PubMed Health⌕ Search

Biomedical subjects

J Rothbard

Publications and source records attributed to J Rothbard.

At least 37 records · Page 2Linked to original sources
PubMed1988

A highly immunogenic region of a human polymorphic epithelial mucin expressed by carcinomas is made up of tandem repeats.

The nucleotide sequences of partial cDNA clones coding for the core protein of a human polymorphic epithelial mucin were determined, and a large domain was found to consist of a 60-base pair tandem repeat sequence. The cDNA clones were originally selected (Gendler, S. J., Burchell, J. M., Duhig, T., Lamport, D., White, R., Parker, M., and Taylor-Papadimitriou, J. (1987) Proc. Natl. Acad. Sci. U. S. A. 84, 6060-6064) using three monoclonal antibodies which show differential reactivity with the mucin produced by normal and malignant breast. Two of the epitopes are exposed in the normally processed and cancer-associated mucin, while one epitope is unmasked only in the cancer-associated mucin (Burchell, J. M., Durbin, H., and Taylor-Papadimitriou, J. (1983) J. Immunol. 131, 508-513; Burchell, J., Gendler, S., Taylor-Papadimitriou, J., Girling, A., Lewis, A., Millis, R., and Lamport, D. (1987) Cancer Res. 47, 5476-5482). We show here that all three antibodies react with a synthetic peptide with an amino acid sequence corresponding to that predicted by the tandem repeat. Identification of the epitopes preferentially expressed on the cancer-associated mucin should allow a directed approach to the development of tumor-specific antibodies using synthetic peptides as immunogens.

Alleles↗
PubMed1988

A monoclonal antibody recognising the site of limited proteolysis of protein kinase C. Inhibition of down-regulation in vivo.

A monoclonal antibody to protein kinase C is described that recognises the site of limited proteolysis on the native enzyme. Binding of the antibody to the purified kinase in vitro blocks partial proteolysis by trypsin, and introduction of the Fab fragment into a rodent glioma cell line inhibits phorbol-ester-induced down-regulation of the kinase. These observations are discussed in the context of the domain structure of protein kinase C and the agonist-induced proteolysis of the kinase in vivo.

Animals↗
PubMed1988

Peptide-mediated modulation of T-cell allorecognition.

Antigen-specific helper T cells recognize a complex of peptide antigen and class II major histocompatibility complex (MHC) gene products. Whether T cells recognize MHC class II alloantigen by a similar mechanism or the native conformation of MHC molecules themselves has yet to be determined. The demonstration that peptide antigens bind directly and specifically to class II molecules has allowed us to examine the influence of foreign peptide binding on T-cell recognition of allogeneic MHC molecules. We report here that an immunodominant, HLA-DR1-restricted peptide of influenza virus hemagglutinin (HA residues 306-320) is able to modulate the recognition of alloantigen by human DR1-specific T-cell clones. For some T-cell clones, but not all, the HA peptide inhibited allorecognition in a dose-dependent manner. However, in one instance, the proliferative response to alloantigen was enhanced in the presence of HA peptide. These results suggest that the specificities of T-cell responses to allogeneic MHC molecules are heterogeneous, which may be influenced by different peptides occupying the class II MHC combining site and by the diversity of antigen-specific receptors of T lymphocytes recognizing the same MHC/peptide complex.

Amino Acid Sequence↗
PubMed1988

A juxta-membrane epitope on the human acetylcholine receptor recognized by T cells in myasthenia gravis.

T cell proliferative responses to synthetic peptides taken from the human nicotinic acetylcholine receptor (AChR) alpha-chain sequence, or to whole AChR purified from electric fish (Torpedo marmorata), have been studied, using blood, thymus, and lymph node cells, from 34 patients with myasthenia gravis (MG) and 17 controls mostly with other neurological diseases. Peptides were selected because they contained amino acid motifs that recur in most defined T cell epitopes. Peptide 257-269 (from the extracellular loop of the AChR alpha-chain between the second and third trans-membrane domains) stimulated cells from six patients and no controls. Peptides from region 125-143 (from the main extracellular 1-210 stretch), which is thought to be an important T cell epitope in rats, provoked responses in 26% of patients and 41% of controls. Two patients responded both to these peptides and to peptide 257-269, thereby implying some heterogeneity of their reacting T cells. Whereas the initial blood T cell samples sometimes responded both to Torpedo AChR and to the 125-143 peptides, T cell lines selected with either antigen subsequently showed no response to the other. This observation suggests that it may be essential to use human AChR sequences for studying truly autoreactive T cells in MG. Finally, no strong association was found between any of the responses to peptides and the HLA types of the responding individuals.

