A polymorphic major histocompatibility complex class II-like locus maps outside of both the chicken B-system and Rfp-Y-system.

Chickens have two major regions encoding major histocompatibility complex (MHC) class Ialpha genes and MHC class IIss genes, the serological and functional B-system and the Rfp-Y-system. Recently, they have been shown to assort in a genetically independent way although still located on the same microchromosome. Moreover, the monomorphic MHC class IIalpha gene maps at a third locus located 5 cM from the nearest class IIss genes, located in the B-system (Kaufman et al., 1995). A pedigree family was studied in three generations in order to assign MHC class IIss restriction fragments observed in Southern blot analyses to either the B-system, the Rfp-Y-system or the B-Lalpha locus. In this study, we demonstrate by classical genetic testing of chickens within this fully pedigreed family the existence of an MHC class II-like polymorphic restriction fragment that segregates independently of the B-system, the Rfp-Y-system and of the B-Lalpha locus.     

Polymorphisms of the equine major histocompatibility complex class II DRA locus

The full extent of the polymorphism of ELA-DRA in Equidae is not yet known. Given the apparent differences in DRA polymorphisms between Equidae and other species, the aims of this study were to more fully characterize ELA-DRA, determine the extent of gene polymorphism and establish the allele-frequency distribution. An allele reference panel for the second exon of ELA-DRA was established by sequence-based typing of 69 equine DNA samples consisting of various breeds of domestic horse (Equus caballus), together with donkeys (Equus asinus), Grant's zebras (Equus boehmi) and one onager (Equus hemionus). Five of the six previously reported alleles detected using single-strand conformation polymorphism were found: ELA-DRA*0101, ELA-DRA*0201, ELA-DRA*0301, ELA-DRA*0501 (Albright-Fraser DG et al. Polymorphism of DRA among equids. Immunogenetics 1996: 43: 315-7) and ELA-DRA*0601 (GenBank accession number AF5419361). In addition to the previously reported alleles, five novel ELA-DRA alleles were detected within the ELA-DRA allele reference panel. One of these was identified in E. caballus (ELA-DRA*JBH11), one in E. boehmi and E. hemionus (ELA-DRA*JBZ185) and three in E. asinus (ELA-DRA*JBD3, ELA-DRA*JBD17 and ELA-DRA*JBH45). A total of 565 equine DNA samples were screened using reference-strand-mediated conformation analysis, a double-stranded conformation-based mutation detection system that can be used to type existing ELA-DRA alleles and identify new variants. Based on our findings, at least 11 ELA-DRA alleles are now known to exist, and this level of polymorphism at the DRA locus appears to be unique to the genus Equus. Both the previously reported alleles and the new alleles displayed a species-specific distribution.     

Conformational isomers of a peptide–class II major histocompatibility complex

Summary: The relative plasticity of peptide binding to class II major histocompatibility complex (MHC) molecules permits formation of multiple conformational isomers by the same peptide and MHC molecule; such conformers are specifically recognized by distinct subsets of T cells. Here, we review current knowledge and recent advances in our understanding of peptide–class II MHC conformational isomerism and the mechanisms that generate distinct MHC–peptide conformers. We focus on our studies of two T‐cell subsets, type A and B, which recognize distinct conformers of the dominant epitope of hen egg white lysozyme presented by I‐A^k. These conformers form via different pathways and in distinct intracellular vesicles: the type A conformer forms in late endosomes upon processing of native protein, while the more flexible type B conformer forms in early endosomes and at the cell surface. In this process, H2‐DM acts as a conformational editor, eliminating the type B conformer in late endosomes. Type B T cells constitute a significant component of the naïve T‐cell repertoire; furthermore, self‐reactive type B T cells escape negative selection and are present in abundance in the periphery. Ongoing studies should elucidate the role of type B T cells in immunity to pathogens and in autoimmune pathology.     

A single nomenclature and associated database for alleles at the major histocompatibility complex class II DRB1 locus of sheep

The development of standardised nomenclatures with associated databases containing reference sequences for alleles at polymorphic loci within the major histocompatibility complex (MHC) has been facilitated by the development of the immuno polymorphism database (IPD). Recently, included within IPD-MHC is information on allelic diversity within sheep species (IPD-MHC-OLA). Here, we present the first report of progress in populating the sheep IPD-MHC database with alleles at the class II MHC DRB1 locus. The sequence of 63 Ovar-DRB1 alleles within 24 allelic families is now held within the database, each meeting the minimum requirement of a complete second exon. These sequences are derived from a combination of genomic and cDNA-based approaches and represent the most extensive collection of validated alleles at the sheep DRB1 locus yet described. Although these 63 alleles probably represent only a fraction of the DRB1 allelic diversity in sheep species worldwide, we encourage the research community to use the official allelic nomenclature and to contribute allelic sequences to the database via its web-based submission tool. In time, the IPD-MHC-OLA resource will underpin population-based MHC genotyping studies and help to simplify meta-analyses of multi-source data from wild and domestic sheep populations.     

