Differential kinase requirements in human and mouse Fc-gamma receptor phagocytosis and endocytosis

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Abstract
Fc gamma receptors (FcγRs) contribute to the internalization of large and small immune complexes through phagocytosis and endocytosis, respectively. The molecular processes underlying these internalization mechanisms differ dramatically and have distinct outcomes in immune clearance and modulation of cell function. However, it is unclear how the same receptors (FcγR) binding to identical ligands (IgG) can elicit such distinct responses. We and others have shown that Syk kinase, Src-related tyrosine kinases (SRTKs) and phosphatidyl inositol 3-kinases (PI3K) play important roles in FcγR phagocytosis. Herein, we demonstrate that these kinases are not required for FcγR endocytosis. Endocytosis of heat-aggregated IgG (HA-IgG) by COS-1 cells stably transfected with FcγRIIA or chimeric FcγRI-γ-γ (EC-TM-CYT) was not significantly altered by PP2, piceatannol, or wortmannin. In contrast, phagocytosis of large opsonized particles (IgG-sensitized sheep erythrocytes, EA) was markedly reduced by these inhibitors. These results were confirmed in primary mouse bone marrow-derived macrophages and freshly isolated human monocytes. Levels of receptor phosphorylation were similar when FcγRIIA was cross-linked using HA-IgG or EA. However, inhibition of FcγR phosphorylation prevented only FcγR phagocytosis. Finally, biochemical analyses of PI3K(p85)-Syk binding indicated that direct interactions between native Syk and PI3K proteins are differentially regulated during FcγR phagocytosis and endocytosis. Overall, our results indicate that FcγR endocytosis and phagocytosis differ dramatically in their requirement for Syk, SRTKs, and PI3K, pointing to striking differences in their signal transduction mechanisms. We propose a competitive inhibition-based model in which PI3K and c-Cbl play contrasting roles in the induction of phagocytosis or endocytosis signaling cascades.
Funding Information
  • National Institutes of Health
  • Natural Sciences and Engineering Council of Canada