Adult↗
PubMed1988

In vitro proliferative responses and antibody titers specific to human acetylcholine receptor synthetic peptides in patients with myasthenia gravis and relation to HLA class II genes.

To investigate which parts of the acetylcholine receptor are involved in the initiation and development of myasthenia gravis (MG), peptides representing different sequences of the human acetylcholine receptor alpha-subunit were synthesized. These peptides were tested for their ability to stimulate T cells of myasthenic patients and healthy control patients in proliferation assays and to bind to sera antibodies. Three of eight peptides discriminated significantly between the two groups in the proliferation assay, as well as in their ability to bind to serum antibodies. HLA-DR3 and DR5 were associated with proliferative responses to specific AChR peptides in the group of myasthenics. Acetylcholine receptor epitopes that might play a specific role in myasthenia gravis thus were demonstrated.

Amino Acid Sequence↗
PubMed1987

The human T3 gamma chain is phosphorylated at serine 126 in response to T lymphocyte activation.

The gamma subunit of the human T lymphocyte T3 antigen is rapidly phosphorylated on serine residues in vivo during the initiation of T cell activation by a polyclonal mitogen (Phaseolus vulgaris phytohemagglutinin), an activator of protein kinase C (phorbol 12,13-dibutyrate), and an elevator of intracellular calcium (ionomycin). The sites of phosphorylation were identified by comparing tryptic peptide analyses of T3 gamma chains labeled in vivo with various synthetic peptides, corresponding to portions of the cytoplasmic domain of the gamma chain that had been labeled in vitro using purified protein kinase C. Two sites, serines 123 and 126, were phosphorylated in response to ionomycin, whereas a single site, serine 126, was phosphorylated when T lymphocytes were stimulated by P. vulgaris phytohemagglutinin or when protein kinase C was directly activated by phorbol 12,13-dibutyrate. Immune activation of T cells via the protein kinase C pathway thus induces phosphorylation of a single site on the T3 gamma chain, namely serine 126.

Amino Acid Sequence↗
PubMed1987

Use of synthetic peptides of influenza nucleoprotein to define epitopes recognized by class I-restricted cytotoxic T lymphocytes.

The conserved epitopes of influenza nucleoprotein (NP) recognized by class I MHC-restricted CTL from CBA (H-2k) and C57BL/10 (H-2b) mice have been defined in vitro with synthetic peptides 50-63 and 365-379, respectively. Two Db-restricted clones were described that recognize different epitopes on peptide 365-379. Finally, the recognition of complete NP was shown to be approximately 200-fold less efficient than peptide in the cytotoxicity assay. These phenomena are closely related to results with class II-restricted T cells and they strengthen the hypothesis that influenza proteins are degraded in the infected cell before recognition by class I-restricted CTL.

Amino Acid Sequence↗
PubMed1986

HLA B37 determines an influenza A virus nucleoprotein epitope recognized by cytotoxic T lymphocytes.

Human influenza A virus-specific, cytotoxic T cells have been shown previously to recognize the virus nucleoprotein on infected cells. CTL preparations from four HLA B37-positive donors were shown to recognize a synthetic peptide that corresponded to amino acids 335-349 of the nucleoprotein sequence. Influenza-specific CTL from 10 donors of other HLA types failed to recognize this epitope. CD8+ CTL lines were derived from lymphocytes of two HLA B37-positive donors and used to show that the peptide was represented on virus-infected cells and to determine the probable boundaries of the epitope.

Cell Line↗
PubMed1986

The epitopes of influenza nucleoprotein recognized by cytotoxic T lymphocytes can be defined with short synthetic peptides.

A proportion of cytotoxic T lymphocytes (CTL) responding to infection by influenza recognize target cells that express the viral nucleoprotein. Recent work showed that CTL can recognize short overlapping regions of large nucleoprotein fragments expressed in transfected L cells. This led to the suggestion that CTL recognize segmental epitopes of denatured or degraded proteins in a similar way to helper T cells. One corollary of this idea is that CTL should recognize appropriate short peptides on the target cell surface. We demonstrate that the epitopes of nucleoprotein recognized by CTL in association with class I molecules of the major histocompatibility complex in both mouse and man can be defined with short synthetic peptides derived from the nucleoprotein sequence.

