Early Events in the Activation of FcγRIIA in Human Neutrophils: Stimulated Insolubilization, Translocation to Detergent-Resistant Domains, and Degradation of FcγRIIA

Abstract

The signal transduction mechanisms associated with the ligation of FcγRIIA in human neutrophils are as yet only incompletely characterized. In the present study, we have investigated the distribution and fate of FcγRIIA following its cross-linking. The results obtained indicate that cross-linking of FcγRIIA led, within a few seconds, to its translocation into a nonionic detergent-insoluble fraction. This was followed, within a couple of minutes, by a substantial loss of immunoreactive FcγRIIA in the cells. The stimulated degradation of FcγRIIA was blocked by the Src kinase inhibitor PP1 but not by wortmannin, ST-638, piceatannol, or cytochalasin B. Cross-linked FcγRIIA could be solubilized by saponin (in the presence of Nonidet P-40) and by β-octylglucoside. Sucrose gradient analysis of the distribution of FcγRIIA revealed that its cross-linking led to its translocation into the pellets and not the light buoyant density fractions classically associated with lipid rafts. Disruption of cholesterol-containing membrane microdomains with filipin prevented the degradation of FcγRIIA but did not inhibit the stimulation of the pattern of tyrosine phosphorylation or the mobilization of calcium that followed FcγRIIA cross-linking. These data suggest that both cholesterol-rich domains and Src kinases are required for the degradation of the activated FcγRIIA and provide new insights into the early events following FcγRIIA cross-linking.