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T Delovitch

Publications and source records attributed to T Delovitch.

9 recordsLinked to original sources
PubMed1997

A novel technique for the study of Ras activity: electroporation of [alpha-32P]GTP.

An efficient, simple, and reproducible procedure for the assessment of Ras activity present in adherent mammalian cells is described. [alpha-32P]GTP was introduced by in situ electroporation into mouse C3H10T1/2 fibroblasts or their ras(val12)-transformed derivatives. After a 3-hr incubation at 37 degrees C, Ras was immunoprecipitated from cell extracts and the Ras-bound GTP/GTP + GDP ratio was determined by thin-layer chromatography. Contrary to Streptolysin-O permeabilization, the cells are not affected in any detectable way by the procedure, so that [alpha-32P]GTP binding and conversion to [alpha-32P]GDP can be studied over a period of time for the measurement of steady-state Ras activity. The results show that careful control of electric field intensity results in a great increase in the efficiency and specificity of labelling compared to the addition of [32P]orthophosphate to the culture medium, while the GTP/GTP + GDP ratios obtained were essentially the same as after in vivo labeling.

3T3 Cells↗
PubMed1995

Both MHC and background gene heterozygosity alter T cell receptor repertoire selection in an antigen-specific response.

Many autoimmune diseases are associated with specific class II MHC alleles; however, this association is not complete. One explanation for the variable expression of disease in susceptible individuals is that variability in the TCR repertoire may alter the potential to generate pathogenic autoreactive T cells. The current study was undertaken to examine the possibility that MHC and background heterozygosity, which is the norm in the outbred human population, alters the expressed TCR repertoire and, if so, whether this has an impact on peptide recognition and antigenic specificity. We, therefore, systematically analysed the beef insulin-specific TCR repertoire in inbred BALB/c mice before and after introduction of MHC heterozygosity (BALB/c x BALB.K)F1 mice, or MHC and background gene heterozygosity (BALB/c x A/J)F1 mice. We show that T cells from all three repertoires are predominantly Ad-restricted and recognize the same immunodominant peptide. Despite this, the beef insulin-specific TCR repertoires in F1 mice differ from those seen in BALB/c mice with the most dramatic changes seen in (BALB/c x A/J)F1 mice. These changes are accompanied by subtle differences in the antigenic specificity of the T cells. The results demonstrate that both MHC and background gene heterozygosity affect TCR repertoire selection, suggesting that the variable expression of autoimmune disease in individuals with a susceptible MHC allele may result, in part, from variability in the TCR repertoire introduced by this heterozygosity.

Amino Acid Sequence↗
PubMed1991

Amino acid residues in the T cell receptor CDR3 determine the antigenic reactivity patterns of insulin-reactive hybridomas.

We examined TCR gene usage in a panel of beef insulin/I-Ad-restricted T cell hybrids obtained from BALB/c mice. These hybrids demonstrated several distinct patterns of reactivity defined by their ability to respond to species variants of insulin. Correlation of TCR-alpha and -beta-gene usage with these patterns of reactivity demonstrated that TCR gene usage was restricted within Ag reactivity groups. In particular, V-J junctional regions (CDR3 equivalent) were restricted with conserved junctional amino acid motifs present in both TCR-alpha- and -beta-chains. Comparison of TCR gene usage in hybrids expressing identical V alpha and V beta gene segments but demonstrating different patterns of reactivity revealed that changes in either J alpha and/or J beta gene segment usage could alter antigenic reactivity. Indeed, single or limited amino acid differences within the CDR3 region were sufficient to markedly alter fine specificity. These data demonstrate the critical role for CDR3 in determining antigenic reactivity in beef insulin-reactive hybrids and are compatible with the current model of TCR/peptide/MHC interaction.

Amino Acid Sequence↗
PubMed1985

Expression of an HLA-Bw6-related specificity by the HLA-Cw3 molecule.