Alleles↗
PubMed1985

Pili of Neisseria meningitidis. Analysis of structure and investigation of structural and antigenic relationships to gonococcal pili.

To provide information useful for the design of a pilus vaccine effective for the prevention of both meningococcal and gonococcal disease, the electron microscopic morphology of meningococcal pili and the structural and antigenic relationships of meningococcal pili to gonococcal pili were investigated. Meningococcal pili were 4-6 nm in width, extended 500-6,000 nm from the organism surface, and occurred singly or in bundles composed of 8-10 pili per bundle. Meningococcal pilin varied between 17,250 and 20,600 daltons. Pilin was present in outer membrane preparations of some meningococcal isolates that were nonpiliated by electron microscopic examination. Antibodies to gonococcal pili, cyanogen bromide cleavage fragments of gonococcal pilin, or synthetic peptide analogues corresponding to regions of the gonococcal pilin sequence, were used to detect common meningococcal and gonococcal antigenic determinants that might indicate the existence of a conserved sequence beyond residue 29. Antibody to intact gonococcal pili or to the variable CNBR-3 region of gonococcal pilin detected little shared antigenicity with meningococcal pilin. However, pilin from all tested meningococcal isolates reacted with antibody to the CNBR-2 fragment of gonococcal pilin, a region highly conserved among gonococcal strains. Meningococcal pilins were also broadly crossreactive with antibody to a synthetic peptide corresponding to residues 69-84 of the gonococcal sequence, a part of the CNBR-2 region that appears to be critical for gonococcal receptor-binding function. If a sequence similar to 69-84 is also important for receptor-binding function in meningococcal pili, a peptide corresponding to this region may elicit antibodies that block the adherence function of pili elaborated by both Neisseria gonorrhoeae and N. meningitidis.

Amino Acid Sequence↗
PubMed1985

Immunogenicity of synthetic peptides from circumsporozoite protein of Plasmodium falciparum.

In a study of recombinant proteins that might be useful in developing a vaccine against malaria, synthetic peptides from the circumsporozoite (CS) protein of Plasmodium falciparum were found to be immunogenic for mice and rabbits. Antibody to peptides from the repeating region of the CS protein recognized native CS protein and blocked sporozoite invasion of human hepatoma cells in vitro. Antibodies to peptides from regions I and II had no biologic activity, although antibody to region I recognized processed CS protein by Western blot analysis. These data support the feasibility of developing a vaccine against the sporozoite stage of the malaria parasite by using synthetic peptides of the repeating region of the CS protein conjugated to a carrier protein.

Amino Acid Sequence↗
PubMed1985

Cloning and sequencing of a Moraxella bovis pilin gene.

Moraxella bovis pili have been shown to play a major role in both infectivity and protective immunity of bovine infectious keratoconjunctivitis. Sonicated M. bovis DNA from the piliated strain EPP63 was inserted into the vector lambda gt11 with EcoRI linkers. Recombinant phage were screened with an oligonucleotide probe based on the amino-terminal portion of the DNA sequence of a Neisseria gonorrhoeae pilin gene. Two candidate phages produced a protein that comigrated with EPP63 beta pilin in sodium dodecyl sulfate-polyacrylamide gels and bound anti-pilus antisera. The 1.9-kilobase insert from one of these, lambda gt11M182, was subcloned in both orientations into pBR322, forming the plasmids pMxB7 and pMxB9, both of which produced beta pilin, as did pMxB12, a HindIII deletion derivative of pMxB7. In HB101(pMxB12), the M. bovis pilin protein was shown to be primarily localized in the inner membrane. The entire 939-base-pair insert of pMxB12 was sequenced, revealing a ribosome binding site just upstream of the coding region and an AT-rich region further upstream containing some potential RNA polymerase recognition sites. The translation of the sequence predicts a six-amino-acid leader sequence preceding the phenylalanine that begins the mature protein. Codon usage analysis of the M. bovis beta pilin gene revealed greater use of the CUA codon for leucine than usual for a well-expressed Escherichia coli gene. Comparisons of the M. bovis EPP63 beta pilin protein sequence with other pilin gene sequences are presented.

