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Rothermel, Ellen; Heine, Lutz; Wurst, Wolfgang; Günther, Eberhard
doi: 10.1007/BF00190895pmid: 8482583
The cDNA and a partial genomic sequence of a rat class I major histocompatibility (RT1) gene, 11/3R, is reported here. The sequence contains several unique amino acid residues at certain positions, mutations in exon 7 (which is not expressed), a mutation of the canonical exon 8 stop codon to a sense codon, and includes a long 3′ unstranslated region (utr). The structure of exon 7 differs from that found in most rat class I genes and resembles exon 7 of most H-2K,D,L.Q genes. Parts of the 3′ noncoding region are homologous to the RT1.A-4 and certain H-2 genes. Expression is detectable by northern blot analysis in mitogen-stimulated lymphocytes only, by polymerase chain reaction (PCR) in each tissue tested. After transfection into L cells 11/3R can be shown to be expressible at the cell surface. Probes derived from the 3′ noncoding part crosshybridize with a number of restriction fragments which map to the RT1.C region, thus defining a subfamily of RT1.C region genes. Several members of this subfamily are deleted in the ιM1 RT1 mutant. The 11/3R gene presents typical features of a class Ib gene. Aspects of evolution and the potential of the gene are discussed.
Charmley, Patrick; Concannon, Patrick
doi: 10.1007/BF00190896pmid: 8482584
The Vb6 subfamily is the largest reported subfamily of human T-cell receptor (Tcr) genes with as many as 14 possible members based on variation in reported DNA sequences. A study of the genomic organization of four distinct Vb6 genes indicated that they contained within their introns theuniterrupted dinucleotide repeat (GT)n, with n>8. DNA amplification primers and conditions were determined which amplified the intron of these four different Vb6 gene segments. All four Vb6 genes tested showed length polymorphism when examined in a group of unrelated individuals. Careful sizing and DNA sequencing showed that the alleles of each gene differed in size by multiples of two base pairs (bp), due to different repeat numbers of the dinucleotide (GT)n. These four microsatellite polymorphisms had from three to ten alleles, and individual heterozygosities of 26% to 83%. The large number of alleles and the high heterozygosity make these polymerase chain reaction (PCR)-based polymorphisms very attractive genetic markers for segregation studies which postulate the presence of autoimmune susceptibility genes within the Tcrb region. Vb6 hybridization to genomic DNA confirmed the relatively large size of the Vb6 subfamily in several hominoid species. Nucleotide sequencing of an intron of the Vb6 genes from other primates revealed the presence of dinucleotide repeats similar to those found in human Vb6 genes. Thus, the (GT)n microsatellite was not only present in the Vb6 intron before Vb6 gene duplication, but was present before speciation of the hominoids.
Schreuder, Geziena; Pool, Jos; Blokland, Els; Els, Cécile; Bakker, Astrid; Rood, Jon
doi: 10.1007/BF00190897pmid: 8482585
An analysis of the genetic traits of human minor histocompatibility (mH) antigens is, unlike with inbred mice, rather complicated. Moreover, the fact that mH antigens are recognized in the context of MHC molecules creates an additional complication for reliable segregation analysis. To gain insight into the mode of inheritance of the mH antigens, we relied upon a series of HLA-A2-restricted cytotoxic T-cell (CTL) clones specific for four mH antigens. To perform segregation analysis independent of HLA-A2 gene: we transfected HLA-A2-negative cells with the HLA-A2 gene: this results in the cell surface expression of the HLA-A2 gene product and, if present, mH antigen recognition. The mode of inheritance of the HLA-A2-restricted mH antigens HA-1, -2, -4, and -5 was analysed in 25 families whoese members either naturally expressed positive. Analysis of distribution of the mH antigens in the parent population among the mating types, together with their inheritance patterns in the families, demonstrated that the four mH antigens behaved as Mendelian traits, whereby each can be considered a product of a gene with two alleles, one expressing and one not expressing the detected specificity. We also showed that the loci encoding the HA-1 and HA-2 antigens are not closely linked to HLa (lod scores Z (0 = 0.05) <−4.0). Some indication was obtained that the HA-4- and HA-5-encoding loci may be losely linked to HLA. While we are aware of the limited results of this nonetheless comprehensive study, we feel the similarity in immunogenetic traits between human and mouse mH antigens is at least striking.
