Phagosomal maturation, acidification, and inhibition of bacterial growth in nonphagocytic cells transfected with Fc gamma RIIA receptors

Phagocytosis and killing of microbial pathogens by professional phagocytes is an essential component of the innate immune response. Recently, heterolo gous transfection of individual receptors into nonmyeloid cells has been us ed successfully to elucidate the early steps that signal phagosome formatio n. It is unclear, however, whether the vacuoles formed by such transfected cells are bona fide phagosomes, capable of fusion with endomembranes, of lu minal acidification, and of controlling the growth of microorganisms, The a im of the current study was to determine whether COS-1 and Chinese hamster ovary cells, rendered phagocytic by expression of human Fc gamma RIIA recep tors, express the cellular machinery required to support phagosomal maturat ion. Immunolocalization studies demonstrated that early endosomes, as well as late endosomes and/or lysosomes, fuse sequentially with phagosomes in th e transfectants, Microfluorescence ratio imaging of particles labeled with pH-sensitive dyes revealed that maturation of the phagosome was accompanied by luminal acidification, The drop in pH, which attained levels comparable to those reported in professional phagocytes, was prevented by inhibitors of vacuolar type HC-ATPases. Optimal phagosomal acidification required elev ation of cytosolic [Ca2+], suggesting that it results from fusion of endome mbranes bearing proton pumps. Moreover, the transfected cells effectively i nternalized live bacteria. Opsonization was essential for bacterial interna lization, implying that it occurred by Fc gamma RIIA-mediated phagocytosis, as opposed to invasion. Uptake into phagolysosomes was associated with inh ibition of bacterial growth, due at least in part to the low intraphagosoma l pH. These studies indicate that the biochemical events that follow recept or-mediated particle internalization in cells transfected with Fc gamma RII A receptors closely resemble the process of phagosomal maturation in neutro phils and macrophages, Fc gamma RIIA-transfected cells can, therefore, be u sed as a model for the study of additional aspects of phagocyte biology.