Chicken MHC Research Articles

Amino acid sequences and structures of chicken and turkey beta2-microglobulin

The complete amino acid sequences of chicken and turkey beta2-microglobulins have been determined by analyses of tryptic, V8-proteolytic and cyanogen bromide fragments, and by N-terminal sequencing. Mass spectrometric analysis of chicken beta2-microglobulin supports the sequence-derived M r of 11,048. The higher apparent M r obtained for the avian beta2-microglobulins as compared to human beta2-microglobulin by SDS-PAGE is not understood. Chicken and turkey beta2-microglobulin consist of 98 residues and deviate at seven positions: 60, 66, 74–76, 78 and 82. The chicken and turkey sequences are identical to human beta2-microglobulin at 46 and 47 positions, respectively, and to bovine beta2-microglobulin at 47 positions, i.e. there is about 47% identity between avian and mammalian beta2-microglobulins. The known X-ray crystallographic structures of bovine beta2-microglobulin and human HLA-A2 complex suggest that the seven chicken to turkey differences are exposed to solvent in the avian MHC class I complex. The key residues of beta2-microglobulin involved in alpha chain contacts within the MHC class I molecule are highly conserved between chicken and man. This explains that heterologous human beta2-microglobulin can substitute the chicken beta2-microglobulin in exchange studies with B-F (chicken MHC class I molecule), and suggests that the MHC class I structure is conserved over long evolutionary distances.

Structure and expression of a chicken MHC class I gene

In the present communication we report the primary structure of an expressed chicken class I gene, B-FIV, of the CB-B 12 haplotype

MHC-like molecules in some nonmammalian vertebrates can be detected by some cross-reactive xenoantisera.

Rabbit antisera raised to human and chicken MHC molecules were used to immunoprecipitate cross-reactive molecules from biosynthetically and cell surface-labeled spleen and/or blood cells of representative vertebrate species. Five major points emerged: 1) There were many nonspecific cross-reactions using these techniques, so various criteria were developed to distinguish these from true MHC-like molecules. 2) Only very small subpopulations of immunogen-specific antibodies cross-reacted with MHC-like molecules in other nonmammalian species. These subpopulations were different for each species and even within a species, sometimes being so limited as to behave like alloantisera. This led to a very scattered pattern of true cross-reactions that sometimes failed to reflect the properties of the bulk antibody population. 3) Antisera containing antibodies to class II beta- and class I alpha-chains cross-reacted better and more widely than those to B-G, class II alpha and, in general, beta 2-microglobulin. 4) Some cross-reactive antibodies were clearly directed to epitopes on the surface of the mature heterodimers, but many seemed to recognize nonlinear cryptic determinants, presumably in the contact regions between the chains. These latter antibodies recognized biosynthetic intermediates and also a variety of unusual cell surface MHC-like molecules present in reptile and amphibian, but absent in the mammal and chicken cells tested. These included E homodimers whose relationship to chicken B-G molecules is unknown. 5) MHC-like molecules were identified in a bird, three reptiles, and two amphibians, but no molecules with the expected properties were found with these reagents in any of the fish tested.

Genetic organization of the chicken MHC

Genetic organization of the chicken MHC

Molecular Genetics of the Chicken MHC: Current Status and Evolutionary Aspects

Etude de l'evolution moleculaire du complexe B, complexe d'histocompatibilite majeur du poulet. Le clonage moleculaire des genes codant pour les antigenes de classe I et une analyse biochimique de la sequence en aminoacides des produits de ces genes ont ete realises

Ontogenic Appearance of MHC Class I (B‐F) Antigens During Chicken Embryogenesis

Expression of chicken MHC class (B-F) antigens during ontogeny was determined by binding of anticlass antibody and appearance of B-F transcripts by Northern blotting in chicken organs during embryogenesis until 2 weeks after hatching. MHC class transcripts first become detectable in day 6.5 of embryogenesis. B-F cell-surface expression first becomes detectable in hemopoietic organs by day 10‑12 of embryogenesis and somewhat later in nonhemopoietic organs. Flow cytometry analysis of hemopoietic cells throughout embryogenesis revealed B-Fhi and B-F1 cell populations. The percentage of B-F cells in spleen and bone marrow decreased around hatching, which could reflect either cell flows in these organs during this period or the sensitivity of hemopoietic cells to hatching stress.

