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J R Parnes

Publications and source records attributed to J R Parnes.

At least 73 records · Page 4Linked to original sources
PubMed1987

The gene encoding the mouse T cell differentiation antigen L3T4 is located on chromosome 6.

We have used Southern blot analysis of DNA from somatic cell hybrids to map the chromosomal location of the mouse L3T4 T cell differentiation antigen gene to chromosome 6. This finding is of interest because both L3T4 and the alternative T cell differentiation antigen Lyt-2 are homologous to kappa-immunoglobulin light chain-variable regions, and the genes encoding kappa and Lyt-2 are also located on mouse chromosome 6.

Animals↗
PubMed1987

Structure of the mouse gene encoding CD4 and an unusual transcript in brain.

The T-cell differentiation antigen CD4 plays an important role in the function of T cells that recognize class II major histocompatibility complex proteins. Mouse CD4 (L3T4) has previously been shown to be evolutionarily related to immunoglobulin variable regions based on the predicted protein sequence from cDNA clones. The gene encoding L3T4 was found to be transcribed not only in a subset of T-lineage cells but also unexpectedly in brain, where a shorter transcript was found. In the present study the gene encoding L3T4 is shown to span 26 kilobases and to contain 10 exons. The structural organization is similar to that of other members of the immunoglobulin gene superfamily except for the striking presence of an intron in the middle of the sequence encoding the amino-terminal immunoglobulin-like homology unit. The structure of the shorter L3T4 transcript in mouse brain has been determined. This mRNA appears to be generated from a transcriptional start site within the coding sequence in exon VI. If translated, this transcript would encode a protein of 217 amino acids that lacks the usual L3T4 signal peptide and the amino-terminal 214 amino acids of the mature protein.

Amino Acid Sequence↗
PubMed1987

L3T4 and the immunoglobulin gene superfamily: new relationships between the immune system and the nervous system.

L3T4 is a mouse cell surface protein expressed on most thymocytes and on the subset of mature T cells that recognizes class II MHC molecules. Its primary function on T cells is most likely that of increasing the avidity of the interaction between T cells and antigen-presenting or target cells. It may accomplish this by binding to a nonpolymorphic region on class II MHC molecules. The cDNA and gene encoding L3T4 have been isolated and sequenced. Analysis of the amino acid sequence predicted by the nucleotide sequence indicates that L3T4 is a member of the Ig gene superfamily. It is most closely related to Ig and Tcr V regions. Although the amino-terminal domain of L3T4 is the portion of the molecule that is most similar to V-regions, L3T4 is one of the polydomain members of the Ig gene superfamily. Studies of the expression of L3T4 mRNA in various tissues led to the surprising finding that this gene is transcribed not only in T lymphoid cells, but also in brain. The predominant form of L3T4 mRNA expressed in brain is foreshortened as compared to that in T lineage cells, and it is most likely the product of a distinct transcriptional start site. If translated, the protein encoded by this brain transcript would be 217 amino acids in length and would lack the signal peptide and the amino-terminal 214 amino acids of the mature protein. It is not known whether a stable protein product is synthesized from this mRNA or what its function might be. However, these findings place L3T4 in an intriguing class of Ig gene superfamily members characterized by coexpression in the immune system and the nervous system.

Animals↗
PubMed1986

Isolation and sequence of L3T4 complementary DNA clones: expression in T cells and brain.

T lymphocytes express on their surface not only a specific receptor for antigen and major histocompatibility complex proteins, but also a number of additional glycoproteins that are thought to play accessory roles in the processes of recognition and signal transduction. L3T4 is one such T-cell surface protein that is expressed on most mouse thymocytes and on mature mouse T cells that recognize class II (Ia) major histocompatibility complex proteins. Such cells are predominantly of the helper/inducer phenotype. In this study, complementary DNA clones encoding L3T4 were isolated and sequenced. The predicted protein sequence shows that L3T4 is a member of the immunoglobulin gene superfamily. It is encoded by a single gene that does not require rearrangement prior to expression. Although the protein has not previously been demonstrated on nonhematopoietic cells, two messenger RNA species specific for L3T4 are found in brain. The minor species comigrates with the L3T4 transcript in T cells, whereas the major species is 1 kilobase smaller.

Amino Acid Sequence↗
PubMed1986

Structure, sequence, and polymorphism of the Lyt-2 T cell differentiation antigen gene.

