title: MHC class I-associated peptides produced from endogenous gene products with vastly different efficiencies.
abstract: We compared the efficiency of generating antigenic peptides from various polypeptide contexts expressed by recombinant vaccinia viruses. These included full-length influenza virus nucleoprotein (NP(1-498)), two truncated forms, and cytosolic and endoplasmic reticulum-targeted minimal peptides. Two peptides were studied, NP(50-57) (Kk-restricted) and NP(147-155) (Kd-restricted). The efficiency of peptide generation was measured in cytotoxicity assays by determining 1) the kinetics of presentation following infection using brefeldin A to block additional presentation and 2) the concentration of anti-class I mAbs required to block presentation. The two determinants behaved similarly, being presented most efficiently from minigene products, with intermediate efficiency from fragments, and least efficiently from NP(1-498). Direct quantitation of HPLC-purified peptides supported the validity of these simple methods to roughly estimate the efficiency of class I Ag presentation. It also surprisingly revealed that 60- to 90-fold more NP(50-57) than NP(147-155) peptide was present in cells expressing NP(1-498) or a rapidly degraded fragment (for NP(1-498), 1800 peptides/cell of NP(50-57) vs 30 peptides/cell of NP(147-155)). By contrast, nearly identical (and much greater) amounts of peptides were recovered from cells expressing minigene products (55,000 copies of either peptide/cell). These findings demonstrate 1) that immunodominant peptides from the same protein can be generated with vastly different efficiencies, and 2) that cytosolic or endoplasmic reticulum-targeted minigene products are presented far more efficiently than longer polypeptides.
authors.affiliation: Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892, USA.
authors.forename: L C
authors.initials: LC
authors.lastname: Anton
authors.forename: J W
authors.initials: JW
authors.lastname: Yewdell
authors.forename: J R
authors.initials: JR
authors.lastname: Bennink
mesh_terms: D000818:Animals; D019138:Antibodies, Blocking / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017951:Antigen Presentation / Q000276:immunology; D000939:Epitopes / Q000378:metabolism; D015395:Histocompatibility Antigens Class I / Q000276:immunology*; D009980:Influenza A virus / Q000276:immunology / Q000378:metabolism; D007700:Kinetics; D007739:L Cells; D051379:Mice; D008807:Mice, Inbred BALB C; D008808:Mice, Inbred CBA; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins / Q000096:biosynthesis / Q000276:immunology / Q000378:metabolism; D010452:Peptide Biosynthesis; D010455:Peptides / Q000032:analysis / Q000276:immunology*; D016601:RNA-Binding Proteins; D014615:Vaccinia / Q000276:immunology; D014758:Viral Core Proteins / Q000096:biosynthesis / Q000276:immunology / Q000378:metabolism
chemical_list: D019138:Antibodies, Blocking; D000911:Antibodies, Monoclonal; D000939:Epitopes; D015395:Histocompatibility Antigens Class I; C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D010455:Peptides; D016601:RNA-Binding Proteins; D014758:Viral Core Proteins