B-G cDNA clones have multiple small repeats and hybridize to both chicken MHC regions.

We used rabbit antisera to the chicken MHC erythrocyte molecule B-G and to the class I alpha chain (B-F) to screen lambda gt11 cDNA expression libraries made with RNA selected by oligo-dT from bone marrow cells of anemic B19 homozygous chickens. Eight clones were found to encode B-G molecules which hybridize with sequences in the chicken MHC as defined by congenic strains; the fusion proteins react with multiple immune but not preimmune sera, they select antibodies from the antisera to B-G, which then react with distinct erythrocyte B-G protein patterns, and they elicit antibodies from mice which in turn react with authentic B-G proteins. None of the clones represent a complete message, some--if not all--bear introns, and none of them match with any sequences presently stored in the data banks. The following new information did, however, emerge. At least two homologous transcripts are present in this homozygous chicken, thereby formally proving the existence of an expressed multigene family. The 3' ends (3'UT) are simple sequences with 80% nucleotide identity between clones, while the 5' ends (either coding or noncoding) are composed of multiple short repeats which are far less similar. These repeats could explain the bewildering variation in size of B-G proteins within and between haplotypes. Southern blots of genomic chicken DNA gave complex patterns for most probes, with many bands in common using different probes, but few bands in common between haplotypes. The sequences detected are all present in the MHC, based on the congenic lines CB and CC. Most of these sequences map into the B-G region, but some map into the B-F/B-L region as defined by the haplotypes B15, B21, and their apparently reciprocal recombinants B21r3 and B15r1.

Antigen-presenting cell-T cell interaction in the chicken is MHC class II antigen restricted.

The involvement of the MHC in the recognition of Ag by avian T lymphocytes was analyzed. PBL from chickens primed with keyhole limpet hemocyanin in vivo were induced to synthesize DNA in an in vitro response to specific Ag. Responding cells were T cells as judged by immunofluorescence staining. In vivo Ag-primed PBL were stimulated in vitro with specific Ag and further propagated in the presence of IL-2. Subsequent Ag-specific T cell proliferation required the presence of Ag-pulsed peripheral blood adherent cells (APC). T cell responses were restricted by the MHC of the APC; Ag presented by allogeneic APC did not support T cell proliferation. By using MHC-recombinant chicken lines, the gene products controlled by MHC class II loci were shown to restrict the T cell-APC interaction. This conclusion was substantiated by the inhibition of the Ag-specific T cell response by a mAb against chicken MHC class II gene products but not by a mAb against chicken MHC class I gene products.

Genetics and Classification of Major Histocompatibility Complex Antigens of the Chicken

The chicken MHC consists of a gene complex divisable into three chromosomal regions, each producing a series of genetically antithetical molecules with regionally specific functions. Most of the genes (alleles) of each region are present in unique haplotype combinations with genes (alleles) from the other two regions; relatively few haplotypes appear to share identical regional genes. Even with the high degree of polymorphism existing within and between regions, general typing of erythrocytes for MHC haplotypes can still be performed as economically as ever with alloimmune reagents of known specificity.

Isolation and characterization of chicken and turkey beta2-microglobulin

Chicken and turkey beta2-m were isolated from citrated plasma in sequential use of three Chromatographie steps: affinity chromatography, gel filtration chromatography and anion-exchange chromatography. The purified protein was identified as beta2-m by reaction with a beta2-m specific monoclonal antibody and by the ability to recombine with the chicken MHC class I heavy chain. The purity was estimated by SDS-PAGE and IEF. The pI was between 5.1 and 5.3 for chicken beta2-m and 4.7 and 4.8 for turkey beta2-m, which fact is reflected in their different electrophoretic mobilities in agarose gel (turkey migrates in the alpha and chicken migrates in the beta region). The mol. wt of both chicken and turkey beta2-m was 14,500 estimated by SDS-PAGE whereas calculations based on the amino acid compositions gave mol. wts of 11,000. E M 280was 15.9 for chicken beta2-m and 16.4 for turkey beta2-m. The amino acid compositions and sequences of the two avian beta2-m molecules have been compared with earlier data from the literature. The sequence of the 23 N-terminal amino acids was found to be identical in our preparations from both chicken and turkey, namely DLTPKVQVYSRFPASAGTKNVLN, and is incompatible with a previously published sequence also thought to be from turkey beta2-m. Reasons for our opinion that the molecules isolated and sequenced in this paper are the correct ones are given.

