Correlation of CT beta phenotype with origins of laboratory mouse strains.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
T J Kindt
Publications and source records attributed to T J Kindt.
Molecular genotyping of human T-cell antigen receptor variable gene segments.
The gene complex encoding the beta chain of the T-cell antigen receptor (Tcr) in man was previously reported to contain a restriction fragment length polymorphism (RFLP) involving a single Bgl II site adjacent to the second constant region gene. This RFLP allowed assignment of Tcr beta genotypes in certain human families. In the present study, two different RFLP in a V beta gene family were detected using the murine probe V8.1 in genomic DNA samples digested with the restriction endonucleases Hind III and Bam HI. Use of these RFLP to mark the V beta gene complex allowed complete haplotype assignment in four of seven families studied and provided support for linkage of the V gene complex to the constant region genes. Different combinations of the C and two V region markers can result in eight possible distinct haplotypes. The observation of all but one of the eight possible haplotypes in parents of the families studied suggests that recombination events occur between the C and V region and among members of the V region subfamily marked by the V8.1 probe. These markers can be used for mapping studies of the V beta gene complex in man and will allow an appraisal of possible associations between Tcr beta genes and disease susceptibility.
DNA↗
Monoclonal antibodies directed against major histocompatibility complex antigens bind to the surface of Treponema pallidum isolated from infected rabbits or humans.
Evidence is presented for the association of class I major histocompatibility complex (MHC) antigens with the surface of Treponema pallidum during infection. A monoclonal antibody (IgG2a) directed against a murine H-2Kb epitope of public specificity reacted with the cell surface of T. pallidum, as assayed by the binding of protein A-colloidal gold in immunoelectron microscopy. Monoclonal antibodies directed against class I rabbit MHC antigens also reacted in immunofluorescence assays with material on the surface of rabbit-cultivated T. pallidum. In addition, impression smears of human syphilitic genital ulcers that were darkfield-positive for the presence of spirochetes were tested in immunofluorescence assays with monoclonal antibodies directed against human MHC antigens; antibody directed against HLA-ABC (class I) was reactive whereas antibody directed against HLA-DR (class II) was nonreactive. Results of the study suggest that the association of host-derived class I MHC antigens or molecular mimicry may play a role in T. pallidum evasion of host immune defenses.
Animals↗
Two distinct T-cell receptor alpha-chain transcripts in a rabbit T-cell line: implications for allelic exclusion in T cells.
Information relevant to allelic exclusion in T cells has been obtained by a study of cDNA clones corresponding to alpha-chain genes of the T-cell receptor in the rabbit T-cell line RL-5. One clone contains a variable-joining-constant (VJC) sequence encoding a complete alpha chain of the T-cell receptor. A second has an identical constant region and includes a distinct variable-joining (VJ) sequence. However, a single-base deletion in the variable region places the remainder of the second transcript out-of-phase and appears to be the product of a rearrangement involving a variable region of the T-cell receptor alpha-chain pseudogene. Presence of two variable-joining-constant (VJC) transcripts in the same cell line indicates that alpha-chain gene rearrangement is not affected by transcription of a complete alpha-chain mRNA and suggests that steps after mRNA synthesis are involved in the allelic exclusion process for alpha-chain genes. Comparison of rabbit alpha-chain sequences with those of man and mouse revealed interspecies conservation in constant and variable regions. Genomic Southern blot analyses using a rabbit constant region of the T-cell receptor alpha-chain probe revealed the presence of a single constant region gene. Hybridization with variable region probes defined two distinct multigenic subfamilies. Homology between certain rabbit and murine variable regions of the T-cell receptor alpha-chain sequences suggests that the existence of subfamilies predated divergence of these species.
Alleles↗
Lambda light chain constant and variable gene complements in wild-derived inbred mouse strains.
