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

Abstract

Fc gamma receptors (FcgammaRs) 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 (FcgammaR) 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 FcgammaR phagocytosis. Herein, we demonstrate that these kinases are not required for FcgammaR endocytosis. Endocytosis of heat-aggregated IgG (HA-IgG) by COS-1 cells stably transfected with FcgammaRIIA or chimeric FcgammaRI-gamma-gamma (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 FcgammaRIIA was cross-linked using HA-IgG or EA. However, inhibition of FcgammaR phosphorylation prevented only FcgammaR phagocytosis. Finally, biochemical analyses of PI3K(p85)-Syk binding indicated that direct interactions between native Syk and PI3K proteins are differentially regulated during FcgammaR phagocytosis and endocytosis. Overall, our results indicate that FcgammaR 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.

Spleen Tyrosine Kinase (Syk) Mediates IL-1β Induction by Primary Human Monocytes during Antibody-enhanced Dengue Virus Infection

“…Syk inhibition with BAY 61-3606 did not reduce the detection of intracellular DENV E protein, indicating that Syk is dispensable for viral replication. This is in agreement with the report that Syk activation is not required for Fc receptor-mediated endocytosis (69).…”

M2 Macrophages Phagocytose Rituximab-Opsonized Leukemic Targets More Efficiently than M1 Cells In Vitro

“…Indeed, human Ig preparations were also strongly inhibitory on phagocytosis. Previous studies have demonstrated that Fc␥RI (CD64), Fc␥RIIA (CD32A), and Fc␥RIIIA (CD16A) can mediate IgG-driven phagocytosis (31,36,37). With regard to the polymorphism of CD16A, previously reported to play a role in the clinical response of some patients to rituximab (38, 39), we did not detect a significant difference in binding or phagocytosis of target cells, using macrophages homozygous for valine or phenylalanine FIGURE 7.…”

The Ubiquitin Ligase c-Cbl Down-Regulates FcγRIIa Activation in Human Neutrophils

“…The TCR is another receptor whose function is modulated by c-Cbl through the ubiquitination of the TCR-chain (74). Finally, c-Cbl was shown to down-modulate the phagocytic function mediated by Fc␥Rs (75,76). These examples illustrate the involvement of the ubiquitin ligase activity of c-Cbl in the down-regulation of various immunoreceptors.…”