title: The transport of class I major histocompatibility complex antigens is determined by sequences in the alpha 1 and alpha 2 protein domains.
abstract: The transport of human-mouse hybrid class I histocompatibility antigens has been studied in a mutant human cell line, 174 X CEM.T2 (T2). T2, a somatic cell hybrid of human B- and T-lymphoblastoid cell lines (B-LCL and T-LCL, respectively), synthesized HLA-A2 and HLA-B5 glycoproteins, but expresses only low levels of A2 and undetectable levels of B5 at the cell surface. We have previously shown that the products of human class I genes introduced into T2 by transfection behave like the endogenous HLA-B5 glycoproteins, while the products of mouse class I alleles similarly introduced are transported normally to the cell surface. We have now determined that the surface expression of class I glycoproteins in T2 depends on the origin of the alpha 1 and alpha 2 domains. Human (HLA-B7) and mouse (H-2Dp) hybrid class I genes, encoding the leader, alpha 1, and alpha 2 sequences of one species fused to the alpha 3, transmembrane, and cytoplasmic domains of the other, were transfected into T2. Normal surface expression of the hybrid class I molecule was observed in T2 only when the leader, alpha 1, and alpha 2-encoding exons were derived from the mouse gene. The reciprocal construct, encoding human leader, alpha 1, and alpha 2 domains fused to the mouse alpha 3, transmembrane, and cytoplasmic regions, resulted in biosynthesis of a hybrid glycoprotein which was not transported to the cell surface. The products of both constructs were expressed normally in control cells. The effects of glycosylation on class I antigen transport were also studied using mutant class I constructs with altered glycosylation sites. Two mutant B7 genes encoding either an extra glycosylation site at position 176 or no glycosylation sites were transfected into T2. These mutant products were expressed at the cell surface in control cells, but were synthesized and not surface-expressed in T2. These data demonstrate that the HLA/H-2 transport dichotomy in T2 is a function of the origin of the alpha 1 and/or alpha 2 domains of the class I glycoprotein, and is not a reflection of glycosylation differences between the human and mouse molecules.
authors.affiliation: Department of Microbiology and Immunology, Duke University Medical Center, Durham, NC 27710.
authors.forename: J
authors.initials: J
authors.lastname: Alexander
authors.forename: J A
authors.initials: JA
authors.lastname: Payne
authors.forename: B
authors.initials: B
authors.lastname: Shigekawa
authors.forename: J A
authors.initials: JA
authors.lastname: Frelinger
authors.forename: P
authors.initials: P
authors.lastname: Cresswell
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000096:biosynthesis; D001692:Biological Transport / Q000235:genetics; D002460:Cell Line; D004560:Electricity; D015180:Electrophoresis, Gel, Two-Dimensional; D005091:Exons / Q000235:genetics; D005434:Flow Cytometry; D006031:Glycosylation; D015793:HLA-B7 Antigen / Q000235:genetics / Q000378:metabolism; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D013601:T-Lymphocytes / Q000378:metabolism*; D014162:Transfection
chemical_list: D000954:Antigens, Surface; D015793:HLA-B7 Antigen; D015395:Histocompatibility Antigens Class I; D011994:Recombinant Proteins