We have determined the nucleotide sequence and genomic organization of the mouse Lyt-2 T lymphocyte differentiation antigen gene. This gene consists of five exons and four introns, and the organization roughly parallels the protein domains. Alternative splicing to include or exclude exon IV (encoding part of the cytoplasmic tail) results in two forms of mRNA and accounts for the difference in size between the alpha- and alpha'-chains of Lyt-2. The gene structure provides further evidence for the evolutionary relationship between Lyt-2 and immunoglobulin genes. Comparison of the nucleotide sequence of the Lyt-2.1 and Lyt-2.2 alleles shows a high degree of conservation, but indicates that a single nucleotide change and consequent amino acid substitution in the variable region-like domain accounts for the serologic difference between these two alleles.

Alleles↗
PubMed1986

A mutational hot-spot within an intron of the mouse beta 2-microglobulin gene.

beta 2-Microglobulin is the smaller, relatively non-polymorphic chain of class I major histocompatibility complex proteins. We have previously described a mutant mouse cell line which had been selected for loss of the class I thymus leukemia (TL) antigen and had concomitantly lost surface expression of H-2k antigens. Expression of class I antigens on the cell surface was restored by fusion to an antigenically distinct mouse lymphoma line, and the defect in the mutant was shown to be the loss of a functional beta 2-microglobulin gene. We now describe three additional mutants with the same phenotype, all selected for loss of TL but after different types of mutagenesis. All of these mutants have genomic rearrangements resulting in the absence of a functional beta 2-microglobulin gene. These data provide strong evidence for the requirement of beta 2-microglobulin for cell surface expression of the heavy chain of class I major histocompatibility complex proteins. We further show that the defects in at least one beta 2-microglobulin gene in each mutant cell line map to the same small DNA segment within the first intron. The breakpoints of these mutations define a hypermutable site within the mouse beta 2-microglobulin gene.

Animals↗
PubMed1985

Gene for the human T cell differentiation antigen Leu-2/T8 is closely linked to the kappa light chain locus on chromosome 2.

We have mapped the gene encoding the T cell differentiation antigen Leu-2/T8 to human chromosome 2 by hybridization of a Leu-2/T8 complementary DNA clone to DNA from a panel of mouse-human cell hybrids. In situ hybridization further localizes the gene to the 2p1 region in close proximity to the Ig kappa light chain gene. The Leu-2/T8 gene translocates with C kappa to chromosome 8 in a Burkitt lymphoma line carrying a t(2;8) translocation. These data support the hypothesis that Leu-2/T8 is the human homologue of the mouse Lyt-2,3 antigen.

Animals↗
PubMed1985

Two Lyt-2 polypeptides arise from a single gene by alternative splicing patterns of mRNA.

The Lyt-2/3 molecule is a glycoprotein expressed on T lymphocytes and has classically been considered a marker for the cytotoxic/suppressor T cell subset. It has been postulated to be a receptor for class I major histocompatibility complex proteins. We have used a cDNA clone encoding the analogous human protein, Leu-2/T8, to isolate mouse cDNA clones, which were used as probes to isolate mouse genomic clones. By transfection we have shown that the mouse homologue of Leu-2/T8 is Lyt-2 and not Lyt-3. We have further demonstrated that two Lyt-2 polypeptide chains are encoded by a single gene and result from alternative modes of mRNA splicing. The nucleotide sequence of cDNA clones encoding each of these polypeptide chains has been determined and shows the difference between the two Lyt-2 polypeptide chains to be in the lengths of their cytoplasmic tails.

Amino Acid Sequence↗
PubMed1985

The T cell differentiation antigen Leu-2/T8 is homologous to immunoglobulin and T cell receptor variable regions.

Leu-2/T8 is a cell surface glycoprotein expressed by most cytotoxic and suppressor T lymphocytes. Its expression on T cells correlates best with recognition of class I major histocompatibility complex antigens, and it has been postulated to be a receptor for these proteins. We have determined the complete primary structure of Leu-2/T8 from the nucleotide sequence of its cDNA. The protein contains a classical signal peptide, two external domains, a hydrophobic transmembrane region, and a cytoplasmic tail. The N-terminal domain of the protein has striking homology to variable regions of immunoglobulins and the T cell receptor. The membrane-proximal domain appears to be a hinge-like region similar to that of immunoglobulin heavy chains. The superfamily of immunologically important surface molecules can now be extended to include Leu-2/T8.

Amino Acid Sequence↗
PubMed1985

Coordinate induction of Ia alpha, beta, and Ii mRNA in a macrophage cell line.

