Impaired intracellular transport and cell surface expression of nonpolymorphic HLA-E: evidence for inefficient peptide binding.
The assembly of the classical, polymorphic major histocompatibility complex class I molecules in the endoplasmic reticulum requires the presence of peptide ligands and beta 2-microglobulin (beta 2m). Formation of this trimolecular complex is a prerequisite for efficient transport to the cell surface, where presented peptides are scanned by T lymphocytes. The function of the other class I molecules is in dispute. The human, nonclassical class I gene, HLA-E, was found to be ubiquitously transcribed, whereas cell surface expression was difficult to detect upon transfection. Pulse chase experiments revealed that the HLA-E heavy chain in transfectants, obtained with the murine myeloma cell line P3X63-Ag8.653 (X63), displays a significant reduction in oligosaccharide maturation and intracellular transport compared with HLA-B27 in corresponding transfectants. The accordingly low HLA-E cell surface expression could be significantly enhanced by either reducing the culture temperature or by supplementing the medium with human beta 2m, suggesting inefficient binding of endogenous peptides to HLA-E. To analyze whether HLA-E binds peptides and to identify the corresponding ligands, fractions of acid-extracted material from HLA-E/X63 transfectants were separated by reverse phase HPLC and were tested for their ability to enhance HLA-E cell surface expression. Two fractions specifically increased the HLA class I expression on the HLA-E transfectant clone.
Ulbrecht
M
M
Institut für Immunologie, Ludwig-Maximilians-Universität München, Germany.
Kellermann
J
J
Johnson
J P
JP
Weiss
E H
EH
D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000276:immunology; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D005805:Genes, MHC Class I; D006680:HLA Antigens / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D015796:HLA-B27 Antigen / Q000096:biosynthesis / Q000235:genetics; D015395:Histocompatibility Antigens Class I / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D046911:Macromolecular Substances; D051379:Mice; D009101:Multiple Myeloma; D011485:Protein Binding; D014162:Transfection; D000097625:HLA-E Antigens
D006680:HLA Antigens; D015796:HLA-B27 Antigen; D015395:Histocompatibility Antigens Class I; D046911:Macromolecular Substances; D000097625:HLA-E Antigens