Showing papers by "Richard A. Flavell published in 1984"

Organization and evolution of the class I gene family in the major histocompatibility complex of the C57BL/10 mouse

Abstract: The major histocompatibility complex (MHC) encodes several classes of protein vital to the regulation of the immune response. We have isolated 26 class I genes that map to this region in the C57BL/10 mouse and linked these into three gene clusters. The number of genes differs from the number found in the BALB/c strain and comparison of the organization of the class I genes in these two strains shows conserved regions and polymorphic regions which probably result from deletions, insertions and translocations within the MHC.

Domain interactions of H–2 class I antigens alter cytotoxic T-cell recognition sites

Abstract: H–2 class I antigens appear to direct the recognition of virus-infected and neoplastic transformed cells by cytotoxic T lymphocytes (CTLs)1,2. Here, to identify the regions of class I antigens involved in CTL recognition, four hybrid class I genes were constructed in which exons were exchanged between the H–2Kband H–2Db genes3,4. These class I genes were expressed in mouse L cells and recognition of the hybrid Kb/Dbantigens by CTLs and monoclonal antibodies specific for either Kb or Db was investigated4. The pattern of CTL and monoclonal antibody recognition obtained indicates three correlations between structure and function of class I antigens. First, most CTL recognition sites and alloantigenic determinants are located on domains 1 and 2 of the antigen molecule. Second, these CTL recognition sites and alloantigenic determinants are not influenced by interaction of domains 1 and 2 with polymorphic regions of domain 3. Third, in contrast, interaction between domains 1 and 2 alters these CTL recognition sites and alloantigenic determinants. The alteration of CTL recognition sites by interaction between domains 1 and 2 suggests that a CTL site may be formed by amino acids from both domains 1 and 2, or that the conformation of amino acids at a CTL site may be altered by interactions between domains 1 and 2. Through these two features, the conformation of CTL recognition sites on H–2 class I antigens may be sensitive to alteration by interaction of either domain 1 or 2 with viral antigens.

Regulated expression of an introduced MHC H-2K bm1 gene in murine embryonal carcinoma cells.

Abstract: The transplantation antigens H-2K, H-2D and H-2L are developmentally regulated, highly polymorphic cell surface proteins encoded by the major histocompatibility gene complex (MHC). First detectable on the early embryo, they are subsequently expressed on most somatic cells of the adult mouse in association with the protein beta2-microglobulin (beta 2 M; ref. 5). Cultured F embryonal carcinoma (EC) cells can be induced to differentiate along alternative pathways to form either parietal or visceral9 extra-embryonic endoderm, each concomitant with a change in morphology and pattern of gene expression. Previous reports have demonstrated an increased level of transplantation antigens in differentiated F9 EC cells, but the cell types expressing them were not defined. Here we show that the level of MHC H-2Kb and beta 2 M transcripts is increased during both pathways of this differentiation. Expression of a foreign MHC H-2Kbm1 gene was found to be regulated in a similar manner when the gene was introduced into EC cells. In contrast, an introduced rabbit beta-globin gene was not regulated but expressed constitutively.

DNA Sequences Required for Regulated Expression of $\beta$ -Globin Genes in Murine Erythroleukaemia Cells

Abstract: We have introduced into murine erythroleukaemia (MEL) cells a series of human globin gene cosmids and two sets of hybrid genes constructed from the human beta-globin gene and the human gamma-globin or murine H-2Kbm1 genes. S1-nuclease analysis of the mRNA products from these genes before and after MEL cell differentiation showed that the human beta-globin gene, but not the human epsilon- or gamma-globin or H-2Kbm1 genes, is induced specifically. Hybrid genes containing human beta-globin DNA sequences from either 5' or 3' side of the translation initiation site were both inducible. Measurement of the relative rate of transcription showed this induction to be the result of transcriptional activation. We therefore suggest that DNA sequences which regulate beta-globin gene expression during MEL differentiation are located both 5' and 3' to the translation initiation site.

Murine leukemia virus sequences are encoded in the murine major histocompatibility complex

Abstract: The studies reported here localize murine leukemia viral sequences to the TL region of the major histocompatibility complex, H-2. We examined a battery of 38 cosmids, isolated from two large genomic libraries constructed from C57BL/10 spleen DNA, that define 25 class I gene sequences. The viral probes used hybridized with only four cosmids, containing overlapping mouse sequences, that define four class I gene-related sequences in a region of 90 kilobases of DNA. The data show that two distinct viral envelope sequences are contained in the cluster. One of these sequences is situated with its 3' end next to the 3' end of a class I sequence. The other sequence, which does not contain the entire viral envelope, is proximal to the 3' end of a different class I sequence. Hybridization of the viral probes with the H-2 cosmid clones does not appear to be due to homology between viral and H-2 sequences. Rather, the viral sequences detected appear to be linked to or inserted amid class I genes. These findings may be significant in understanding molecular mechanisms involved in the generation of H-2 class I gene diversity.

A new determinant, Qa-m208, detected on T lymphocytes and transfected L cells by a Kb-specific monoclonal antibody.

Abstract: A Kb-specific monoclonal antibody, 6.3.4, defining a new class I specificity, m208, reacts with some K and D region products and with a Qa determinant present on T lymphocytes but not detectable on thymocytes and B lymphocytes. The strain distribution of reactions indicates that this determinant is controlled by the Qa-Tla region, and shows no concordance with the strain distribution pattern of any of the known Qa antigens. The antibody reacts also with L cells expressing a cloned H-2b class I gene mapping in the Qa-Tla region.

Molecular Cloning of H-2 Class I Genes in the H-2b Haplotype

Abstract: We have cloned H-2 class I genes of the C57BL/10 (B10, H-2b haplotype) mouse using cosmid cloning techniques developed in our laboratory (1). Cosmids, which contain 40–45 kilobasepairs (kbp) of mouse DNA per clone, were chosen since there are about 20–30 class I genes (see below) in the mouse genome and most of these genes map between the H-2K and TL loci on chromosome 17 (Figure 1); a region of some 1000 kbp. of DNA. Estimates of the number of class I genes in the mouse genome are based on Southern blot analysis (2) in which restriction enzyme digested mouse genomic DNA is hybridised to a class I H-2 cDNA probe (Figure 2). In all digests shown, and in all three H-2 haplotypes tested, the cDNA probe hybridises to multiple bands in the digests indicating that the H-2 class I are multiple copy and highly homologous. Using either H-2 cDNA or H-LA genomic probes we have isolated 90 cosmids containing class I genes from a mouse genomic library constructed using B10 spleen DNA. These cosmids have been organised into 5 distinct clusters of overlapping mouse DNA on the basis of restriction enzyme mapping and Southern blot hybridisation analysis (Table 1). The DNA regions cloned in each cosmid cluster have been mapped to one of the four genetic loci known, from immunological analysis, to control the expression of class I cell surface antigens in the B10 mouse (see Figure 1).