Treatment of the BAC1.2F5 macrophage cell line with Macrophage Colony Stimulating Factor (M-CSF) resulted in a rapid induction of vesiculation that was reminiscent of macropinocytosis. Time-lapse micrography showed that these vesicles initiated as small vesicles at the cell periphery, but grew in size and migrated with time to a perinuclear localisation after growth factor stimulation. Immunofluorescence showed that the M-CSF receptor (c-fms) associated with the small vesicles and also the larger phase-bright vesicles. Treatment with two distinct inhibitors showed that the rapid initiation of vesicle formation was not dependent on phosphatidylinositol-3′ (PI-3) kinase activity; however, the subsequent maintenance, maturation and translocation of the large, phase-bright, c-fms-containing vesicles was dependent on PI-3 kinase activity. The inhibitors could also reverse the further maturation of preformed vesicles. The inhibition of vesicle trafficking and maturation correlated with ablation of M-CSF-induced PI-3 kinase activity associated with p110(alpha). These data demonstrate a role for PI-3 kinase in vesicle trafficking and maintenance. PI-3 kinase activity was also necessary for the macropinocytotic response in macrophages, a process that is essential for efficient antigen processing and presentation in macrophage-like cells.

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