STRUCTURE OF THE GENE OF TUM- TRANSPLANTATION ANTIGEN-P198 - A POINT MUTATION GENERATES A NEW ANTIGENIC PEPTIDE .12.

Mutagen treatment of mouse tumor cell line P815 produces turn - variants that are rejected by syngeneic mice because they express new transplantation antigens. These turn - antigens are recognized by cytotoxic T lymphocytes (CTL) but induce no detectable antibody response. By transfecting P815 cell line P1.HTR with DNA of turn- variant P198, we obtained transfectants expressing turn - antigen P198 that could be identified on the basis of their ability to stimulate anti-P198 CTL. This was repeated with DNA of a cosmid library derived from variant P198, and a cosmid carrying the sequence encoding antigen P198 was recovered from a transfectant. GeneP198 is 3 kb long and contains eight exons. It shows no homology with previously identified turn - gene P91A, nor with any gene presently recorded in the data banks. The long open reading frame codes for a 23.5-kD protein. The antigenic allele of gene P198 differs from the normal allele by a point mutation located in exon 7. This mutation causes an Ala to Thr change, and was shown by site-directed mutagenesis to be responsible for the expression of the antigen. An 11-amino acid synthetic peptide covering the sequence surrounding the turn - mutation rendered P815 cells sensitive to lysis by anti-P198 CTL. The homologous peptide corresponding to the normal sequence of the gene did not, but it was able to compete for binding to major histocompatibility complex molecule Kd. We conclude that turn - mutation P198 generates a new epitope recognized by syngeneic T cells. As observed with gene P91A, we found that a fragment of gene P198 that contained only exons 3-7, cloned in nonexpression vectors, transferred efficiently the expression of the antigen. © 1990, Rockefeller University Press., All rights reserved.