer and thymic-graft models, we demonstrate that the proliferation of the first thymic emigrants reaching the periphery requires T cell antigen receptor-self-peptide/self-MHC interactions and is regulated by the size of the peripheral T cell pool.

Genes of the major histocompatibility complex (MHC) play a central role in immune recognition, yet they also influence the odor of individuals. Mice can be trained to distinguish odors mediated by classical MHC loci; however, training can introduce confounding behavioral artifacts. This study demonstrates that mice can distinguish some, but not all, naturally occurring allelic variants at classical MHC loci without prior training. This result suggests that MHC-disassortative mating preferences might operate by means of small MHC-based odor differences, and could therefore contribute to diversifying selection acting on MHC loci. Here we show that odors of two MHC mutant mouse strains (bm1 and bm3) can be distinguished, even after genetic background is controlled by intercrossing strains. These two strains differ by five amino acids, three of which are predicted to chemically contact peptides bound to the peptide-binding region (PBR), the site of antigen presentation for T cell recognition. However, the odors of neither bm1 nor bm3 were distinguished from their parental B6 haplotype after randomizing genomic background, despite discrimination of purebred B6 and bm1 strain odors. These combined results suggest that (i) there may be an MHC odor discrimination threshold based on divergence in PBR residues, providing a more logical pattern of MHC-based odor discrimination than found in previous training studies, where discrimination ability was not correlated with PBR divergence; and (ii) additional (non-MHC) mutations that influence odor have accumulated in these strains during the 100 generations of divergence between pure B6 and bm1 strains.

Research demonstrates binding of murine and human beta2-microglobulin to the major histocompatibility complex class I heavy chain alpha3 domain as revealed by surface plasmon resonance technique. Data show that both beta2-microglobulins exhibit similar binding kinetics, temperature dependence, and beta2-microglobulin refolding.

Crystal structures of alphabeta T cell receptors, bound to MHC I and II molecules along with bound antigenic peptides, demonstrate the atomic contacts, which illustrate MHC restricted T cell recognition sites.

Immunoglobulin G binding properties of the human major histocompatibility complex-related Fc receptor and its crystal structure were determined. The neonatal Fc receptor (FcRn) has two related functions separate from each other. It transports maternal immunoglobulin G to pre- or neonatal mammals and thus gives passive immunity. It also protects IgG from normal serum protein catabolism. Current efforts to use FcRn-mediated regulation of IgG half-life in serum o increase lifetimes of antibody-based therapeutics are related to this work. Binding of human FcRn to the four human IgG subclasses shows subclass and allotype variations, but no clear subclass affinity differences that are related to serum half-lives.

The evolution of immunology as a scientific discipline is reviewed with discussion of the future of the field. The manipulation of the adaptive immune response to overcome many human diseases may be used to advantage in the future. Drugs cannot exploit the specificity of adaptive immunity. The study of the immune system has led to discovery of principles with broad implications for diverse biological processes, some of which are discussed. The field is ripe for taking basic research to clinical applications. Topics include the basis for specificity and memory in adaptive immune responses, initiation of immune responses from the perspective of lymphocytes, innate immunity, effector mechanisms.

The transport of major histocompatibility complex (MHC) class II molecules are hypothesized to play a role in developmental regulation. This was tested in mouse dendritic cells (DCs) as they are the most potent of all antigen presenting cells. Investigations show that the transport of MHC class II molecules are controlled by DCs through the cleavage of the third polypeptide invariant (Ii) chain. Cathepsin S is the enzyme thought to be responsible for cleaving the Ii chain. This in turn is controlled by the down-regulation of the cysteine protease inhibitor cystatin C.

The protein structure of major histocompatibility complex (MHC) class I antigens contains a range of information suitable for achieving a number of cellular functions common to epithelial cells. Among these functions are cellular transport, lymphocyte response regulation to altered epithelium and foreign antigens. The MHC class I antigens can link specific ligand binding to a host of pathways in the cell that ultimately invoke lymphocyte response.

