CROSS-LINKING OF CD59 AND OF OTHER GLYCOSYL PHOSPHATIDYLINOSITOL-ANCHORED MOLECULES ON NEUTROPHILS TRIGGERS CELL ACTIVATION VIA TYROSINE KINASE

Many membrane proteins are attached via a glycosyl phosphatidylinositol (GPI) anchor. Proteins anchored in this way make no direct contact with the interior of the cell, therefore a role in signaling or activation would seem unlikely. Nevertheless, cross-linking of GPI-anchored proteins on human and murine T lymphocytes has been shown to cause calcium transients and cell activation. Our studies address the non-lethal events caused by the membrane attack complex of complement, which include release of Ca2+ from intracellular stores, and have suggested that the GPI-anchored complement inhibitor CD59 may be involved in signaling these events. We here report that cross-linking of CD59 on human neutrophils using specific monoclonal antibody and second antibody caused a rapid increase in intracellular free Ca2+ concentration (Ca2+ transient) due to release of Ca2+ from stores and also caused neutrophil oxidase activation. All antibodies against CD59 tested were effective and cross-linking of any other GPI-anchored protein expressed on neutrophils also initiated an increase in intracellular free Ca2+ concentration, whereas cross-linking of transmembrane proteins caused little or no response. A tyrosine kinase-dependent activation pathway was indicated by the demonstration of tyrosine phosphorylation on cross-linking and by blocking of the Ca2+ transient with the tyrosine kinase inhibitor herbimycin.