We demonstrated a tightly coordinated timing in the appearance of mRNA for the four class II (Ia) MHC chains, A alpha, A beta, E alpha, and E beta, and the Ia-associated invariant chain in a murine macrophage cell line after the addition of immune interferon (IFN-gamma) or of IFN-gamma-containing supernatants from Con A-stimulated spleen cells. The marked increase in mRNA levels for these molecules at approximately 8 hr after IFN-gamma addition contrasts sharply with the earlier, more gradual kinetics observed for class I (H-2) and beta 2-microglobulin mRNA. The difference in kinetics of IFN-gamma induction of class I and class II mRNA suggests differential regulation of the expression of Ia and H-2 antigens. The long lag period preceding detection of Ia mRNA raises the possibility that IFN-gamma may not directly mediate the increase in mRNA expression, but may act through an additional cellular intermediate.

Animals↗
PubMed1985

Structure of Leu-2/T8 as deduced from the sequence of a cDNA clone.

We have determined the primary structure of the Leu-2/T8 T lymphocyte differentiation antigen from the complete nucleotide sequence of a cDNA clone. The protein consists of a classical signal peptide, two external protein domains, a hydrophobic transmembrane segment, and a highly charged intracytoplasmic tail. The N-terminal external domain of the mature protein is homologous to immunoglobulin and T cell receptor variable regions. Protein structural predictions suggest that this domain can indeed fold like an immunoglobulin domain. The Leu-2/T8 protein has no segment that is homologous to a constant region. However, the membrane-proximal domain appears to serve as a hinge. These studies indicate that Leu-2/T8 is another member of the immunoglobulin supergene family.

Amino Acid Sequence↗
PubMed1984

Transcriptional control of HLA-A,B,C antigen in human placental cytotrophoblast isolated using trophoblast- and HLA-specific monoclonal antibodies and the fluorescence-activated cell sorter.

Human placental cell suspensions prepared by trypsin digestion were analyzed with several monoclonal antibodies on a multiparameter fluorescence-activated cell sorter (FACS). Five distinct cell populations were isolated on the basis of size and quantitative differences in the coordinate expression of cell surface antigens detected by monoclonal antibodies against an HLA-A,B,C monomorphic determinant (MB40.5) and against human trophoblasts (anti-Trop-2). By FACS analysis and after sorting we clearly identified the major cell population as cytotrophoblasts based on several independent criteria, including presence of trophoblast-specific surface antigens, Trop-1, and Trop-2; absence of all HLA class I, class II, and beta 2-microglobulin (beta 2m) antigens; absence of the pan-leucocyte and monocyte antigens, HLe1 and LeuM1, respectively; presence of Y-chromatin in a male placenta; presence of placental and not liver alkaline phosphatase; and a large, mononuclear morphology. These procedures provide a reproducible method for obtaining highly purified human cytotrophoblast populations for further studies. We measured by molecular hybridization (RNA or Northern blots) the HLA-A,B,C and beta 2m mRNA in total RNA extracted from sorted cytotrophoblasts. We find that normal human cytotrophoblasts have extremely small amounts of HLA-A,B,C mRNA: approximately 300 times less than that in the lymphoid cell line LCL-721 or normal lymphocytes. In contrast, they have approximately 11% the level of beta 2m mRNA present in LCL-721 cells. Thus, HLA-A,B,C antigen expression on human cytotrophoblasts is limited by the level of HLA heavy chain mRNA.

Alkaline Phosphatase↗
PubMed1984

Isolation of the gene encoding the human T-lymphocyte differentiation antigen Leu-2 (T8) by gene transfer and cDNA subtraction.

We report the isolation of genomic and cDNA clones encoding the human T-lymphocyte cell-surface differentiation antigen, Leu-2 (T8), by use of a combination of transfection, fluorescence-activated cell-sorting, and subtractive cDNA hybridization. We constructed a cDNA library with mRNA from a mouse L-cell transfectant in which the human Leu-2 gene is expressed and amplified. We identified Leu-2 cDNA clones by screening with a selected cDNA probe from a second amplified Leu-2 transfectant. This probe contained cDNA species not removed by hybridization with L-cell mRNA. A Leu-2 cDNA clone was used to isolate a genomic clone. Transfection with DNA from this clone resulted in a high number of Leu-2 transfectants. This approach can be used to clone genes coding for other cell-surface molecules.

Animals↗
PubMed1984

Limitation of differential expression of HLA-A,B,C antigens on choriocarcinoma cell lines by messenger RNA for HLA heavy chain but not by beta 2-microglobulin.