A first map of the porcine major histocompatibility complex class I region

Abstract: A map of the SLA complex, or swine major histocompatibility complex (MHC), class I region was constructed by alignment of yeast artificial chromosomes (YACs) harboring MHC class I genes as well as anchor genes already mapped within the human MHC complex (HLA). Five YACs containing 9 anchor genes built a contig of about 1.0–1.2 Mb between the SLA class HI BAT1 locus and the olfactory receptor-like genes OLF42. Ten different SLA class I sequences, including putative allelic forms of published classical and non-classical SLA class I genes, were assigned to the 400-kb enclosing centromeric part of the contig. Three additional YACs comprising the OLF89 genes and two YACs containing the butyrophilin gene were located telomeric to the contig. Comparison between the human and porcine MHC complexes showed a perfect conserved order of anchor genes, whereas no orthologous relationships were found for the class I loci.     

Interferon-induced expression of class II major histocompatibility antigens in the major histocompatibility complex (MHC) class II deficiency syndrome

Class II antigens encoded by genes of the major histocompatibility complex (MHC) are expressed by a variety of cell types and have a vital role in the cellular interactions required for an effective immune response. We have analyzed the regulation of HLA-DR, DP, and DQ class II antigen expression on cells of different lineage from an immunodeficient patient with the MHC class II deficiency syndrome. T and B lymphocytes, monocytes, and fibroblasts, which initially expressed no class II antigens, were treated with inductive stimuli that normally lead to enhanced expression of class II antigens. Monocytes, but not fibroblasts, cultured for 48–96 hr in the presence of recombinant gamma interferon expressed all three types of class II antigens. In contrast, T lymphocytes did not express class II antigens following their exposure to a variety of stimuli, including activation with phytohemagglutinin and culture in the presence of interleukin-2, transformation by the retrovirus HTLV-1 or HTLV-2, or exposure to the demethylating agent 5-azacytidine. Similarly, class II antigens were not induced on B cells by cross-linkage of surface immunoglobulin molecules with anti-mu, exposure to Epstein-Barr virus, or treatment with soluble factors secreted by activated T cells. These results demonstrate that the regulation of class II MHC antigen expression by monocytes and lymphocytes is dissimilar and suggest that different regulatory genes are involved in the control of class II antigen expression by cells of different lineage.     

The outermost N‐terminal region of tapasin facilitates folding of major histocompatibility complex class I

Tapasin (Tpn) is an ER chaperone that is uniquely dedicated to MHC‐I biosynthesis. It binds MHC‐I molecules, integrates them into peptide‐loading complexes, and exerts quality control of the bound peptides; only when an “optimal peptide” is bound will the MHC‐I be released and exported to the cell surface for presentation to T cells. The exact mechanisms of Tpn quality control and the criteria for being an optimal peptide are still unknown. Here, we have generated a recombinant fragment of human Tpn, Tpn_1–87 (representing the 87 N‐terminal and ER‐luminal amino acids of the mature Tpn protein). Using a biochemical peptide–MHC‐I‐binding assay, recombinant Tpn_1–87 was found to specifically facilitate peptide‐dependent folding of HLA‐A*0201. Furthermore, we used Tpn_1–87 to generate a monoclonal antibody, αTpn_1–87/80, specific for natural human Tpn and capable of cellular staining of ER localized Tpn. Using overlapping peptides, the epitope of αTpn_1–87/80 was located to Tpn_40–44, which maps to a surface‐exposed loop on the Tpn structure. Together, these results demonstrate that the N‐terminal region of Tpn can be recombinantly expressed and adopt a structure, which at least partially resembles that of WT Tpn, and that this region of Tpn features chaperone activity facilitating peptide binding of MHC‐I.     

Mutations in the major histocompatibility complex class I antigen-presenting groove affect both negative and positive selection of T cells

In several transgenic mouse models T cell development was shown to be controlled by the binding of the alpha/beta T cell receptor (TcR) to ligands in the thymus. In transgenic mice expressing a male-specific TcR alpha/beta, the presence of the restricting D major histocompatibility complex (MHC) molecule plus the male specific peptide deleted thymocytes at an early stage of development. On the other hand, maturation of T cells required an interaction of the TcR with the thymic D MHC molecules in the absence of specific peptides. This could imply that negative and positive selection of this receptor are affected differently by mutations in the HY peptide-binding groove of the D MHC molecule. Such mutants have been isolated and were shown to affect the response to HY antigen in that both the bm14 (residue Glu70----Asp) and the bm13 (residue Leu114----Glu, Phe116----Tyr and Glu119----Asp) strains do not normally mount cytotoxic responses to male cells. Here we show that these mutations affect antigenicity of male cells, as well as negative and positive selection of T cells in TcR alpha/beta transgenic mice.     