Hermel, Evan; Robinson, Peter; She, Jin-Xiong; Fischer Lindahl, Kirstein
doi: 10.1007/BF00190898pmid: 8482575
Mouse β2-microglobulin (β2m) is polymorphic. Sequences of five allelic wild mouse B2m genes have been determined from the large exons of genomic DNA using the polymerase chain reaction. Relative to the standard B2m a allele, the products of four alleles of Mus musculus origin (w2, w3, w4, and w5), differ by only one or two amino acids. w5 has a single nucleotide change, Asp85 → Val, and is identical to the c allele. w2 differs at Arg81 → Thr and w4 at His34 → Gln, and they share the Asp85 → Val change with B2m c and B2m w5.w5 and c cells are lysed by S19.8, a monoclonal antibody specific for β2mb (Ala85), in a complement-mediated cytotoxicity assay, whereas w4 cells are not. Thus, distant changes appear to introduce subtle conformational effects on β2m structure. Five independent isolates of Mus spretus (w1) differ the most from B2m a, with 12 amino acid changes and only one silent substitution. Replacements predicted from the nucleotide sequence occur in loops of the molecule facing away from the class I heavy chain and not in regions where β2m associates with class 1 α3 domains. Concordantly, the w1 – 5 allelic forms of β2m associate well with H-2 heavy chains. The many amino acid changes in the spretus sequence and the paucity of silent substitutions suggest that B2m has been subject to positive selection.
Boucraut, José; Guillaudeux, Thierry; Alizadeh, Mehdi; Boretto, Joelle; Chimini, Giovanna; Malecaze, François; Semana, Gilbert; Fauchet, Renée; Pontarotti, Pierre; Bouteller, Philippe
doi: 10.1007/BF00190899pmid:
Roopenian, Derry; Christianson, Greg; Davis, Allan; Zuberi, Aamir; Mobraaten, Larry
doi: 10.1007/BF00190900pmid: 7683307
The purpose of this study was to elucidate the genetic origin of minor histocompatibility (H) antigens. Toward this end common inbred mouse strains, distinct subspecies, and species of the subgenus Mus were examined for expression of various minor H antigens. These antigens were encoded by the classical minor H loci H-3 and H-4 or by newly identified minor H antigens detected as a consequence of mutation. Both minor H antigens that stimulate MHC class I-restricted cytotoxic T cells (Tc) and antigens that stimulate MHC class II-restricted helper T cells (Th) were monitored. The results suggested that strains of distinct ancestry commonly express identical or cross-reactive antigens. Moreover, a correlation between the lack of expression of minor H antigens and ancestral heritage was observed. To address whether the antigens found on unrelated strains were allelic with the sensitizing minor H antigens or a consequence of antigen cross-reactivity, classical genetic segregation analysis was carried out. Even in distinct subspecies and species, the minor H antigens always mapped to the site of the appropriate minor H locus. Together the results suggest: 1 minor H antigen sequences are evolutionarily stable in that their pace of antigenic change is slow enough to predate subspeciation and speciation; 2 the minor H antigens originated in the inbred strains as a consequence of a rare polymorphism or loss mutation carried in a founder mouse stock that caused the mouse to percieve the wild-type protein as foreign; 3 there is a remarkable lack of antigenic cross-reactivity between the defined minor H antigens and other products.
doi: 10.1007/BF00190901pmid: 8482576
Nucleotide sequence analysis of rhesus macaque major histocompatibility complex class I cDNAs allowed the identification of the orthologue of HLA-F, designated Mamu-F. Comparison of Mamu-F with earlier published human and chimpanzee orthologues demonstrated that these sequences share a high degree of similarity, both at the nucleotide and amino acid level, whereas a New World monkey (cotton-top tamarin) equivalent is more distantly related. Exon 7, encoding one of the cytoplasmatic domains, is absent for all primate Mhc-F cDNA sequences analyzed so far. In contrast to the human, chimpanzee, and rhesus macaque equivalents, the cotton-top tamarin Saoe-F gene seems to have accumulated far more nonsynomynous than synonymous differences.
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Polymorphic as well as HLA-F and -G genes are repressed in the human cell line JAR, derived from a tumor of trophoblast origin. By contrast, the HLA-E gene as well as the non-HLA novel coding-sequence, R1, located 5′ to HLA-E, both remain transcriptionally active. We first demonstrated the role of DNA methylation in the repression of class I genes (except HLA-E) in JAR by the use of the 5-Azacytidine demethylating agent. Following treatment, JAR clones reexpressed polymorphic class I transcripts and cell surface α chains. Using methylation-sensitive rare cutter enzymes on JAR genomic DNA, followed by classical or pulse field gel electrophesis and hybridization with HLA locus-specific probes, we found methylated CpG islands in the 5′ region of all class I genes, except for HLA-E. These results, establishing an inverse relationship between states of methylation and transcriptional activity within the MHC class I chromosomal region in JAR, and the observations that the HLA-E and R1 genes were ubiquitously expressed, suggest that the HLA-E chromosomal domain might have functional importance including the presence of housekeeping genes.