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C LeGuern

Publications and source records attributed to C LeGuern.

At least 55 records · Page 3Linked to original sources
PubMed1992

Characterization of a polymorphism of CD4 in miniature swine.

A polymorphism of CD4 in miniature swine has been identified by failure of cells from some animals to react with mAb 74-12-4. The phenotypic, molecular genetic, and functional characteristics of these animals have been defined. Cells from heterozygous animals bear approximately 50% the number of 74-12-4-reactive molecules on their surface as do cells from animals homozygous for the wild type. Animals of both phenotypes demonstrate similar flow cytometric profiles for CD8+ T cells. Northern blot analysis confirms the presence of mRNA for CD4 among PBL of animals failing to stain with 74-12-4. CD4 allelism is confirmed by Southern blot analysis, revealing RFLP. Function of the CD4 subset in vivo, as demonstrated by antibody production against a T cell-dependent Ag, is similar between animals of both phenotypes. Proliferative responses to PHA and alloantigen stimulation by a full haplotype mismatch or a class II mismatch alone are equivalent for animals of both phenotypes. These data suggest that the allelic form of CD4, designated CD4.2 in contrast to the wild-type CD4.1, is capable of performing normally as an accessory molecule in the generation of immune responses. Furthermore, antixenogeneic responses to C57B10.BR were equivalent, suggesting that both CD4 molecular types may be capable of interacting with xenogeneic class II molecules. Although the polymorphism includes differences in exons 3 and 4, regions thought to encode portions of the molecule interacting with MHC class II, these results imply that this naturally occurring CD4 polymorphism does not affect the interaction with class II molecules.

Animals↗
PubMed1991

Expression of a swine class II gene in murine bone marrow hematopoietic cells by retroviral-mediated gene transfer.

As a first step in assessing the efficacy of a gene transfer approach to the induction of transplantation tolerance in our miniature swine model, double-copy retroviral vectors engineered to express a drug-resistance marker (neomycin) and a swine class II DRB cDNA were constructed. Infectious particles containing these vectors were produced at a titer of greater than 1 x 10(6) G418-resistant colony-forming units/ml using both ecotropic and amphotropic packaging cell lines. Flow cytometric analysis of DRA-transfected murine fibroblasts subsequently transduced with virus-containing supernatants demonstrated that the transferred sequences were sufficient to produce DR surface expression. Cocultivation of murine bone marrow with high-titer producer lines leads to the transduction of 40% of granulocyte/macrophage colony-forming units (CFU-GM) as determined by the frequency of colony formation under G418 selection. After nearly 5 weeks in long-term bone marrow culture, virus-exposed marrow still contained G418-resistant CFU-GM at a frequency of 25%. In addition, virtually all of the transduced and selected colonies contained DRB-specific transcripts. These results suggest that a significant proportion of very primitive myelopoietic precursor cells can be transduced with the DRB recombinant vector and that vector sequences are expressed in the differentiated progeny of these cells.

Animals↗
PubMed1991

A gene remnant in the rabbit MHC related to HLA-DR alpha.

The screening of a rabbit genomic library yielded 2 HLA-DR alpha-related clones. One of these was previously shown to contain a functional gene (RLA-DR alpha) closely related to HLA-DR alpha. The second DR-related clone, designated DF, was mapped to the rabbit MHC class II region by genetic studies. DF contains contiguous sequences that have significant homology to the RLA-DR alpha transmembrane-coding region (contained in exon 4 of RLA-DR alpha) and to part of intron 4. These 2 stretches were reversed in DF when compared to other class II genes, and were flanked on both sides by direct repeats of 130 bp with 85% sequence identity. The presence of the transmembrane sequence without the coding sequences that accompany it in DR alpha, and the observation of the repeats, suggests that the DF region played a role in evolution of class II genes. Genomic blots revealed sequences hybridizing to DF in all rabbits tested and in other lagomorph species. The ratio of replacement to silent substitutions in the DF transmembrane region differed significantly from functional class II genes suggesting an absence of recent selective pressure. An additional direct repeat of the C type, similar to that found in rabbit cytochrome p450, certain class I MHC, TcR beta and uteroglobin genes, was also present in the DF fragment.

Amino Acid Sequence↗
PubMed1990

Class II genes of miniature swine. IV. Characterization and expression of two allelic class II DQB cDNA clones.