Variation and linkage disequilibrium between a structurally polymorphic Alu located near the OR12D2 gene of the extended major histocompatibility complex class I region and HLA‐A alleles

We investigated the genetic structure and population frequency of an Alu repeat dimorphism (absence or presence) located near the OR12D2 gene within the olfactory receptor gene region telomeric of the alpha HLA class I region (HLA‐J, ‐A, ‐G, ‐F). The structurally polymorphic Alu insertion (POALIN) locus rs33972478 that we designated as AluOR and its allele and haplotype frequencies and association with HLA‐A and six other structurally polymorphic retroelements (3 Alu, 2 SVA and an HERVK9) were determined in 100 Japanese, 174 Caucasians and 100 African American DNA samples. The AluOR insertion varied in population frequency between 14.4% and 31.5% with significant differences between the Japanese and Caucasians, but not between the Caucasian and African Americans. Although AluOR is located 600 kb from the HLA‐A gene, there was a significant linkage disequilibrium between the two loci and a high percentage association of the AluOR insertion with HLA‐A29 (79%) in Caucasians and HLA‐A31 (69.4%) in Japanese. Inferred haplotypes among three‐locus to eight‐locus haplotype structures showed maximum differences between the populations with the eight‐locus haplotypes. The most frequent multilocus haplotype shared between the populations was the HLA‐A2 allele in combination with the AluHG insertion. The AluOR whether investigated alone or together with the HLA class I alleles and other dimorphic retroelements is an informative ancestral marker for the identification of lineages and variations within the same and/or different populations and for examining the linkage and crossing‐over between the HLA and OR genomic regions in the extended MHC.     

Genes outside the major histocompatibility complex control resistance and susceptibility to experimental Lyme arthritis

The mechanism controlling the development of experimental Lyme arthritis remains poorly understood. Mice with the H-2^k haplotype have generally been thought to be more susceptible to Lyme arthritis development than those with the H-2^d haplotype. In the present study the role of genes within the major histocompatibility complex (MHC) in determining resistance or susceptibility to the development of experimental Lyme arthritis was investigated. We found that C3H congenic mice were equally susceptible, and DBA congenic mice equally resistant, to arthritis development regardless of their H-2 haplotype (H-2^k or H-2^d). These results indicate that genes outside the murine MHC are the major determinants of both resistance and susceptibility to the development of experimental Lyme arthritis. Received: 23 March 2000     

Properties and applications of single-chain major histocompatibility complex class I molecules

Stable major histocompatibility complex (MHC) class I molecules at the cell surface consist of three separate, noncovalently associated components: the class I heavy chain, the β(2)-microglobulin light chain, and a presented peptide. These three components are assembled inside cells via complex pathways involving many other proteins that have been studied extensively. Correct formation of disulfide bonds in the endoplasmic reticulum is central to this process of MHC class I assembly. For a single specific peptide to be presented at the cell surface for possible immune recognition, between hundreds and thousands of peptide-containing precursor polypeptides are required, so the overall process is relatively inefficient. To increase the efficiency of antigen presentation by MHC class I molecules, and for possible therapeutic purposes, single-chain molecules have been developed in which the three, normally separate components have been joined together via flexible linker sequences in a single polypeptide chain. Remarkably, these single-chain MHC class I molecules fold up correctly, as judged by functional recognition by cells of the immune system, and more recently by X-ray crystallographic structural data. This review focuses on the interesting properties and potential of this new type of engineered MHC class I molecule.     

Delayed onset of graft‐versus‐host disease in immunodeficent human leucocyte antigen‐DQ8 transgenic,murine major histocompatibility complex class II‐deficient mice repopulated by human peripheral blood mononuclear cells