Structure, biosynthesis, and polymorphism of chicken MHC class II (B-L) antigens and associated molecules.

Chicken MHC class II (B-L) antigens were immunoprecipitated by the monoclonal antibody TaP1 from inbred chicken splenic leukocytes and a lymphoblastoid B cell line (RP9), and were studied by two dimensional gel electrophoresis. B-L antigens are composed of one alpha and one beta chain that are noncovalently bound at the cell surface. In all haplotypes studied, a single acidic 34,000 dalton non-polymorphic chain was observed, whereas two polymorphic chains could be distinguished, differing in both pH and m.w. The alpha-beta heterodimer is associated during its maturation in the cytoplasm with several basic invariant molecules with m.w. ranging from 30,000 to 42,000 daltons. Treatment of cells with tunicamycin and treatment of immunoprecipitated molecules with several glycosidases revealed a complex process of maturation for all of these molecules. The alpha and beta chains undergo a N-glycosylation of complex type, whereas the invariant molecules bear N-linked high mannose glycans, and perhaps also O-linked glycans in the RP9 lymphoblastoid line. Overall, the B-L antigens appear very similar to the HLA-DR and I-E antigens.

Analysis of chickens for recombination within the MHC (B-complex).

In an attempt to further map the chicken MHC (the B complex), a systematic search for genetic recombinants within the B complex was performed by serotyping the progeny from F2 crosses of chickens by means of specific anti-class I, anti-class II, and anti-class IV alloantisera. Two recombinant B-haplotypes (B21r and B15r) were found by analysing 2,656 F2 chickens representing 5,312 informative typings. In either case, the B-G (class IV) allele was recombined with both the B-F and B-L alleles of the opposite haplotype. MLC typings, tests for direct compatibility by GVH reactions, and absorption analyses confirmed the original serological typing of the two recombinant B haplotypes. No recombination between B-F (class I) and B-L (class II) loci was found. This very low frequency of recombination within the B complex as compared with recombination frequencies found in mammalian MHC's is discussed.

Mouse monoclonal antibodies to class I and class II antigens of the chicken MHC. Evidence for at least two class I products of the B complex.

Mouse monoclonal antibodies to class I and class II antigens of the chicken MHC. Evidence for at least two class I products of the B complex.

A monoclonal antibody against chicken MHC class I (B-F) antigens.

A monoclonal antibody against chicken MHC class I (B-F) antigens.

Chicken major histocompatibility complex and disease.

The chicken MHC (B complex) initially described by Briles as controlling blood antigens, is now known to be composed of at least three regions, L, F and G. Two of these, F and G, were described on the basis of recombinants found in a study of over 10,000 chickens. On the basis of biochemical, tissue distribution and functional analyses, F corresponds to the murine H-2 K/D regions. The G region is unique to the chicken since the antigenic product is expressed only on erythrocytes and their progenitors. L was identified by serological studies and corresponds to the H-2 I region; the L antigen is expressed predominantly on B lymphocytes, monocytes and 10% of T lymphocytes, and differences in the L region result in variations in immune responsiveness. A number of functional similarities exist between the chicken MHC and that of other species such as regulation of graft rejection, graft-versus-host reaction (GVHR) and mixed lymphocyte reactions (MLR), mitogenic and immune responsiveness and resistance to RNA and DNA virus infection. The chicken MHC also controls the severity of autoimmune disease, as exemplified by the spontaneous thyroiditis of Obese strain (OS) chickens. It differs from mammalian MHC's by having of lower crossing-over frequency and no apparent gene duplication.

Glyoxalase 1 (GLO) in the chicken: genetic variation and lack of linkage to the MHC.