Previous investigations of lambda light (L) chains in wild-derived inbred (WDI) mouse strains revealed large variations in serum levels of this L chain type as well as differences in antibody responses in which lambda chains predominate. In the present study a diverse group of WDI strains was analyzed by Southern blot analysis using probes for V lambda, C lambda 1 and C lambda 2 genes in an attempt to correlate the complement of lambda genes present in these strains with their lambda expression and with their taxonomic classification. All strains studied had two or three DNA sequences that strongly hybridized with a V lambda probe derived from BALB/c; most strains had V lambda sequences on restriction fragments similar in size to V lambda 1 and V lambda 2 sequences from BALB/c. Comparison of C lambda 1 and C lambda 2 genes in WDI mice to those of BALB/c revealed extensive variation both in number of hybridizing fragments and in their sizes. There were no obvious correlations observed between C lambda and V lambda patterns obtained for mouse strains of any phylogenetic group suggesting that constant and variable regions of lambda have evolved independently. In contrast to variations found for C lambda, a single kappa constant region gene appears to be conserved throughout the various mouse species.
Animals↗
Localization of C4 genes within the HLA complex by molecular genotyping.
Segregation of the complement component, C4, was analyzed in six families that each included an individual who inherited an HLA haplotype where a crossover event had occurred in the region between HLA-B and HLA-DR. Two cDNA clones corresponding to the C4 gene were utilized as probes in Southern blot analysis of DNA from members of each family. Restriction fragment length polymorphisms (RFLP) were observed and were assigned to haplotypes. In one family RFLP, hybridizing with the C4 probes, segregrated with HLA-B, and in four families RFLP segregated with HLA-DR; one family was not informative in this respect. These analyses have made it possible to localize the genes for C4 between HLA-B and HLA-DR by molecular genotyping and to characterize three different genomic configurations of C4 genes by limited restriction mapping.
Chromosome Mapping↗
Structure of a functional rabbit class I MHC gene: similarity to human class I genes.
Studies of rabbit major histocompatibility complex proteins have suggested that rabbits express only a single class I antigen, in contrast to most mouse strains (H-2K, D, and L) and man (HLA-A, -B, and -C), which express three. To explore the significance and the molecular basis of this apparent species difference, we have characterized the expressed class I protein and a corresponding cDNA clone from the rabbit cell line RL-5, which is derived from the inbred B/J rabbit strain. As an extension of these analyses, this report documents the genomic sequence of a gene, designated 19-1, which encodes the same histocompatibility antigen expressed in RL-5. The availability of the corresponding full-length cDNA and amino-terminal protein sequence indicates the fully functional nature of the 19-1 gene and allows presumptive assignment of the transcription start site and delineation of exon/intron boundaries. Comparisons of the rabbit gene with homologous human and mouse sequences reveal a striking similarity between 19-1 and human genes in both exon/intron organization and specific nucleotide sequences; this close similarity allows tentative identification of previously unrecognized transcriptional start sites in the human genes.
Animals↗
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↗
Rabbit-mouse hybridomas secreting intact rabbit immunoglobulin.
Rabbit-mouse hybridomas offer the potential for production of monoclonal rabbit antibodies by immortal cell lines. In previous studies, it was possible to produce and stabilize rabbit-mouse hybrid cells secreting either a rabbit heavy or light chain. These have been useful for structural characterization of the individual rabbit immunoglobulin polypeptides and for isolation of large amounts of immunoglobulin mRNA for molecular studies. For some studies, however, it would be useful to have intact rabbit immunoglobulin molecules comparable to the myeloma proteins available in the human and mouse. The availability of rapid, sensitive and specific assays for rabbit heavy and light chains and allotypes located on specific chains has now permitted the early identification of clones secreting intact rabbit immunoglobulin. Vigorous cloning efforts have resulted in isolation and partial stabilization of three such clones. The first, H105, secretes a product with a kappa light chain bearing the b6 allotype and a mu-chain bearing the a1 allotype. Biochemical and serologic analyses of the product show that it is secreted as a fully assembled IgM pentamer and that the rabbit heavy and light chains are covalently associated. No rabbit J-chain gene was detected in H105 by Southern blot analysis. The second hydridoma, H134, secretes a product with a mol. wt of 150 K, consisting of a b4 light chain and an a1 heavy chain. The third, H171, secretes an alb4 IgG with antibody specificity for group C streptococcal carbohydrate. An additional rabbit-mouse hybridoma, H89, have been produced which secretes a rabbit heavy chain lacking group a allotypic activity. The rabbit heavy chain, which is associated with a mouse light chain, has an N-terminal amino acid sequence identical to a2-positive molecules although thorough serologic analysis revealed no group a allotypic activity.
