Soluble tapasin restores MHC class I expression and function in the tapasin-negative cell line .220.

Tapasin forms a bridge between TAP (transporters associated with antigen processing) and MHC class I molecules and plays a critical role in class I assembly. In its absence, TAP and class I do not associate, and class I cell surface expression is reduced. We now identify two independent functions for tapasin. Tapasin increases TAP levels and allows more peptide to be translocated to the endoplasmic reticulum. Furthermore, when expressed in the tapasin-negative .220 cell line, recombinant soluble tapasin retains its association with class I and restores class I cell surface expression and function, even though it no longer binds TAP or increases TAP levels. This finding suggests that the association of tapasin with class I is sufficient to facilitate loading and assembly of class I molecules.

Negative regulation by HLA-DO of MHC class II-restricted antigen processing.

HLA-DM is a major histocompatibility complex (MHC) class II-like molecule that facilitates antigen processing by catalyzing the exchange of invariant chain-derived peptides (CLIP) from class II molecules for antigenic peptides. HLA-DO is a second class II-like molecule that physically associates with HLA-DM in B cells. HLA-DO was shown to block HLA-DM function. Purified HLA-DM-DO complexes could not promote peptide exchange in vitro. Expression of HLA-DO in a class II+ and DM+, DO- human T cell line caused the accumulation of class II-CLIP complexes, indicating that HLA-DO blocked DM function in vivo and suggesting that HLA-DO is an important modulator of class II-restricted antigen processing.

HLA-DR molecules from an antigen-processing mutant cell line are associated with invariant chain peptides.

The invariant chain, which associates with the major histocompatibility complex (MHC) class II molecules in the endoplasmic reticulum, serves two functions important in antigen processing. First, it prevents class II molecules from binding peptides in the early stages of intracellular transport. Second, it contains a cytoplasmic signal that targets the class II-invariant chain complex to an acidic endosomal compartment. Proteolytic cleavage and subsequent dissociation of the invariant chain then occurs, allowing peptides derived from endocytosed proteins to bind to released class II molecules before their expression at the cell surface. Certain human cell lines that are mutant in one or more MHC-linked genes are defective in class II-restricted antigen processing. Here we show that in transfectants of one of these cell lines, T2, this deficiency results in the association of a large proportion of class II molecules with a nested set of invariant-chain-derived peptides (class II-associated invariant chain peptides, or CLIP). HLA-DR3 molecules isolated from T2 transfectants can be efficiently loaded with antigenic peptides by exposure to a low pH in vitro, perhaps reflecting the in vivo conditions in which peptides associate with class II molecules. Addition of synthetic CLIP inhibits the loading process, indicating that CLIP may define the region of the invariant chain responsible for obstructing the class II binding site.