Electrophoretic variation of glyoxalase 1 (GLO) has been detected in chicken red-cell lysates. Three phenotypes are shown to be inherited through a diallelic system, just as in humans and mice. The chicken GLO phenotype differ from their mammalian counterparts in that one of the homozygotes is devoid of GLO activity. The heterozygote produces two bands, while the other homozygote yields a single band of GLO activity with mobility equal to the faster of these two bands. In noninbred White Leghorn birds, the GLO2 allele occurred significantly more often in birds homozygous for the B1 allele at the chicken MHC than in those homozygous for B19, suggesting that the products of these loci may have population associations in the chicken. Absence of close linkage between the GLO and B loci was, however, demonstrated by appropriate test crosses.

Structure and properties of the major histocompatibility complex of the chicken. Speculations on the advantages and evolution of polymorphism.

Our knowledge of the chicken MHC has greatly expanded since the last review article on the subject (Pazderka et al; 1975 a) through the great progress that has been made in the areas of genetic structure, biochemistry, disease resistance and serology of the chicken MHC. It is becoming increasingly apparent that the chicken MHC is very similar in most respects to mammalian MHC's. Most of the immunologically related phenomena or functions which are known for H-2 alleles are now known for the chicken MHC. These include the determination of serologically detectable antigens, histocompatibility antigens, immune responses, serum proteins, resistance to virus-induced malignancy and cell-cell interactions. The finding of all these immunological functions grouped together in close linkage disequilibrium in species as different as chickens, mice, and humans, as well as the striking homologies between MHC antigens of these species strongly favors the view that MHC's evolved and persisted in association with some functional advantages to the species. Apart from the interesting phylogenetic aspects one might naturally ask the question"why study the chicken MHC?" The answer to this question is quite simply that basic studies on the chicken MHC have given uniquely important clues to the functional advantage of MHC polymorphism as well as the survival value for the species of certain MHC alleles. The study of inbred mice has told us a great deal about the biochemistry of MHC antigens and the fine structure of the MHC but tells us little about natural selection. The herpes virus causing Marek's disease is an important agent of natural selection in the chicken and one MHC haplotype is strongly associated with resistance to this contagious lymphoma in several unrelated populations of chickens, including the progenitor of the species. Other MHC haplotypes of the chicken are associated with long life span and general fitness, or with high adult mortality. Other basic studies on the chicken MHC have revealed the widespread existence of natural antibodies against MHC as well as other polymorphic cell-surface antigens. These studies have also shown that the immune response of mice against

Role of the B-G-region antigen in the humoral immune response to the B-F-region antigen of chicken MHC.

In the chicken MHC there exist two regions, designated F and G, which were separated by crossing-over. The F region contains genes controlling all functions characteristic of the MHC. So far only one gene has been assigned to the G region and it is responsible for the presence of an RBC antigen. When cross-immunizing animals of the congenic lines CB and CC with erythrocytes, we have found that both F- and G-specific antibodies were produced. By using the recombinant haplotypes Br1 and Br2 we were able to dissociate the F from the G antigen and immunize with them separately. It was found that production of F antibodies required the copresence of the G antigen, whereas G antibodies were formed regardless of the presence of absence of the F-region antigen. It could be demonstrated that a prerequisite of the role of the G antigen with respect to the F antigen was the localization of both antigens on the same erythrocyte. Possible mechanisms underlying this phenomenon are discussed.

Evidence for two populations of B-L (Ia-like) molecules encoded by the chicken MHC.

Two specific alloantisera detecting B-L (Ia-like) antigens on chicken lymphocytes of the B6 and B15 haplotypes were found to cross-react strongly. Anti-B-L6 and anti-B-L15 alloantisera both reacted with B-L molecules on B6 and B15 lymphocytes as demonstrated by immunofluorescence and SDS-PAGE analysis of 125I-labeled B-L antigens isolated by incubation with anti-B-L alloantisera. Absorption studies showed that the anti-B-L alloantisera reacted with at least two kinds of antigenic determinant, one set shared by B-L6 and B-L15 molecules and another set specific for each haplotype. In spite of the absence of genetic evidence for more than one B-L locus in the chicken B complex, it was shown by sequential antibody incubations that these two different B-L antigenic determinants are associated with at least two separate species of B-L molecules, indicating the presence of at least two B-L loci within the MHC of the chicken.

Nomenclature for chicken MHC (B) antigens defined by monoclonal antibodies.

Nomenclature for chicken MHC (B) antigens defined by monoclonal antibodies.