Biomedical subjects

J Maryanski

Publications and source records attributed to J Maryanski.

5 recordsLinked to original sources

PubMed1995

Cells expressing a major histocompatibility complex class I molecule with a single covalently bound peptide are highly immunogenic.

The major histocompatibility complex (MHC) class I molecules expressed at the cell surface are associated with a large number of different peptides so that the density of a given MHC-peptide complex is relatively low. Here we describe the properties of MHC class I molecules genetically attached to a single antigenic peptide. Cells expressing these fusion proteins are recognized by T cells specific for the particular MHC-peptide complex. Coculture of naive splenocytes with cells expressing these MHC-peptide fusion proteins and the B7.1 antigen allows the induction of primary cytotoxic T lymphocytes (CTL) in vitro. Injection of these cells into naive mice enhances the frequency of specific CTL precursors and protects against a subsequent challenge with a tumor or a virus bearing the antigenic peptide. Soluble MHC-peptide fusions were also produced in which all three components, that is, the heavy chain, beta 2-microglobulin and the peptide, have fused into a single-chain protein. The availability of MHC class I molecules bound to a single peptide provides valuable tools for the manipulation of CTL responses and the analysis of the selection processes in the thymus.

Amino Acid Sequence↗

PubMed1989

Expression of mutant H-2d proteins encoded by class I genes which alternatively process the 5' end of their transcripts.

Mutation of the 3' splice sites bordering exon 2 of the H-2Dd and H-2Kd genes generated alternatively spliced transcripts when the constructs were transfected into L cells (J. Immunol. 143:1018). The H-2Dd transcripts contained an additional 84 nucleotides derived from the first intervening sequence, whereas 60 extra bases were included in the H-2Kd mRNA. Proteins derived from these transcripts were recognized by mAb. Moreover, both Ag served as recognition elements for CTL, and the mutant H-2Kd molecule functioned as a restricting element for an Ag peptide. As a result of alternative splicing, the mutant proteins should have additional residues at their NH2 termini to increase their lengths by 28 (Dd) or 20 (Kd) amino acids. Immunoprecipitation and analysis on SDS-PAGE demonstrated that the mutant H-2Kd molecule was indeed larger than the normal H-2Kd protein, but the mutant and wild-type H-2Dd Ag were the same size. In addition, treatment of H-2Dd mutant and normal Ag with N-glycanase produced molecules of equal size, demonstrating that the mutant protein was completely glycosylated. Limited amino acid sequencing of this Ag indicated that it was normal H-2Dd. Therefore, before its transfer to the cell surface, post-translational modifications remove the additional NH2-terminal residues of the mutant Dd but not Kd protein.

Amino Acid Sequence↗

PubMed1985

Tumour cell variants with increased immunogenicity obtained by mutagen treatment.

By treating mouse tumour cells in vitro with mutagens it is possible to obtain at high frequency variants that are rejected by normal syngeneic mice. Such variants have been designated 'tum-' to distinguish them from the original tumorigenic (tum+) cell lines. Most tum- variants appear to express new, individual antigens that can be defined in vivo by cross-protection experiments or in vitro by cytolytic T lymphocytes. The failure of tum- variants to form progressive tumours is apparently the result of an immune rejection response. Mice that have rejected a tum- variant clone are often protected against a subsequent challenge with the original tum+ cells. This was even demonstrated for two spontaneous mouse tumours for which no immunogenicity could be demonstrated otherwise. An analysis of 21 variants derived from mastocytoma P815 with cytolytic T lymphocyte populations showed that the repertoire of tum- antigens probably exceeds 50 specificities. No two variants have been found to express the same new antigen. Variant-specific long-term cytolytic T lymphocyte clones were isolated and used for the immunoselection of secondary antigen-loss variants that could then be analysed for the presence of previously undefined residual variant antigenic determinants. Somatic cell hybrids were prepared between different P815 tum- variants or between tum- variants and the original P815 cells. An analysis of the hybrids with variant-specific cytolytic T lymphocyte clones showed that the expression of tum- antigens was dominant. The intriguing paradox of the high frequency of tum- variants in populations of mutagen-treated tumour cells may require new genetic and immunological approaches before being understood.

Animals↗

PubMed1983

Escape of mouse mastocytoma P815 after nearly complete rejection is due to antigen-loss variants rather than immunosuppression.

Even though mastocytoma P815 often undergoes a nearly complete rejection in syngeneic mice, the tumor cells almost always escape to form progressive tumors. We found that this was not due to the establishment of an immunosuppressed state because genetically marked P815 cells, that were injected in mice where tumor escape was occurring, were readily rejected. An analysis of escaping tumor cell populations with anti-P815 cytolytic T lymphocyte (CTL) clones showed the presence of stable resistant variants. Using antigen-loss variants found in escaping populations or selected in vitro with CTL clones, we were able to define four different tumor-associated antigenic specificities, each recognized by a specific CTL clone. One of these specificities was absent from all escaping tumor cells and another had been lost by some of them.

Animals↗

PubMed1982

Immunogenic variants obtained by mutagenesis of mouse mastocytoma P815. V. H-2 associativity of variant-specific antigens.

By in vitro mutagenesis of mastocytoma P815, it is possible to obtain tumor cell variants that are rejected by syngeneic mice (tum-). Most of these variants carry new individual antigens and stimulate a specific cytolytic T cell (CTL) response in mixed leukocyte tumor cell culture (MLTC). The H-2 associativity of this response was examined for six different variants by measuring the inhibition of cell-mediated cytolysis by antibodies directed against products of the K or the D end of the H-2d complex. The lysis was either not inhibited (variants P91 and P116) or inhibited selectively by anti-Kd (variants P21, P32 and P198) or anti-Dd antibodies (variant P35). All these tum- variants expressed Kd and Dd antigens as measured by absorption of H-2 alloantisera. Long-term CTL clones can be obtained that are specific for individual tum- antigens. The pattern of H-2 associativity obtained with MLTC-derived CTL against four tum- variants was verified with CTL clones directed against the specific antigens of these variants. Concordant results were observed in all cases. In addition to CTL clones specific for tum- antigens, it is possible to isolate clones against a P815 tumor-associated antigen found on all P815 tum- variants. For these clones no clear associativity with either Kd or Dd products was found.

Animals↗