Two cDNA clones coding for allelic miniature swine MHC class II Ag DQB chains have been isolated, characterized, and shown to be expressed after transfection into mouse fibroblasts. The two alleles differ at the nucleotide level by an overwhelming proportion of replacement substitutions, suggesting the influence of selection for polymorphism. Most of the resulting predicted amino acid replacements are in regions commonly polymorphic in mouse Ab and human DQB sequences, corresponding to the predicted Ag recognition site. Nucleotide and amino acid sequence comparisons to homologous mouse and human sequences show more similarity between swine and man than between either swine and mouse or man and mouse. This tendency is most pronounced when comparing the 3' untranslated regions. However, an examination of unique cross-species sharing of amino acid residues suggests a closer relationship between both man and miniature swine and man and mouse than between miniature swine and mouse. The simplest explanation we can envision for these findings is that the mouse DQB gene homologue (Ab) has been subject to a higher substitution rate than either swine or human DQB genes. An additional cytoplasmic exon expressed in mouse Ab gene products and in putative human DQB2 gene products is lacking in both swine and human DQB cDNA clones. Its absence suggests either that the expression of this exon in mouse Ab genes was activated after mammalian speciation or that the expression of this exon was independently inactivated in swine DQB and human DQB1 genes. Alternatively, the mouse Ab gene may be derived from the same primordial gene as human DQB2, whereas the pig DQB gene may be derived from the same primordial gene as the human DQB1 gene.

Alleles↗
PubMed1990

Structure of class II genes in wild mouse Mus saxicola: functional and evolutionary implications.

Class II beta genes have been characterized from Mus saxicola, a wild mouse which diverged five millions years ago from the house mouse. The nucleotide sequences of the exons 2 and 3 of E beta 1, A beta 2 and E beta 2 genes have been determined in order to define the molecular mechanisms of interspecies variability. The E beta 1 and A beta 1 (J. X. She, personal communication) genes from Mus saxicola display extensive sequence variability in exon 2 when compared to those of the house mouse, in association with the variability of residues probably involved in antigen binding. On the other hand, most putative T cell contact residues from the I-E beta chain, and to a lesser extent from the I-A beta chain, were found conserved in mice. T cell contact residues from Mus saxicola I-A beta and I-E beta chains are identical to those of the b and q, and of the k haplotypes of the house mouse, respectively. Comparisons performed with A beta 2 and E beta 2 genes revealed that both are highly conserved and, in contrast to A beta 1 and E beta 1 genes, that a strong selective pressure for conservation occurs in the A beta 2 second exon. The different aspects of A beta 2 and E beta 2 genes evolution may correlate with the original function of their products. Comparisons of the intervening sequences revealed that the repeated motifs of the hotspot region present in the E beta 1b gene are also conserved in Mus saxicola. Recombinations may have occurred, in some cases in association with chi-like sequences.

Amino Acid Sequence↗
PubMed1990

Class II genes of miniature swine. II. Molecular identification and characterization of B (beta) genes from the SLAc haplotype.

Genomic clones corresponding to class II beta genes of the SLAc haplotype of miniature swine have been isolated and characterized. These genes have been grouped into seven non-overlapping clusters on the basis of restriction mapping. Ordering of exons within each cluster was accomplished by hybridization of Southern blots of restriction fragments with exon-specific probes. The two clusters (clusters 2 and 3) encoding the DRB and DQB genes were identified on the basis of hybridization with locus-specific 3' untranslated cDNA probes. Cluster 4 contained exons of both DOB and DQB genes, the basis for which remains to be determined. The remaining four clusters (1, 5, 6, 7) were identified as containing DP, DR, and DO coding sequences, respectively, on the basis of sequence analysis. The porcine class II region appears very similar to that of man in number and nature of the class II genes identified and in the intron/exon organization of corresponding genes.

Animals↗
PubMed1990

Structure of miniature swine class II DRB genes: conservation of hypervariable amino acid residues between distantly related mammalian species.