Amino Acid Sequence↗
Molecular determination of T-cell receptor alpha and beta chain genotypes in human families.
Polymorphism in genes encoding the alpha and beta chain of the human T cell receptor has been detected by Southern blot analysis. Genomic DNA samples were isolated from B lymphoblastoid cell lines derived from members of families, each family including at least one individual with a recombinant HLA haplotype. T cell receptor alpha and beta chain haplotypes could be assigned in the families on the basis of observed restriction fragment length polymorphism (RFLP). Polymorphism in the alpha chain gene was detected in BglII digests using an alpha chain probe that included the V, J, C, and 3' untranslated sequences. A probe consisting of only the constant region (C alpha) revealed no polymorphism indicating that the polymorphic fragment hybridized to V, J, or 3' untranslated sequences of the alpha chain. Polymorphism in beta chain genes was observed in BglII digested DNA samples using a probe that corresponds to the constant region (C beta). Polymorphic C beta restriction fragments of 10.0 and 9.2 kilobase segregated in six of the eight families studied. Recent structural data for the C beta region suggest that the polymorphic BglII site lies in the region 5' to the C beta 2 gene. These polymorphisms should serve as markers for alpha and beta chain complexes allowing genetic studies of these immunologically important gene families.
Cloning, Molecular↗
Cellular distribution of a human I-A-like (DS/DC) antigen on normal and neoplastic lymphoid cells.
Previous biochemical studies have shown that B cell specific, monoclonal antibody, McAb 33.1, reacts with a class II antigen that represents a human analogue of murine I-A (DS/DC) antigens (J. Exp. Med. 158:1924, 1983). McAb 33.1 recognizes a polymorphic human B lymphocyte specific antigen present on mu+, B1+ peripheral B cells, B lymphoid cell lines, activated B cells, and neoplastic B lymphoid cells. Of 100 HLA-D/DR typed EBV-transformed lymphoblastoid cell lines tested, only those from DR3,3 and DR7,7 individuals failed to react with McAb 33.1. The 33.1 antigen is present at lower concentrations on B cells from blood, tonsil, spleen, and lymph node when compared to B cell lines. By contrast, the antigen is not detectable on blood T lymphocytes, T cell lines, or mitogen activated T cells and it is absent on monocytes of some individuals or is present only on a minor subpopulation (approximately 20%). McAb 33.1 should facilitate the functional, structure, and molecular dissection of the human Ia system.
Animals↗
Segregation of polymorphic T-cell receptor genes in human families.
Polymorphism in the genes encoding the constant (C) region of the beta chain of the T-cell antigen receptor (CT beta, also called C beta) has been detected by molecular genotyping analyses. In initial screenings, a panel of restriction endonucleases was used to digest DNA samples from two individuals; the digested samples were subjected to Southern blot analyses using a CT beta probe. The enzyme Bgl II revealed restriction-fragment-length polymorphism in these samples and was subsequently used to test 59 individual members of eight different families. Polymorphic fragments detected in six of the families could be used to follow the segregation of T-cell receptor genes; in many cases maternal and/or paternal haplotypes could be assigned. All members of two additional families displayed a single CT beta hybridizing fragment. In one family the DNA sample from one of the children lacked an expected Bgl II restriction fragment. On the basis of analyses with other restriction enzymes, the most likely explanation is that the lymphoblastoid B-cell line used as a source of genomic DNA for this individual had rearranged or altered CT beta genes. Restriction-fragment-length polymorphisms used to discriminate CT beta haplotypes in families provide useful markers that will facilitate linkage studies and genetic analyses of T-cell function.
DNA↗
Recombination within the HLA-D region. Correlation of molecular genotyping with functional data.