Radioimmunoassay of HLA-transformed mouse L cells expressing A3, A24, B7, or Cw3 HLA class I molecules with a set of monomorphic monoclonal antibodies distinguishes between A3-A24 and B7-Cw3 patterns of reactivity. Analyses with Bw6-specific monoclonal antibodies and a human alloantiserum demonstrate the expression by the HLA-Cw3 molecules of a Bw6 public specificity related to but not identical with that expressed by the HLA-B7 molecules. Exon-shuffling experiments and inhibition studies of monoclonal antibody cell-surface fixation indicate that similar parts of B7 and Cw3 molecules account for their serological cross-reactivity.

Animals↗
PubMed1985

HLA class I genes: from structure to expression, serology and function.

HLA class I genes have been isolated from phage and cosmid libraries and assayed by transfection into murine L cells. The transfection step proved to be very important because of the large number of genes (and pseudogenes) in this family. All functional genes characterized so far in this way are "classical" class I genes, i.e. members of the HLA-A, -B or -C families. Three of these have been sequenced (HLA-A3, -Aw24; HLA-Cw3) in addition to the pHLA 12.4 pseudogene. Sequence comparisons indicate, in particular, extreme conservation of the 3' non-coding region between allelic HLA-A locus genes; the general organization of all these genes (8 exons) is very similar. Restriction mapping around the functional genes has been performed to investigate the degree of conservation (e.g. between HLA-A3 regions from 2 different individuals) and examine allelism at the DNA level (e.g. between HLA-A3 and HLA-Aw24 regions). Exon shuffling experiments followed by serological analysis of the expressed product indicate that, as expected, specificities are determined by the first two domains of the molecule. However, further constructs show that as soon as a single exon is exchanged most specific reactivities disappear. CTL analysis of murine cells expressing HLA molecules has run into many difficulties but still holds promise for the study of structure-function relationships in this system.

Alleles↗
PubMed1979

Immunosuppressive ALS. II. Antibody to Ia antigens in heterologous anti-lymphocyte serum.

The relationship between Ia alloantigens and xenoantigens detected by immunosuppressive heterologous anti-lymphocyte sera has been investigated. Three rabbit anti-rat lymphocyte sera were examined for the presence of antibodies to Ia antigens. Two of these sera, an anti-thymus membrane and anti-lymphocyte sera detected labelled cell-surface Ia antigens (mol. wt 35,000 and 27,000) present on rat spleen B cells. The third antiserum, prepared against fractionated soluble lymphocyte antigens, was essentially non-reactive with these antigens. Of these three heterologous antisera, the anti-membrane serum reacted with the same antigens detected by two alloantisera. It seemed possible that such an antiserum could modify a recipient animal's immune response in vivo in a fashion identical to alloantibody to Ia antigens. In fact, all three heterologous antisera, including one devoid of antibody to Ia proved immunosuppressive in vivo. These results suggest that antibodies to antigens other than Ia can induce prolonged allograft survival. Since heterologous sera bind Ia antigens, it remains to be determined whether monospecific heterologous antisera to Ia antigens can allograft survival. The results raise the prospect that more than one antibody specificity may contribute to the immunosuppression achieved with ALS.

Animals↗
PubMedDate not supplied

Rearrangement and transcription of the beta-chain genes of the T-cell antigen receptor in different types of murine lymphocytes.

Rearrangements of T-cell receptor beta-chain genes are usually found on both chromosomal homologues, occurring by both deletional and non-deletional mechanisms. Two constant-region (C beta) genes have been identified previously and at least one is transcribed in every helper or cytotoxic T cell tested, but the choice of C beta gene expression is not correlated with the specialized functions of these T lymphocytes. By contrast, four of five suppressor T-cell hybridomas examined have deleted all known joining (J beta) gene segments and C beta genes and therefore may have antigen receptors encoded by different T-cell receptor gene families.

Animals↗