As part of our studies of the class II genes of miniature swine, we have isolated and characterized cDNA clones corresponding to DRB genes from two major histocompatibility complex homozygous strains. Comparison of the sequences of these clones to those of human DRB genes revealed a striking amino acid homology between the hypervariable residues of SLA-DRBc and the human DRB1-0101 allele. The percentage of differences in these residues between the pig DRBc allele and the human DRB1-0101 allele was significantly lower (29%) than that between the DRB1-0101 allele and all other human alleles (average, 66.2%). This similarity was not seen in a comparison of the number of silent substitutions, by which the swine DRBc and the human DRB-0101 differed. Since phenotypic selection operates at the level of protein products rather than nucleotide sequences, these data suggest the existence of selective mechanisms that have resulted in similar hypervariable regions in certain alleles even in these widely disparate species. Consistent with this hypothesis, an examination of available murine and bovine class II sequences revealed a homology in hypervariable residues between the human DRB1-1401 allele and the mouse E beta s allele as well as a cow DRB allele. Consideration of these data along with intraspecies allelic sequence comparisons suggests that at least some of the interspecies similarities have emerged as the result of convergent evolution, possibly as the result of a need to react to common pathogens.

Amino Acid Sequence↗
PubMed1987

Expression patterns of MHC class II genes in rabbit tissues indicate close homology to human counterparts.

Genomic clones corresponding to five distinct major histocompatibility complex class II alpha-chains have been described for the rabbit; four of these encode complete, potentially functional alpha-chains. Hybridization analysis and preliminary sequence analysis indicate that one of these clones is structurally related to HLA-DP alpha, one to -DR alpha, one to -DQ alpha and one to -DZ alpha. Probes specific for the four class II genes were used to screen RNA samples from normal rabbit tissues to determine which of these genes are transcribed and whether expression of any particular gene is tissue specific. All four of the genes are transcribed, but there are variations in the levels of expression, in tissue distribution, and in transcript size. The highest levels of RLA-DR alpha, -DQ alpha, and -DP alpha transcription were found in lymphoid tissues. Lower levels of transcription were also detectable in several nonlymphoid tissues. Transcripts observed were about 1.3 kb, a size expected for these class II alpha-chain genes based on experience with their human homologues. The RLA-DZ alpha probe corresponding to HLA-DZ alpha hybridized weakly with a band of 3.6 kb; its expression could be detected only in lymphoid tissues. The size of the DZ alpha transcript, its tissue distribution, and partial sequence data confirm its homology with the human gene DZ alpha. In blots of total cellular RNA, a probe for a recently described human beta-chain, DO beta, hybridized to a transcript of about 1.3 kb in lymphoid tissues. These data indicate that RNA transcripts corresponding to all HLA class II loci described to date can be detected in rabbit tissues.

Animals↗
PubMed1987

Sequence determination of a transcribed rabbit class II gene with homology to HLA-DQ alpha.

The human HLA-DQ alpha probe was used to screen genomic and cDNA libraries constructed from a rabbit T-cell line. Clones containing highly homologous sequences were obtained from both libraries and their sequences were determined. The organization of the RLA-DQ gene was determined by comparison of the nucleotide sequences of the genomic clone to that of the corresponding cDNA clone. This analysis allowed assignment of the complete structure of the RLA-DQ alpha chain. Comparisons with human and mouse class II products revealed that RLA-DQ is more closely related to HLA-DQ/DX than to H-2A. In contrast to the DQ/DX region of man, which contains at least two distinct alpha genes, the rabbit genome contains a single DQ gene which is equally distant from the HLA-DQ or -DX genes. The rabbit DQ alpha gene, like human HLA-DQ, is transcribed in T cells.

Amino Acid Sequence↗
PubMed1985

Molecular evidence for five distinct MHC class II alpha genes in the rabbit.

Human HLA class II gene probes were used to identify five distinct genes encoding the class II heavy chain (alpha chain) in the rabbit. The rabbit genes were defined by both mapping data and hybridization studies of genomic clones derived from the inbred B/J rabbit strain. Analysis of the clones by hybridization at graded stringencies indicated that one group of clones corresponded to HLA-DR, one group to HLA-DQ, and two groups to HLA-DP. Clones within a fifth group, designated DN, hybridized weakly to HLA-DR and may carry a fourth species of class II alpha genes in the rabbit. Clones within the group showing high homology to HLA-DR alpha were found to also contain sequences hybridizing with a probe for HLA-DR beta. No HLA-DP, -DQ, or -DR beta sequences were detected in any of the other class II alpha clones. Distinct banding patterns observed in Southern blot analyses using either human or rabbit class II alpha probes revealed restriction fragment length polymorphism for the different rabbit haplotypes studied. The DN, DQ, and DR alpha genes appear to be present as single copies whereas there are two distinct DP-like genes in the rabbit.

Animals↗