Role of COPI in phagosome maturation

Phagosomes mature by sequentially fusing with endosomes and lysosomes. Vesi cle budding is presumed to occur concomitantly, mediating the retrieval of plasmalemmal components and the regulation of phagosomal size. We analyzed whether fission of vesicles from phagosomes requires COPI, a multimeric com plex known to be involved in budding from the Golgi and endosomes. The role of COPI was studied using ldlF cells, that harbor a temperature-sensitive mutation in E-COP, a subunit of the coatomer complex. These cells were made phagocytic toward IgG-opsonized particles by heterologous expression of hu man Fc gamma RIIA receptors. Following incubation at the restrictive temper ature, epsilon-COP was degraded in these cells and their Golgi complex disp ersed. Nevertheless, phagocytosis persisted for hours in cells devoid of ep silon-COP. Retrieval of transferrin receptors from phagosomes became ineffi cient in the absence of epsilon-COP, while clearance of the Fc gamma RIIA r eceptors was unaffected. This indicates that fission of vesicles from the p hagosomal membrane involves at least two mechanisms, one of which requires intact COPI. Traffic of fluid-phase markers and aggregated IgG-receptor com plexes along the endocytic pathway was abnormal in epsilon-COP-deficient ce lls. In contrast, phagosome fusion with endosomes and lysosomes was unimpai red. Moreover, the resulting phagolysosomes were highly acidic. Similar res ults were obtained in RAW264.7 macrophages treated with brefeldin A, which precludes COPI assembly by interfering with the activation of adenosine rib osylation factor. These data indicate that neither phagosome formation nor maturation are absolutely dependent on COPI. Our findings imply that phagos omal maturation differs from endosomal progression, which appears to be mor e dependent on COPI-mediated formation of carrier vesicles.