The relative amounts of HLA-A,B,C antigens, beta 2-microglobulin (beta 2m), and trophoblast antigens (Trop-1 and Trop-2) were determined on nine choriocarcinoma cell lines including seven lines of gestational origin and two lines of nongestational origin (from ovary and stomach) by quantitative immunofluorescence analysis using a fluorescence-activated cell sorter. Most of these lines expressed surface HLA to variable extents, but one had none detectable. However, all lines secreted readily measurable amounts of beta 2m. We analyzed total RNA extracted from these lines using northern blot molecular hybridization with HLA-A,B,C- and beta 2m-specific complementary DNA probes. We found no messenger RNA species which hybridized with the HLA probe in cells with no detectable HLA surface antigen and only small amounts of HLA-specific RNA in cells with low levels of HLA membrane antigen. Cells exhibiting surface HLA levels greater than about 30% of that on lymphocytes had much higher amounts of HLA-specific RNA than did choriocarcinoma cells with no or low HLA antigen expression. In contrast, RNA hybridizing with beta 2m-specific probes was present at the 20% level or higher (relative to lymphocytes) in all the cell lines tested. Thus, the expression of HLA-A,B,C is apparently limited in choriocarcinoma cells by the level of HLA heavy-chain RNA and not by the level of beta 2m RNA. We discuss these findings in relation to the normal trophoblastic or other origins of this tumor type and with respect to the regulation and function of HLA in trophoblasts.

Antibodies, Monoclonal↗
PubMed1983

Control of H-2 antigen and beta 2-microglobulin gene expression in mouse trophoblast cell clones.

We have investigated the expression of H-2 antigen and beta 2-microglobulin in recently established mouse trophoblast cell clones. These clones, derived from C57BL/6 (H-2b) or BALB/c (H-2d) strains, synthesized extra-embryonic endoderm- and trophectoderm-specific cytoskeletal proteins, termed "Endo A" and "Endo B," and no detectable SSEA-1 embryonic antigen. Flow microfluorometry indicated that H-2 antigen expression on the cell surface of trophoblast cells was very low, corresponding to 2-5% of the amounts found on differentiated teratocarcinoma cells (12-1a) and BALB/c 3T3 cells, respectively. Expression of beta 2-microglobulin was reduced to approximately equal to 14% of the amounts found on 12-la cells. Immunoprecipitation and polyacrylamide gel electrophoretic analysis indicated that the synthesis of H-2 antigen and beta 2-microglobulin in the trophoblast cells was lower than that found in normal spleen cells. In addition low, but unequal, levels of mRNA specific for H-2 antigen and beta 2-microglobulin were found in trophoblast cells by blot hybridization with cDNA probes. The low mRNA levels may be due to transcriptional control of the genes encoding H-2 antigen and beta 2-microglobulin.

Animals↗
PubMed1983

Linkage of beta 2-microglobulin and ly-m11 by molecular cloning and DNA-mediated gene transfer.

beta 2-Microglobulin (beta 2m) is expressed on the cell surface after introduction of a beta 2mb (C57BL/6N) genomic clone into thymidine kinase-deficient mouse L cells by cotransformation using the calcium phosphate precipitate method. Stable transformant cell lines were identified that express the beta 2mb allele, as determined by reaction of the cells with appropriate monoclonal antibodies and by two-dimensional gel electrophoresis of endogenously labeled immunoprecipitates of cell extracts. These beta 2mb transformants now express ly-m11.2, as detected by an indirect radioimmunoassay. A plasmid subclone of the beta 2mb gene that contains an 8.4-kilobase insert, after introduction into mouse L cells, similarly directs the synthesis of both the beta 2mb and the ly-m11.2 antigens. Thus, the beta 2mb and ly-m11.2 determinants most likely represent sites on the same protein structure.

Animals↗
PubMed1982

Structure of wild-type and mutant mouse beta 2-microglobulin genes.

beta 2-Microglobulin is encoded by a single gene per haploid genome in the mouse. This gene was cloned and sequenced and was shown to consist of four coding regions separated by three intervening sequences. Most of the protein (amino acids 3-95) is encoded in a single uninterrupted unit flanked by intervening sequences. In this respect, the genetic organization is similar to that of the evolutionarily related H-2 and immunoglobulin domains. A mutant cell line selected for loss of cell-surface expression of the thymus leukemic antigen and failing to express surface beta 2-microglobulin and H-2 was found to contain no detectable cytoplasmic beta 2-microglobulin. The genetic defect in this mutant was analyzed by Southern blots, which indicated that there are defects in both copies of the beta 2-microglobulin gene present in this cell. The concomitant loss of three cell-surface antigens was thereby linked to genetic defects in the beta 2-microglobulin locus.

Animals↗