CONSTITUTIVE MACROPINOCYTOSIS ALLOWS TAP-DEPENDENT MAJOR HISTOCOMPATIBILITY COMPLEX CLASS-I PRESENTATION OF EXOGENOUS SOLUBLE-ANTIGEN BY BONE-MARROW-DERIVED DENDRITIC CELLS

Dendritic cells expanded from mouse bone marrow (BMDC) with granulocyt e/macrophage-colony-stimulating factor have potent T cell-stimulatory properties both in vitro and in vivo. This has been well documented fo r major histocompatibility complex (MHC) class II-restricted responses , and more recently using peptide-loaded and protein-pulsed DC for CD8 responses following adoptive transfer in mice. An unresolved question concerns the capacity of BMDC to present exogenous antigen on MHC cla ss I molecules, an unconventional mode of MHC class I loading for whic h there is now considerable evidence, particularly in macrophages. Her e, we show that BMDC exhibit high levels of macropinocytosis driven by constitutive membrane ruffling activity. Up to one-third of actively ruffling and macropinocytosing BMDC transferred pinocytosed horseradis h peroxidase into the cytosol following a 15-min pulse, suggesting tha t they might be capable of presenting exogenous soluble antigen on MHC class I molecules. We show that BMDC presented exogenous ovalbumin to a T cell hybridoma more effectively, more rapidly, and at lower exoge nous antigen concentrations than BM macrophages on a cell-for-cell bas is. Presentation was TAP dependent, brefeldin A sensitive, and blocked by inhibitors of proteasomal processing, demonstrating use of the cla ssical MHC class I pathway. Although effective presentation of exogeno us antigen by BMDC occurred in the absence of agents which stimulate m acropinocytosis, treatment with phorbol myristate acetate (PMA) enhanc ed both pinocytosis and MHC class I presentation by BMDC. Finally, PMA -stimulated BMDC exposed to exogenous ovalbumin in vitro were able to prime an antigen-specific cytotoxic T lymphocyte response following ad optive transfer in vivo.