Characterization of the nover late T cell activation proteins, 519 and 520

ABSTRACT

The late T cell activation gene, 519, is expressed in antigen specific, growth factor dependent T cell lines and clones but not in T or B cell tumors, other hematopoietic cells, tonsil, muscle, lung, or liver. Resting peripheral blood lymphocytes (PBLs) express little or no 519mRNA, but levels increase dramatically 5~7 days after activation with alloantigen or mitogen. Four alternatively spliced transcripts of the gene exist. 520 cDNA, the predominate gene transcript, encodes a protein of 144 amino acids and has a potentially cleavable hydrophobic leader of 15 amino acids at the amino terminus. The 519 transcript encodes a 129 amino acid product. An polyclonal antiserum was produced by immunizing a rabbit with a synthetic peptide corresponding to the second hydrophobic region common to both 519 and 520. 14.3 and 15.5kD protein bands were immunoprecipitated (IP) from radiolabled in-vitro translated products, 519 and 520 respectively. A protein of 14.3kD was detected by IP of in-vivo 35S cysteine and methionine radiolabled cytotoxic T lymphocytes (CTL) cells. Western blotting of lysates from a CTI line and a B cell tumor line revealed a distinct band at 14.3kD only in CTL lanes. Both CTL and peripheral blood lymphocytes expressed the greatest amount of 519/520 protein in cells 5~7 days after stimulation. Western blotting of differential centrifugation samples localized 519/520 protein to the granule layer. Comparison of 519/520 proteins with other known protin sequences identified homology with surfactant associated and sphyngolipid activating proteins. A homologous gene(NKC5) is present in natural killer cells (NK). The increase in the amount of the 519/520 protein product in CTL and PBL parallel increases in mRNA expression. 519/520 and related gene products are present in CTL, PBL and NK cells, have structural similarity with lipid associating proteins.