Enhancement of Fcγ Receptor-Mediated Phagocytosis by Transforming Mutants of Cbl

Abstract

AbstractPhagocytosis mediated by FcγR plays an important role in host defense. The molecular events involved in this process have not been completely defined. The adapter protein Cbl has been implicated in FcγR signaling, but the function of Cbl in phagocytosis is unknown. Here we show that overexpression of the transforming mutants of Cbl, Cbl-70Z, and v-Cbl, but not wild-type (wt) Cbl, enhance phagocytosis mediated by FcγR in COS cells. Cbl-70Z, but not Cbl-wt, also enhanced FcγR-mediated phagocytosis in P388D1 murine macrophage cells. Cbl-70Z did not affect tyrosine phosphorylation or in vitro kinase activity of Syk, indicating that Syk may not be the direct target of Cbl-70Z in the enhancement of phagocytosis. A point mutation (G306E) in the phosphotyrosine domain of Cbl-70Z, as well as a C-terminal 67-aa deletion, partially abolished the enhancing effect on FcγR-mediated phagocytosis. A double mutant of Cbl-70Z containing both the G306E mutation and the C-terminal deletion completely lacked the ability to enhance phagocytosis. Thus, both the phosphotyrosine binding domain and the carboxyl-terminal tail were required for optimal enhancement of phagocytosis by Cbl-70Z. Functional phosphatidylinositol 3-kinase was required for Cbl-70Z to enhance phagocytosis, since wortmannin, a phosphatidylinositol 3-kinase inhibitor, inhibited FcγR-mediated phagocytosis in the presence of Cbl-70Z. These studies demonstrate that mutants of Cbl can modulate the phagocytic pathway mediated by FcγR and imply a functional involvement of c-Cbl in Fcγ receptor-mediated phagocytosis.