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J L Strominger

Publications and source records attributed to J L Strominger.

At least 289 records · Page 16Linked to original sources
PubMed1986

Structure and evolution of the HLA class II SX beta gene.

The class II major histocompatibility complex antigens are cell-surface heterodimers consisting of an alpha and a beta chain. Cosmid cloning has shown that the three families of class II antigens, DR, DQ, and DP, are encoded within the HLA-D region of chromosome 6 as a series of discrete gene clusters. The DP cluster contains two pairs of alpha and beta genes, one of which encodes the biochemically-defined DP antigen. In order to assess whether the other two genes, SX alpha and SX beta, are also expressed, potential coding regions have been subcloned and sequenced. The SX3 beta gene is shown to contain regions closely homologous to all six exons of DP beta. A 1 bp deletion in the beta 2 exon, also observed for the SX4 beta allele, causes a translational frameshift, suggesting that SX beta is a pseudogene. However, all the other exons, as well as their splice sites and the putative promoter region, appear to be intact.

Alleles↗
PubMed1986

The complete primary structure of the T-cell receptor genes from an alloreactive cytotoxic human T-lymphocyte clone.

The complete primary structure of the cDNAs encoding the alpha and beta chains of the T-lymphocyte receptor for antigen from a human alloreactive, cytotoxic T-cell clone, L17, is presented. Sequence analysis of these genes reveals that both are related to immunoglobulins and are composed of variable, diversity (at least in the case of the Ti beta clone), joining, and constant region sequences. Comparison of the sequence of the alpha-chain cDNA to that of previously sequenced mouse and human alpha cDNAs suggests the presence of human T-cell receptor alpha D-region sequences. Southern blot analysis confirms the finding that these cDNAs represent the functional receptor genes expressed by the L17 cytotoxic T-cell clone. The availability of these full-length T-cell receptor cDNA clones from a human T-lymphocyte clone of known antigen specificity should allow an analysis of the relationship between T-cell receptor structure and function.

Amino Acid Sequence↗
PubMed1986

Immunochemically purified DR antigens in liposomes stimulate xenogeneic cytolytic T cells in secondary in vitro cultures.

A monoclonal antibody directed against the human class II major histocompatibility antigen DR was generated. Use of this antibody, LB3.1, allowed isolation of large amounts of highly purified DR by immunoaffinity chromatography. The DR was reconstituted into liposomes and shown to stimulate secondary xenogeneic cytolytic T lymphocytes (CTL) specific for targets expressing DR antigens. DR digested with neuraminidase was equally as effective as native DR at stimulating CTL, while denatured DR and other purified membrane proteins were much less effective. The DR liposome-induced CTL lysed only target cells expressing class II antigens. Cytolysis of targets bearing class II antigens was blocked by DR-specific antisera.

Animals↗
PubMed1986

A monoclonal antibody that recognizes the alpha chain of HLA-DR antigens.

A monoclonal antibody, HC2.1, has been generated that specifically reacts with both the denatured and the in vitro translated alpha chain of the DR antigen. Although HC2.1 antibody reacted with the alpha chain of protein immunoprecipitated by two DR-specific monoclonal antibodies, L227 and LB3.1, it did not react with the alpha chain of the DQ1 antigen immunoprecipitated by the monoclonal antibody, Genox 3.53. The isoelectric focusing pattern of the alpha chain precipitated by HC2.1 antibody was invariant across a range of DR specificities within a panel of lymphoblastoid cells. The alpha chain of DR antigen from a B cell line was purified by HC2.1-Sepharose immunoaffinity chromatography and limited amino acid sequence analysis was carried out with Staphylococcus aureus SV8 protease fragments purified by high-pressure liquid chromatography. The sequence analysis confirmed that the antigen reactive with HD2.1 antibody is encoded by the DR alpha chain gene.

Amino Acid Sequence↗
PubMed1986

DNA restriction fragment length polymorphism of HLA-DR2: correlation with HLA-DR2-associated functions.

HLA-DR2 can be divided into at least 3 distinct HLA-D clusters which correlate with structural differences within the HLA-D region. Further, a functional counterpart of this subdivision has been previously identified. The presence of a particular DR beta 2 polypeptide chain correlated precisely with the susceptibility of measles virus-infected HLA-DR2 homozygous typing cell lines to lysis by measles virus-specific, HLA class II-restricted CTL clones. To determine if a genetic basis for these functional differences could be detected, the degree of polymorphism at the DNA level within the serologically defined HLA-DR2 haplotype has been examined. By using DNA probes for DR beta and DQ beta 4 of the 5 HLA-D clusters of HLA-DR2 could be distinguished and a RFLP pattern was identified which correlates with known immunological functions associated with these various D types. In addition, this technique of 'molecular genotyping' was used to investigate a limited panel of patients with multiple sclerosis (MS) who were HLA typed as either HLA-DR2,2 or HLA-DR2,blank. The RFLP profile of the HLA-DR2 Dw2 D type was found in all of these MS patients.