Haematopoietic humanization of mice is used frequently to study the human immune system and its reaction upon experimental intervention. Immunocompromised non‐obese diabetic (NOD)‐Rag1^–/– mice, additionally deficient for the common gamma chain of cytokine receptors (γc) (NOD‐Rag1^–/– γc^–/– mice), lack B, T and natural killer (NK) cells and allow for efficient human peripheral mononuclear cell (PBMC) engraftment. However, a major experimental drawback for studies using these mice is the rapid onset of graft‐versus‐host disease (GVHD). In order to elucidate the contribution of the xenogenic murine major histocompatibility complex (MHC) class II in this context, we generated immunodeficient mice expressing human MHC class II [human leucocyte antigen (HLA)‐DQ8] on a mouse class II‐deficient background (Aβ^–/–). We studied repopulation and onset of GVHD in these mouse strains following transplantation of DQ8 haplotype‐matched human PBMCs. The presence of HLA class II promoted the repopulation rates significantly in these mice. Virtually all the engrafted cells were CD3⁺ T cells. The presence of HLA class II did not advance B cell engraftment, such that humoral immune responses were undetectable. However, the overall survival of DQ8‐expressing mice was prolonged significantly compared to mice expressing mouse MHC class II molecules, and correlated with an increased time span until onset of GVHD. Our data thus demonstrate that this new mouse strain is useful to study GVHD, and the prolonged animal survival and engraftment rates make it superior for experimental intervention following PBMC engraftment.     

The "adjuvant effect" of the polymorphic B-G antigens of the chicken major histocompatibility complex analyzed using purified molecules incorporated in liposomes

The polymorphic B-G region of the chicken major histocompatibility complex has previously been shown to mediate an "adjuvant effect" on the humoral response to other erythrocyte alloantigens. We demonstrate here that B-G molecules purified with monoclonal antibodies exert this adjuvant effect on the production of alloantibodies to chicken class I (B-F) molecules, when the two are in the same liposome. The adjuvant effect may in part be mediated by antibodies, since the antibody response to B-G molecules occurs much faster than the response to B-F molecules, and conditions in which antibodies to B-G are present increase the speed of the response to B-F molecules. We also found that the presence of B-G molecules in separate liposomes results in a lack of response to B-F molecules. In the light of this and other data, we consider the possible roles for the polymorphic B-G molecules, particularly for the generation of B cell diversity, in the immune systems of birds and other animals.     

Structures and homology modeling of chicken major histocompatibility complex protein class I (BF2 and beta2m)

In order to elucidate the two-dimensional (2D) and three-dimensional (3D) structures of chicken major histocompatibility complex (MHC) class I protein (BF2 and beta2m) and further reconstruct their complex identifying the virus-derived antigenic peptides, the mature protein of BF2 and beta2m genes were expressed solubility in pMAL-p2X/Escherichia coli. TB1 system. The expressed MBP-BF2- and MBP-beta2m-fusion proteins were purified, and cleaved by the factor Xa protease. Subsequently, the monomers were further separated, and the purified MBP-BF2, -beta2m, and MBP were analyzed by circular dichroism (CD) spectrum. The contents of alpha-helix, beta-sheet, turn, and random coil in BF2 protein were 72, 102, 70, and 90 amino acids (aa), respectively. The beta2m proteins displayed a typical beta-sheet and the contents of alpha-helix, beta-sheet, turn, and random coil were 0, 46, 30, and 22 aa, respectively. Homology modeling of BF2 and beta2m proteins were similar as the 3D structure of human MHC class I (HLA-A2). The results showed that pMAL-p2X expression and purification system could be used to obtain the right conformational BF2 and beta2m proteins, and the 2D and 3D structures of BF2 and beta2m were revealed to be similar to human's. The recombinant BF2 and beta2m-based proteins might be a powerful tool for further detecting antigenic peptides.     

Transfection of major histocompatibility complex class I and class II genes causes tumour rejection

Many human and mouse tumours do not express MHC class II antigens and have reduced levels of class I antigens. Because of the requirement for class I and/or class II antigen for antigen presentation to Th and Tc cells, these phenotypes may enable tumour cells to 'escape' the host's immune response. Experiments presented here are designed to assess the role of MHC class I and class II antigens in tumour immunity, and to overcome the MHC class I- or class II-negative phenotype. When transfected with the syngeneic H-2Db gene, the MHC antigen-negative 402AX teratocarcinoma expresses high levels of H-2Db antigen. 402AX/Db cells are rejected by MHC allogeneic and some MHC syngeneic 402AX-susceptible mice, however the fully syngeneic strain of origin (129) remains tumour-susceptible. Induction of MHC class I gene products on class I antigen-negative embryonal carcinoma cells therefore increases tumour immunogenicity in some hosts, but not in the fully syngeneic mouse. In an attempt to enhance antigen presentation of tumour-associated antigens to Th cells, MHC class I antigen-positive SaI (KkDd) sarcoma cells were transfected with syngeneic A alpha k and A beta k genes to generate Iak-expressing tumour cells. SaI/Ak cells are efficiently rejected by syngeneic A/J (KkDd) mice, while untransfected SaI cells are lethal. Induction of MHC class II antigen expression on the class I antigen-positive SaI sarcoma therefore completely abrogates malignancy.