title: A novel ligand in lymphocyte-mediated cytotoxicity: expression of the beta subunit of H+ transporting ATP synthase on the surface of tumor cell lines.
abstract: Extracellular adenosine triphosphate (eATP) has been suggested to play a role in lymphocyte-induced tumor destruction. We now provide evidence that a protein responsible for ATP synthesis in mitochondria may also play a physiologic role in major histocompatibility complex-independent, lymphocyte-mediated cytotoxicity. A 51.5-kD protein (p51.5) bearing structural and immunologic characteristics of the beta subunit of H+ transporting ATP synthase (E.C. 3.6.1.34, beta-H+ATPase, published molecular mass of 51.6 kD) was detected on the plasma membrane of three different human tumor cell lines studied. NH2-terminal amino acid sequence analysis of purified p51.5 from K562 tumor cells revealed 100% homology of 16 residues identified in the first 21 positions to the known sequence of human mitochondrial beta-H+ ATPase. Antibody directed against a 21-mer peptide in the ATP binding region of beta-H+ ATPase (anti-beta) reacted with only one band on Western blots of whole tumor extracts and tumor membrane extracts suggesting that the antiserum reacts with a single species of protein. Anti-beta reacted with the cell membranes of tumor cells as determined by fluorescence-activated flow cytometry and immunoprecipitated a 51.5-kD protein from surface-labeled neoplastic cells (but not human erythrocytes and lymphocytes). Purified p51.5 bound to human lymphocytes and inhibited natural killer (NK) cell-mediated cytotoxicity. Furthermore, anti-beta treatment of the K562 and A549 tumor cell lines inhibited NK (by > 95%) and interleukin 2-activated killer (LAK) cell (by 75%) cytotoxicity, respectively. Soluble p51.5 upon binding to lymphocytes retained its reactivity to anti-beta suggesting that the ATP binding domain and the lymphocyte-receptor binding domain reside in distinct regions of the ligand. These results suggest that beta-H+ ATPase or a nearly identical molecule is an important ligand in the effector phase (rather than the recognition phase) of a cytolytic pathway used by naive NK and LAK cells.
authors.affiliation: Department of Medicine, State University of New York Health Science Center at Brooklyn 11203.
authors.forename: B
authors.initials: B
authors.lastname: Das
authors.forename: M O
authors.initials: MO
authors.lastname: Mondragon
authors.forename: M
authors.initials: M
authors.lastname: Sadeghian
authors.forename: V B
authors.initials: VB
authors.lastname: Hatcher
authors.forename: A J
authors.initials: AJ
authors.lastname: Norin
mesh_terms: D000595:Amino Acid Sequence; D003602:Cytotoxicity, Immunologic; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism; D008024:Ligands; D008565:Membrane Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D008024:Ligands; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D006180:Proton-Translocating ATPases