Cell Line↗
PubMed1986

Generation of diversity of the beta chain of the human T-lymphocyte receptor for antigen.

Human T-cell receptor beta-chain (Ti beta) genes are formed by the rearrangement of variable (V), diversity (D), and joining (J) gene segments. A comparison of the nucleotide and deduced amino acid sequences of the variable regions of six human Ti beta cDNAs reveals that they display a level of homology similar to that shared by human immunoglobulin heavy chain V genes. In contrast to immunoglobulin V regions, which contain three discrete regions of hypervariability, the Ti beta V regions display a more widely distributed pattern of variability. Southern blot analyses show that most human Ti V beta gene families contain one to three members. However, a single family containing at least eight members is identified. This analysis allows the identification of at least 15 human Ti V beta germ-line genes. The sequence data show that at least one germ-line Ti beta J gene is used preferentially in Ti beta cDNAs. Moreover, they suggest the presence of at least four human germ-line Ti beta D genes. At least three mechanisms are involved in generating the diversity of human Ti beta genes: (i) the combinatorial rearrangement of different V, D, and J genes; (ii) imprecise V-D-J joining, including V-D joining in any of three translational reading frames; and (iii) the addition of extra nucleotides at the V-D-J joints (N-region diversity).

Base Sequence↗
PubMed1986

Recombinant human tumor necrosis factor increases mRNA levels and surface expression of HLA-A,B antigens in vascular endothelial cells and dermal fibroblasts in vitro.

Recombinant human tumor necrosis factor (TNF), purified to greater than 99% homogeneity, increases surface expression of class I major histocompatibility complex (MHC) antigens to a maximum of 9-fold on cultured human endothelial cells (HEC) and human dermal fibroblasts (HDF). The increase is concentration dependent (peak 20-100 units/ml) and time dependent (nearly maximal by 4 days); expression remains elevated in the continued presence of TNF and requires greater than 7 days to return to basal levels upon TNF withdrawal. The increase in surface expression appears to result from increases in steady-state mRNA levels for the class I antigens, although the increase in mRNA is proportionately greater than for surface expression. No surface expression of or mRNA for class II MHC antigens is detectable in either control or TNF-treated HEC or HDF. These effects are similar to those produced by leukocyte or fibroblast (type I) interferons (IFNs). The protein synthesis inhibitor cycloheximide (CHX), when added coincidentally with type I IFNs, leads to superinduction of mRNA for class I MHC antigens and, unexpectedly, leads to the appearance of mRNA for class II MHC antigens. CHX has no effect by itself upon mRNA levels for class I or class II MHC antigens, nor does it modulate the increases in mRNA produced by immune (type II) IFN. Most interesting, CHX blocks the increase in mRNA for class I MHC antigens induced by TNF. Thus TNF appears to act on MHC gene expression through a newly synthesized protein intermediate. Our results provide direct evidence that TNF can modulate gene expression in normal (untransformed) cell types and contribute to understanding the complex nature of MHC gene regulation. Finally, they suggest that TNF may act in vivo as an immunoregulatory molecule.

Cell Membrane↗
PubMed1986

Genes for the tumor necrosis factors alpha and beta are linked to the human major histocompatibility complex.

The human major histocompatibility complex (MHC) includes the closely linked genes for the tumor necrosis factors alpha and beta. Their location is within the chromosomal segment between HLA-DR and HLA-A or centromeric of HLA-DP. This assignment is based on Southern blot analysis of a number of different MHC deletion mutants and is corroborated by chromosome in situ hybridization.

Chromosome Deletion↗
PubMed1986

Regulation of a transfected human class II major histocompatibility complex gene in human fibroblasts.

To investigate the cis-acting DNA elements that are involved in regulation of class II major histocompatibility complex genes, including gamma-interferon (gamma-IFN) induction, 5' flanking DNA deletions of a DQ beta "minigene" were analyzed in stable transfected cell lines. At least four elements 5' to the gene were found to be involved in DQ beta regulation. Deletion of sequences from -2500 to -159 base pairs (bp) resulted in increased transcription, suggesting that negative regulatory elements resided in the deleted region. These clones were all capable of responding to gamma-IFN. Further deletion of sequences from -159 to -128 bp resulted in constitutive high level transcription and the inability of these constructions to respond to gamma-IFN. A deletion to -107 bp resulted in a decrease in the basal level of expression that was restored by removal of the 5' DNA sequence to -82 bp, suggesting the presence of a second negative element. Finally, deletion to -64 bp caused a marked decrease in expression, suggesting the loss of an element necessary for high levels of transcription. The gamma-IFN control and the transcription control elements contain the conserved upstream sequences found in all class II genes, suggesting a role for these sequences.

Fibroblasts↗
PubMed1986

An Epstein-Barr virus transcript from a latently infected, growth-transformed B-cell line encodes a highly repetitive polypeptide.

