title: In vivo MHC class II presentation of cytosolic proteins revealed by rapid automated tandem mass spectrometry and functional analyses.
abstract: We report a strategy for high through-put sequence analyses of large MHC class II-bound peptide repertoires which combines automated electrospray ionization tandem mass-spectrometry with computer-assisted interpretation of the tandem mass spectra using the algorithm SEQUEST. This powerful approach discerned 128 peptide sequences displayed by the murine MHC class II molecule I-Ab in activated B cells and macrophages, including a surprisingly large number of peptides derived from self cytosolic proteins. Mice lacking the chaperone molecule H-2M were used to generate T cells specific for selected self peptides. Functional T cell analyses of ex vivo antigen-presenting cells indicated that peptides originating from cytosolic proteins are efficiently presented by splenic and thymic dendritic cells, but less so by resting B cells or thymic cortical epithelial cells. These results suggest that central tolerance to at least some MHC class II-bound self peptides derived from cytosolic proteins exists in vivo.
authors.affiliation: Howard Hughes Medical Institute, University of Washington School of Medicine, Seattle 98195, USA.
authors.forename: A R
authors.initials: AR
authors.lastname: Dongre
authors.forename: S
authors.initials: S
authors.lastname: Kovats
authors.forename: P
authors.initials: P
authors.lastname: deRoos
authors.forename: A L
authors.initials: AL
authors.lastname: McCormack
authors.forename: T
authors.initials: T
authors.lastname: Nakagawa
authors.forename: V
authors.initials: V
authors.lastname: Paharkova-Vatchkova
authors.forename: J
authors.initials: J
authors.lastname: Eng
authors.forename: H
authors.initials: H
authors.lastname: Caldwell
authors.forename: J R
authors.initials: JR
authors.lastname: Yates
authors.forename: A Y
authors.initials: AY
authors.lastname: Rudensky
mesh_terms: D000465:Algorithms; D000595:Amino Acid Sequence; D000818:Animals; D017951:Antigen Presentation / Q000276:immunology*; D000938:Antigen-Presenting Cells / Q000276:immunology* / Q000378:metabolism; D001324:Autoantigens / Q000737:chemistry* / Q000276:immunology* / Q000378:metabolism; D001331:Automation / Q000379:methods; D001402:B-Lymphocytes / Q000276:immunology / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003600:Cytosol / Q000737:chemistry*; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D004847:Epithelial Cells / Q000276:immunology / Q000378:metabolism; D000949:Histocompatibility Antigens Class II / Q000276:immunology* / Q000378:metabolism; D006825:Hybridomas / Q000276:immunology; D007108:Immune Tolerance / Q000276:immunology; D008264:Macrophages / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013601:T-Lymphocytes / Q000276:immunology; D013950:Thymus Gland / Q000166:cytology / Q000276:immunology
chemical_list: D001324:Autoantigens; D000949:Histocompatibility Antigens Class II; D011506:Proteins