INTRACELLULAR ROUTES AND SELECTIVE RETENTION OF ANTIGENS IN MILDLY ACIDIC CATHEPSIN D LYSOSOME-ASSOCIATED MEMBRANE PROTEIN-1/MHC CLASS II-POSITIVE VESICLES IN IMMATURE DENDRITIC CELLS/

Immature dendritic cells (DC) use both macropinocytosis and mannose re ceptor-mediated endocytosis to internalize soluble Ags efficiently. Th ese Ags are ultimately presented to T cells after DC maturation and mi gration into the lymph nodes. We have previously described the immorta lized myeloid cell line FSDC as displaying the characteristics of earl y DC precursors that efficiently internalize soluble Ags. To describe the different routes of Ag uptake and to identify the Ag retention com partments in FSDC, we followed the intracellular fate of FITC-coupled OVA, dextran (DX), transferrin, and Lucifer Yellow using flow cytometr y, confocal microscopy, and immunoelectron microscopy. OVA and DX gain ed access into macropinosomes and early endosomes. DX was preferential ly sorted into endosomal compartments, while most of the OVA entered m acropinosomes via fluid phase uptake. We found a long-lasting retentio n of DX and OVA of up to 24 h. After 6 h of chase, these two molecules were concentrated in common vesicular compartments. These retention c ompartments were distinct from endosomes and lysosomes; they were much larger, only mildly acidic, and lacked the small GTP binding protein rab7. However, they were positive for lysosome-associated membrane pro tein-1, the protease cathepsin D, and MHC class II molecules, thus rep resenting matured macropinosomes. These data suggest that the activity of vacuolar proteases is reduced at the mildly acidic pH of these ves icles, which explains their specific retention of an Ag. The retention compartments might be used by nonlymphoid tissue DC to store peripher al Ags during their migration to the lymph node.