In human B cells, class II molecules of the major histocompatibility complex (MHC-II) accumulate in an endosomal/lysosomal compartment, the MIIC, in which they may encounter and bind peptides. An additional molecule required for MHC-II peptide binding, HLA-DM (DM), has also been localized to the MIIC. Neither the relationship of the MIIC to the endosomal system nor the mechanisms by which DM localizes to the MIIC are understood. To address these issues, DM localization was analyzed in cells that do or do not express MHC-II. DM alpha beta heterodimers were localized in transfected MHC-II-negative HeLa and NRK cells, in the absence of the MHC-II-associated invariant chain, to a prelysosomal/lysosomal compartment by immunofluorescence microscopy. To identify a potential targeting determinant, we analyzed the localization of a chimeric protein, T-T-Mb, in which the cytoplasmic tail of murine DM beta (Mb) was appended to the lumenal and transmembrane domains of a cell surface protein, Tac. Like intact DM, T-T-Mb was localized to a lysosomal compartment in HeLa and NRK cells, as judged by immunofluorescence and immunoelectron microscopy. T-T-Mb was rapidly degraded in this compartment by a process that was blocked by inhibitors of lysosomal proteolysis. The DM beta cytoplasmic tail also mediated internalization of anti-Tac antibody from the cell surface and delivery to lysosomes. Deletion from the DM beta cytoplasmic tail of the tyrosine-based motif, YTPL, resulted in cell surface expression of T-T-Mb and a loss of both degradation and internalization; alanine scanning mutagenesis showed that the Y and L residues were critical for these functions. Similarly, mutation of the same Y residue within full-length DM beta resulted in cell surface expression of DM alpha beta heterodimers. Lastly, T-T-Mb was localized by immunoelectron microscopy to the MIIC in a human B lymphoblastoid cell line. Our results suggest that a motif, YTPL, in the cytoplasmic tail of the beta chain of DM is sufficient for targeting either to lysosomes or to the MIIC.

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