Amino Acid Sequence↗
PubMed1984

Gonococcal pili. Primary structure and receptor binding domain.

The complete amino acid sequence of pilin from gonococcal strain MS11 and the sequence of constant and variable regions from strain R10 pilin have been determined in order to elucidate the structural basis for adherence function, antigenic diversity, and polymeric structure. The MS11 pilin sequence consists of 159 amino acids in a single polypeptide chain with two cysteines in disulfide linkage and serine-bonded phosphate residues. TC-2 (31-111), a soluble monomeric pilus peptide prepared by arginine-specific digestion, bound human endocervical, but not buccal or HeLa cells and therefore is postulated to encompass the receptor binding domain. Variable regions of CNBr-3 appear to confer antigenic diversity and comprise segments in which changes in the position of charged residues occur in hydrophilic, beta-turns. Residues 2-21 and 202-221 of gonococcal pilins and lower eucaryotic actins, respectively, exhibit 50% homology. When these residues are arranged at intervals of 100 degrees of arc on "helical wheels," the identical amino acids comprise a hydrophobic face on one side of the helix. This observation, the hydrophobic character of this region and the tendency for TC-1 (residues 1-30) to aggregate in water, suggest that this stretch interacts with other subunits to stabilize polymeric structure.

Amino Acid Sequence↗
PubMedDate not supplied

Structure, expression and divergence of T-cell receptor beta-chain variable regions.

Analysis of three new T-cell receptor beta-chain variable regions together with those in the literature indicates that they have both remarkable similarities and differences with those of immunoglobulin. Less than 10 V regions appear to predominate in the thymus. V beta sequences are much more heterogeneous at the amino acid level than are immunoglobulin V regions and they appear to diverge between species much more quickly, apparently the result of additional hypervariable regions. Three of these putative new hypervariable regions lie outside of the classical immunoglobulin binding site, an indication that important interactions may be occurring in these regions with polymorphic MHC determinants.

Amino Acid Sequence↗
PubMedDate not supplied

Cytotoxic T lymphocytes recognize a fragment of influenza virus matrix protein in association with HLA-A2.

Both human and murine cytotoxic T cells (CTL) elicited in response to infection with influenza A viruses have been shown to be specific for internal viral proteins, such as the matrix and nucleoprotein. Individual CTL epitopes have been identified in the nucleoprotein by successfully substituting short synthetic peptides for the intact virus in the preparation of target cells in cytotoxicity assays. The defined peptide epitopes have each been recognized by CTL in association with individual class I major histocompatibility complex (MHC) proteins, H-2Db, H-2Kk, H-2Kd (Taylor, P. et al., unpublished data) and HLA-B37. A logical strategy to investigate the molecular details of the interaction between antigen and MHC class I proteins would be to define an epitope recognized by the MHC class I molecule HLA-A2. This is because the amino-acid sequence is known, several variants of A2 have been characterized and the protein has been purified and crystallized. Here we describe a peptide derived from the influenza matrix protein that is recognized by human CTL in association with the HLA-A2 molecule.

Animals↗
PubMedDate not supplied

Novel chimaeric protein expressed in Philadelphia positive acute lymphoblastic leukaemia.

Cytogenic changes are becoming increasingly important in understanding the pathogenesis of human malignancies. The t(9;22) (q34;q11) translocation is one of the most consistent and generates the Philadelphia chromosome (Ph1) (ref. 1) in chronic myeloid leukaemia (CML); it has also been observed in some acute lymphoblastic leukaemias (ALL) (ref. 2). In CML the breakpoints occur on chromosome 22 in the region designated bcr (ref. 3) and result in the expression of a bcr-abl fusion product of relative molecular mass (MT) 210,000 (210K) with associated in vitro tyrosine kinase activity (P210bcr-abl). In some cases of Ph1-positive ALL, a novel abl-related protein (P190all-abl) of 190K has been shown to have tyrosine kinase activity. In this report we demonstrate that the P190all-abl protein has a bcr determinant from the amino-terminal region, but is lacking a bcr determinant normally found in the P210bcr-abl near the bcr-abl junction. The chimaeric nature of the P190all-abl was confirmed by sequential immunoprecipitation with antisera against abl and bcr peptides.

Cell Line↗