Molecular genotyping of the HLA-D/DR region in a family correlated with serologic and cellular typing data. It was further possible to predict a subtle difference in SB region-related functions from such molecular studies. A family that included an individual who inherited an HLA haplotype with a paternal recombination between HLA-B and the HLA-D/DR region was identified by classic HLA typing techniques. Segregation of HLA-D/DR region genes in this family was studied by Southern blot analysis using cDNA probes for DR alpha, DR beta, DC alpha, DC beta, and SB beta. Restriction enzyme fragment polymorphisms observed for every gene tested were in concordance with assigned HLA haplotypes (including the individual known to have inherited a paternal recombinant haplotype) with one exception: two HLA identical siblings were observed to have different SB beta restriction fragment patterns. Further testing revealed that one individual inherited a maternal HLA haplotype recombinant between the HLA-D/DR region and SB beta. Although both maternal SB alleles typed as SB4, allelic differences could be detected cellularly by primed lymphocytes and by the differential expression of a class II cell surface antigen using monoclonal antibody. Therefore, predicted and nonpredicted recombinant haplotypes were detected in a family by molecular genotyping.
DNA Restriction Enzymes↗
Specificity of the HLA class II reactive monoclonal antibody 33.1.
Explore the source record for details and available documents.
Antibodies, Monoclonal↗
Primary structural sequence polymorphism in the human class II MHC antigen 33.1.
Monoclonal antibody 33.1 defines a non-DR, class II, human major histocompatibility complex antigen, 33.1, which appears to be distinct from other class II antigens in its cellular distribution and primary structure. To characterize the structure more fully and to determine the degree of polymorphism within 33.1, a comparative N-terminal sequence study has been undertaken using a series of ten B lymphoblastoid cell lines with different DR and MB types. The results confirm that both the alpha and beta chains of 33.1 are homologues of the corresponding chains of the murine I-A antigen and indicate that while 33.1 does not appear to be identical with MB, it is closely related. Sequence analyses revealed two major variants of 33.1, corresponding to cells with specificities MB1 and MB3, respectively. Within each MB type, other polymorphisms have been detected. Cells that are MB2 do not react with monoclonal antibody 33.1. Suggestive evidence is presented that monoclonal antibody 33.1 reacts predominantly with the beta chain of the antigen. The preferential expression of 33.1 on activated B cells suggests that expression of at least the 33.1 beta chain gene is greatly enhanced in the course of B-cell activation, but the specific function of 33.1 remains to be determined.
Amino Acid Sequence↗
Cell surface glycoproteins of rabbit lymphocytes: characterization with monoclonal antibodies.
The structural characteristics of antigens recognized by a panel of monoclonal antibodies prepared against a rabbit T-lymphocyte cell line have been investigated. Those antigens which could be isolated using immunoadsorbents prepared from the monoclonal antibodies had mol. wts of 42,000, 90,000 and 120,000. The 42,000 mol. wt molecule is similar or identical to a rabbit class I major histocompatibility complex antigen and its characterization has been reported elsewhere. Three different 90,000 mol. wt proteins can be distinguished by their reactivity with lectins and by sequential immunoprecipitation. The 120,000 mol. wt protein is a very abundant surface glycoprotein that appears to be a specific marker for T-cells in the rabbit. It is the immunodominant antigen in a lentil lectin bound glycoprotein pool. Over half of the antibodies were directed against this antigen. All antigens detected by the panel of monoclonal antibodies have been detected on normal lymphoid cells.
Animals↗
A genomic gene encoding the b5 rabbit immunoglobulin kappa constant region: implications for latent allotype phenomenon.
We previously reported that domestic rabbits of five immunoglobulin kappa allotype strains (b4v, b4, b5, b6, and b9) harbor at least two DNA sequences that hybridize strongly to kappa constant region probes in Southern blots. One of these sequences ("type A") has been identified as encoding the constant region of the kappa 2 isotype, an immunoglobulin chain that most rabbits express only at low levels, if at all. We identified the second sequence--for rabbits of the b4 allotype--as encoding the nominal b4 kappa chain (or kappa 1 isotype), but for rabbits of other allotypes no definite identification for this "type B" sequence could be made. Here we suggest that the type B sequence in rabbits of the other domestic allotypes also encodes the nominal kappa 1 immunoglobulin chain. We show this directly for the b5 allotype; a type B sequence cloned from b5 DNA has been found to contain an apparently functional gene encoding the b5 constant region sequence. Indirect arguments suggest the corresponding conclusion for the b4v, b6, and b9 allotypes. We have considered the implications of these results for the phenomenon of "latent allotype" expression.
Animals↗