Identification and characterization of a membrane-bound cytotoxin of murine cytolytic lymphocytes that is related to tumor necrosis factor/cachectin.
Cytotoxic T lymphocytes (CTLs) kill their targets by a contact-dependent mechanism. We investigated the possibility that the CTL membranes themselves could exert direct cytotoxic activity. Murine CTLs that had been fixed with paraformaldehyde retained a slow cytotoxic activity toward various target cells that are also sensitive to another cytokine, tumor necrosis factor (TNF)/cachectin. This cytotoxic activity was neutralized by antibodies specific for TNF. Membrane fractions obtained from CTLs were cytotoxic to TNF-sensitive targets but not to several TNF-resistant cell lines. Immunoblot analysis revealed a membrane protein band of 50-60 kDa from CTLs that reacts with anti-TNF antibodies. The surface localization of this cytokine was further ascertained by flow cytometry, indirect immunofluorescence, and immunoelectron microscopy studies using TNF-specific antibodies. Radioiodination of CTL surface proteins followed by immunoprecipitation with anti-TNF antibodies confirmed the presence of a TNF-related cytokine in the plasma membranes of CTLs that migrated with an apparent molecular mass of 50-60 kDa under disulfide-reducing conditions. This cytokine can be removed from membranes by treatment with detergents but not with high-salt buffers, suggesting that it may be an integral membrane protein.
Liu
C C
CC
Laboratory of Cellular Physiology and Immunology, Rockefeller University, New York, NY 10021.
Detmers
P A
PA
Jiang
S B
SB
Young
J D
JD
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000032:analysis; D003603:Cytotoxins / Q000032:analysis*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D007150:Immunohistochemistry; D008565:Membrane Proteins; D051379:Mice; D008854:Microscopy, Electron; D008970:Molecular Weight; D013602:T-Lymphocytes, Cytotoxic / Q000032:analysis* / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis*
D003603:Cytotoxins; D008565:Membrane Proteins; D014409:Tumor Necrosis Factor-alpha