HLA-DM is a major histocompatibility complex (MHC) class II-like molecule that facilitates antigen processing by catalyzing the exchange of invariant chain-derived peptides (CLIP) from class II molecules for antigenic peptides. HLA-DO is a second class II-like molecule that physically associates with HLA-DM in B cells. HLA-DO was shown to block HLA-DM function. Purified HLA-DM-DO complexes could not promote peptide exchange in vitro. Expression of HLA-DO in a class II+ and DM+, DO- human T cell line caused the accumulation of class II-CLIP complexes, indicating that HLA-DO blocked DM function in vivo and suggesting that HLA-DO is an important modulator of class II-restricted antigen processing., Mhc class II molecules assemble in the endoplasmic reticulum (ER) as a nonameric complex consisting of an invariant chain trimer associated with three class II [Alpha]Β dimers(1). Signals in the [...]

Bacterial superantigens (SAgs) bind to major histocompatibility complex (MHC) class II molecules and activate T cells in a V[Beta]-restricted fashion. We recently identified subsets of HLA-DR1 molecules that show selectivity for SAgs. Here, we extend these observations by showing that different cell lineages demonstrate distinct SAg-binding specificities although they all express HLA-DR1. Indeed, B cells bind staphylococcal enterotoxin A (SEA) and toxic shock syndrome toxin 1 (TSST-1) with high affinity while staphylococcal enterotoxin B (SEB) binding is barely detectable. In contrast, DR1-transfected HeLa cells show efficient binding of SEB, but not of SEA or TSST-1. We investigated the class II maturation events required for efficient interaction with SAgs and found that the ability of cells to bind and present the toxins can be drastically modulated by coexpression of the class II-associated invariant chain (Ii) and HLA-DM. SEA binding to DR1 molecules required coexpression of Ii, whereas TSST-1 binding was selectively enhanced by DM. Binding of SEB was affected by cell type-specific factors other than Ii or DM. The selectivity of SAgs for different MHC class II populations was minimally affected by HLA-DR intrinsic polymorphism and could not be explained by binding to alternative sites on DR molecules. Our results indicate that SAgs are sensitive to structural heterogeneity in class II molecules, which is consequent to the differential regulation of expression of antigen processing cofactors. Therefore, we speculate that Staphylococcus aureus have retained the ability to express numerous SAgs in adaptation to the micro-heterogeneity displayed by MHC class II molecules and that this may relate to their ability to infect different tissues.

Myasthenia gravis (MG) is a T cell-regulated, antibody-mediated autoimmune disease. Two pep-tides representing sequences of the human acetylcholine receptor [Alpha]-subunit, p195-212 and p259-271, were previously shown to stimulate peripheral blood lymphocytes of patients with MG and were found to be immunodominant T cell epitopes in SJL and BALB/c mice, respectively. Single amino acid substituted analogs of p195-212 (analog Ala-207) and p259-271 (analog Lys-262) were synthesized. We showed that analogs Ala-207 and Lys-262 inhibited, in vitro and in vivo, the proliferative responses of T cell lines specific to the relevant peptide and lymph node cells of mice immunized to p195-212 and p259-271, respectively. To inhibit T cell responses to both peptides (p195-212 and p259-271), we synthesized dual analogs composed of the tandemly arranged two single (Ala-207 and Lys-262) analogs (dual analog) either sequentially (Ala-207-Lys-262) or reciprocally (Lys-262-Ala-207). In the present study, we report that both dual analogs could bind to major histocompatibility complex class II molecules on antigen-presenting cells of SJL and BALB/c mice. Analog Lys-262-Ala-207, which bound more efficiently to major histo-compatibility complex class II molecules, was found to inhibit the proliferative responses of both p195-212- and p259-271-specific T cell lines. Furthermore, the analog inhibited the in vivo priming of lymph node cells of both SJL and BALB/c mice when administered i.v., i.p., or per os. The dual analog Lys-262-Ala-207 could also immunomodulate myasthenogenic manifestations in mice with experimental autoimmune MG induced by inoculation of a pathogenic T cell line. Thus, a single peptide that is composed of analogs to two epitope specificities can be used to regulate T cell responses and disease associated with each epitope.

Mice deficient in major histocompatibility complex (MHC) molecules I and II were analyzed to determine T cell specificities in the absence of MHC expression. Analysis of the MHC reactivity of germline T cell receptor (TCR) sequences indicated the presence of inherent MHC reactivity in TCRs due to the specific interaction of the receptors with MHC. Coreceptor signaling induced by anti-CD4 antibody promoted positive selection in a manner that was similar to MHC-mediated positive selection in vivo. Furthermore, the preselection repertoire exhibited high reactivity with MHC-expressing cells.