Cell-specific heterogeneity in sensitivity of phosphatidylinositol-anchored membrane antigens to release by phospholipase C.
Cell surface antigens thought to be linked to the membrane via phosphatidylinositol (PI) are incompletely, and variably, released by treatment with phosphatidylinositol-specific phospholipase C (PI-PLC). The basis for this was investigated with cloned tumor cell lines and PI-PLCs isolated from two species of bacteria. Residual Thy-1 antigen, which was detectable by flow cytometry, remained on all thymoma cell lines after exposure to very high concentrations of either purified enzyme. A majority of the presumptive PI anchored molecules on all of the cell lines was sensitive to release by PI-PLC derived from Bacillus thuringiensis. However, cell lines differed dramatically in the ease with which PI-PLC from Staphylococcus aureus liberated the same surface antigens. This heterogeneity was determined at the single cell level because at least five different PI-anchored antigens exhibited similar behavior on a given cell line or transfected subclones of it. The two phospholipases differed with respect to molecular mass, serological cross-reactivity and sensitivity to inhibition by NaCl and detergents. These observations suggest that the two types of PI-PLC may have distinct substrate specificities or sensitivities to environmental conditions which account for the difference in their ability to act on PI-anchored proteins in particular cell types. Such enzymes should continue to be important tools for investigating the method and significance of attachment of lymphocyte surface glycoproteins. In particular, the S. aureus PI-PLC can be used to demonstrate and investigate a previously unrecognized heterogeneity in cells which express PI-anchored molecules.
Low
M G
MG
Oklahoma Medical Research Foundation, Oklahoma City 73104.
Stiernberg
J
J
Waneck
G L
GL
Flavell
R A
RA
Kincade
P W
PW
D000818:Animals; D000954:Antigens, Surface / Q000378:metabolism*; D001413:Bacillus thuringiensis / Q000201:enzymology; D002460:Cell Line; D003429:Cross Reactions; D004306:Dose-Response Relationship, Immunologic; D000939:Epitopes / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D010716:Phosphatidylinositols / Q000378:metabolism*; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism / Q000494:pharmacology*; D013211:Staphylococcus aureus / Q000201:enzymology; D018800:Thy-1 Antigens
D000954:Antigens, Surface; D000939:Epitopes; D010716:Phosphatidylinositols; D018800:Thy-1 Antigens; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D043265:Phosphatidylinositol Diacylglycerol-Lyase