Immunogenic (tum-) variants of mouse tumor P815: cloning of the gene of tum- antigen P91A and identification of the tum- mutation.
Mutagen treatment of mouse P815 tumor cells produces tum- variants that are rejected by syngeneic mice because these variants express new surface antigens. These "tum- antigens" are recognized by cytolytic T lymphocytes but induce no detectable antibody response. Transfection of P815 cell line P1.HTR with DNA of tum- variant P91 yielded transfectants expressing tum- antigen P91A. They were detected by their ability to stimulate proliferation of cytolytic T lymphocytes [Wölfel, T., Van Pel, A., De Plaen, E., Lurquin, C., Maryanski, J. L. & Boon, T. (1987) Immunogenetics 26, 178-187]. A cosmid library of a cell line expressing antigen P91A was transfected into P1.HTR. Transfectants expressing the antigen were obtained. By packaging directly the DNA of a transfectant with lambda phage extracts, we obtained a small cosmid population containing as major component a cosmid that transferred the expression of P91A. The assay of various restriction fragments of this cosmid led to the isolation of an 800-base-pair fragment containing the P91A sequence required for transfection. Comparison with a homologous cDNA showed that this fragment contained only one of the several exons of the P91A gene. The normal and the tum- forms of the gene differ by one nucleotide located in this 137-base-pair exon. The essential role of this mutation, which produces an amino acid change, was confirmed by site-directed mutagenesis. No significant sequence similarity was found between the 800-base-pair fragment and any recorded gene.
D000595:Amino Acid Sequence; D000818:Animals; D000951:Antigens, Neoplasm / Q000235:genetics*; D001483:Base Sequence; D012515:Mast-Cell Sarcoma / Q000235:genetics / Q000276:immunology* / Q000473:pathology; D051379:Mice; D008811:Mice, Inbred DBA / Q000276:immunology; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D014407:Tumor Cells, Cultured / Q000276:immunology
D000951:Antigens, Neoplasm; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins