Indirect role for COPI in the completion of Fc gamma receptor-mediated phagocytosis

Recent evidence suggests that extension of pseudopods during phagocytosis r equires localized insertion of endomembrane vesicles. The nature of these v esicles and the processes mediating their release and insertion are unknown . COPI plays an essential role in the budding and traffic of membrane vesic les in intracellular compartments. We therefore assessed whether COPI is al so involved in phagosome formation. We used IdlF cells, a mutant line deriv ed from Chinese hamster ovary cells that express a temperature-sensitive fo rm of epsilon COP, To confer phagocytic ability to IdlF cells, they were st ably transfected with Fc receptors type IIA (Fc gamma RIIA), In the presenc e of functional COPI, Fc gamma RIIA-transfected IdlF cells effectively inte rnalized opsonized particles. In contrast, phagocytosis was virtually elimi nated after incubation at the restrictive temperature. Similar results were obtained impairing COPI function in macrophages using brefeldin A. Notably , loss of COPI function preceded complete inhibition of phagocytosis, sugge sting that COPI is indirectly required for phagocytosis, Despite their inab ility to internalize particles, COPI-deficient cells nevertheless expressed normal levels of Fc gamma RIIA, and signal transduction appeared unimpeded , The opsonized particles adhered normally to COPI-deficient cells and were often found on actin-rich pedestals, but they were not internalized due to the inability of the cells to extend pseudopods. The failure to extend pse udopods was attributed to the inability of COPI-deficient cells to mobilize endomembrane vesicles, including a VAMP3-containing compartment, in respon se to the phagocytic stimulus.