Cytoplasts from cytotoxic T lymphocytes are resistant to perforin-mediated lysis.
Cytotoxic T lymphocytes (CTL) contain a potent cytolytic pore-forming protein (PFP, perforin or cytolysin) localized in their cytoplasmic granules. In the presence of calcium, perforin lyses a variety of target cells (TC) non-specifically. CTL, however, are generally resistant to the lytic effect of perforin. In this work, cytoplasts from CTL and susceptible TC were made by centrifuging cells on a Ficoll density gradient in the presence of cytochalasin B. Characterization by electron microscopy and a serine esterase assay established that both CTL and TC cytoplasts were completely devoid of nuclei and CTL cytoplasts contained essentially no granules. CTL cytoplasts are just as resistant to perforin-mediated lysis as the intact CTL, and both TC and their corresponding cytoplasts are very sensitive to lysis. Furthermore, CTL cytoplasts are less effective than TC cytoplasts in inhibiting hemolysis, a property shared by the respective intact cells. These results indicate that soluble granular components do not confer resistance on CTL, and suggest that the protective agent(s) acts by impeding perforin binding to the CTL membrane.
Laboratory of Cellular Physiology and Immunology, Rockefeller University, New York, NY 10021.
D M
DM
Ojcius
S B
SB
Jiang
P M
PM
Persechini
P A
PA
Detmers
J D
JD
Young
D000818:Animals; D002499:Centrifugation, Density Gradient; D004950:Esterases / Q000096:biosynthesis; D066298:In Vitro Techniques; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000494:pharmacology*; D051379:Mice; D008854:Microscopy, Electron; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects*
D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D052899:Pore Forming Cytotoxic Proteins; D054353:Perforin; D004950:Esterases; C022635:serine esterase