Characterization of a novel function-associated molecule (FAM) on murine natural killer cells.
Committee ChairHarris, David T.
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PublisherThe University of Arizona.
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AbstractTrue antigen receptors on murine natural killer (NK) cells have not been identified. In this study, using a novel monoclonal antibody (mAb) directed against a 43 kDa function-associated molecule (FAM) specific to murine NK cells, a "receptor-like" structure has been characterized. It was observed by two-color flow cytometric analysis that the anti-FAM mAb specifically binds to a subpopulation of nylon wool nonadherent (NWNA) lymphocytes (19-20%). The expression of FAM was restricted to NK cells that expressed the NK1.1 antigen. Analysis of FAM expression in various lymphoid tissues revealed that slenocytes expressed the greatest numbers of mAb(+) cells. Lymphokine activated killer (LAK) cells expressed higher levels of FAM. It was demonstrated that the anti-FAM mAb inhibited the lysis and the conjugate formation (i.e. recognition) of target cells by NK cells. Stimulation of NK cells with anti-FAM mAb resulted in enhanced cytotoxicity as well as lymphokine production by NK cells. Redirected lysis of the anti-FAM mAb bearing hybridoma indicated that this structure is an activating (i.e., signal-transducing) molecule. Modulation of FAM from the surface of NK cells resulted in a significant loss of cytotoxicity as well as conjugate formation. A partial amino acid sequence of the anti-FAM mAb recognized molecule from rat NK cells was found to be closely related to an intermediate filament, vimentin. Anti-vimentin mAb reacted with NK cells, as shown by flow cytometry, and further, inhibited the cytotoxicity and the conjugate formation between NK cells and target cells. Moreover, northern blot analysis and polymerase chain reaction (PCR) analysis revealed no differences in the size of the mRNA between NK cells that express this molecule and cells that are negative for the expression of this molecule. Treatment of NK cells with anti-sense oligonucleotides against the vimentin gene inhibited the expression and the lytic activity of NK cells against target cells. Taken as a whole, these results demonstrated that the "vimentin-like" NK cell surface molecule FAM may play an important role in the recognition, signal-transduction, and lytic mechanism of murine NK cells. The ligand for FAM on fish and human NK target cells has been shown to be a 43 kDa molecule. An anti-idiotypic (id) mAb against the NK target cell antigen-specific mAb (18C2) inhibits the cytotoxicity of rat NK cells by binding to their membrane. It has been shown that the anti-id mAb recognizes a vimentin-like FAM on the membrane of NK cells. This study describes that the ligand for FAM is also present on murine transformed cell lines. The mAb 18C2 recognized a 43 kDa molecule on murine NK target cells and inhibited the lysis and conjugate formation (i.e. recognition) of target cells by NK cells. Further, the role of MHC class I antigens in NK cell cytotoxicity was examined. It was determined that target cells that express low levels of MHC class I antigens in association with the novel target cell antigen were more sensitive to NK cell lysis.
Degree ProgramMicrobiology and Immunology