Extracellular proteins are not generally presented on class I MHC molecules in vitro, yet many studies show that a pathway exists in vivo for the presentation of extracellular material on class I molecules to prime CDS+ T cell responses. Here, we provide morphological evidence that proteins taken up by macropinocytosis can gain access to the cytosol and therefore into the conventional class I MHC pathway. Class I presentation of soluble ovalbumin by mouse bone marrow macrophages was dramatically enhanced by MCSF or phorbol ester and blocked by amiloride, which stimulate and inhibit membrane ruffling and macropinocytosis, respectively. Brefeldln A, gelonin, and a pep tide aldehyde inhibitor of proteasomal processing each blocked presentation of macropinocytosed antigen, demonstrating that unusual access to the conventional class I MHC pathway was occurring. This novel cell type-specific endocytic pathway may facilitate presentation of exogenous material on class I MHC molecules, allowing induction of CDS+ T cell responses to soluble protelns, tumor cell fragments, and some pathogens. introduction

The conventional view of antigen presentation to T cells is that endogenous proteins are presented on class I major histocompatibility complex (MHC) molecules and exogenous proteins on class II MHC molecules (Morrison et al., 1986; Bevan, 1987; Townsend and Bodmer, 1989; Germain and Marguiles, 1993). However, an increasing number of reports demonstrate that this division is not absolute and that a significant level of cross-over can occur, such that peptides from endogenous cytosolic proteins may appear on class II MHC molecules (Jaraquemada et al., 1990; Dodi et al., 1994) and peptides from exogenous proteins on class I MHC molecules. CDS+ T cell responses can be elicited in vivo in the absence of de novo antigen synthesis within host antigen-presenting cells (APCs). These responses have been observed following immunization with purified proteins (Wraith et al., 1967; Staerz