By screening a cDNA library prepared from poly(A)+ RNA isolated from an Epstein-Barr virus latently infected, growth-transformed human B-lymphoblastoid cell line, we have recovered a clone corresponding to a highly spliced viral transcript encoded largely by the major internal repeat (IR1). The 5' region contains one copy of a 26-base-pair (bp) exon (W0) [which is 28 bp downstream from a CAATT-(N)34TATAAA sequence (N, unspecified base)] and seven copies of two small exons (W1, 66 bp; W2, 132 bp). In addition, there are three exons from the "unique" region of the BamHI Y fragment of the viral genome. Two other cDNA clones that have been described, corresponding to latent viral transcripts, share homology in their 5' regions with this clone and are clearly divergent at their 3' ends. The cDNA clone described in this paper contains one long open reading frame that extends through the repeat element. In vitro transcription and translation of this open reading frame yielded a 62-kDa polypeptide that could be immunoprecipitated by an Epstein-Barr virus-positive human serum.

B-Lymphocytes↗
PubMed1986

Cloning and sequence analysis of complementary DNA encoding an aberrantly rearranged human T-cell gamma chain.

Complementary DNA (cDNA) encoding a human T-cell gamma chain has been cloned and sequenced. At the junction of the variable and joining regions, there is an apparent deletion of two nucleotides in the human cDNA sequence relative to the murine gamma-chain cDNA sequence, resulting simultaneously in the generation of an in-frame stop codon and in a translational frameshift. For this reason, the sequence presented here encodes an aberrantly rearranged human T-cell gamma chain. There are several surprising differences between the deduced human and murine gamma-chain amino acid sequences. These include poor homology in the variable region, poor homology in a discrete segment of the constant region precisely bounded by the expected junctions of exon CII, and the presence in the human sequence of five potential sites for N-linked glycosylation.

Amino Acid Sequence↗
PubMed1986

Rearrangement and expression of T-cell antigen receptor genes in human T-lymphocyte tumor lines and normal human T-cell clones: evidence for allelic exclusion of Ti beta gene expression and preferential use of a J beta 2 gene segment.

The gene encoding the beta chain of the human T-cell receptor for antigen is composed of variable (V), diversity (D), joining (J), and constant (C) gene segments which undergo specific rearrangements during T-lymphocyte ontogeny. Southern blot analyses of seven human T-cell tumor lines and normal human T-lymphocyte clones revealed that most of these T-cell lines rearrange their Ti beta genes differently. The T-cell tumor line HPB-MLT rearranges and transcribes both of its Ti beta genes. Cloning and sequencing of the Ti beta cDNAs corresponding to these rearrangements revealed that one of the rearranged Ti beta genes is defective, while the other is functional and corresponds to the Ti beta protein expressed on the surface of these cells. Thus, this cell line displays a pattern of allelic exclusion of Ti beta gene expression. A comparison of four C beta 2-containing Ti beta cDNAs from three different cell lines revealed that three of the four utilize the same J beta 2.5 gene segment joined to different D beta and V beta genes, suggesting that there may be preferential use of this J gene during J beta 2 rearrangements. Hybridization analyses with probes for the alpha and beta genes of the T-cell receptor and the T-cell-specific T gamma gene revealed that HPB-MLT cells appear to express approximately equivalent amounts of RNA corresponding to each of the rearranged Ti alpha and Ti beta genes. However, they express a much lower level of T gamma RNA.

Alleles↗
PubMed1985

Biochemical characterization of VLA-1 and VLA-2. Cell surface heterodimers on activated T cells.

The very late antigen complexes VLA-1 and VLA-2 which appear on long-term activated human T cells have been characterized with respect to 1) subunit arrangement, 2) location of monoclonal antibody (MAb) binding sites, 3) carbohydrate content, and 4) protein homology. Cross-linking experiments showed that the VLA-1 complex is a heterodimer composed of an Mr 210,000 subunit (alpha 1) in acid-labile association with an Mr 130,000 subunit (beta). The VLA-2 complex is a heterodimer with an Mr 165,000 subunit (alpha 2) in base-labile association with the Mr 130,000 beta subunit. The subunits of VLA-1 (alpha 1 beta) and VLA-2 (alpha 2 beta) each appear to be arranged with 1:1 stoichiometry. The MAb A-1A5 has been shown to bind to an epitope on the common beta subunit, consistent with its recognition of both the VLA-1 and VLA-2 heterodimers. On the other hand, MAb TS2/7 bound to an epitope of the alpha 1 subunit, thus explaining the specific recognition of the VLA-1 heterodimer by TS2/7. Digestion of the alpha 1, alpha 2, and beta subunits with neuraminidase and with endoglycosidase F revealed that each subunit contains substantial sialic acid and N-linked carbohydrate. By one-dimensional peptide mapping, the alpha 1, alpha 2, and beta subunits were shown to be highly nonhomologous with respect to each other, although each subunit from different T cell sources appeared highly homologous if not identical.

Antibodies, Monoclonal↗