title: Mass tag-assisted identification of naturally processed HLA class II-presented meningococcal peptides recognized by CD4+ T lymphocytes.
abstract: The meningococcal class I outer membrane protein porin A plays an important role in the development of T cell-dependent protective immunity against meningococcal serogroup B infection and is therefore a major component of candidate meningococcal vaccines. T cell epitopes from porin A are poorly characterized because of weak in vitro memory T cell responses against purified Ag and strain variation. We applied a novel strategy to identify relevant naturally processed and MHC class II-presented porin A epitopes, based on stable isotope labeling of Ag. Human immature HLA-DR1-positive dendritic cells were used for optimal uptake and MHC class II processing of (14)N- and (15)N-labeled isoforms of the neisserial porin A serosubtype P1.5-2,10 in bacterial outer membrane vesicles. HLA-DR1 bound peptides, obtained after 48 h of Ag processing, contained typical spectral doublets in mass spectrometry that could easily be assigned to four porin A regions, expressed at diverging densities ( approximately 30-4000 copies/per cell). Epitopes from two of these regions are recognized by HLA-DR1-restricted CD4(+) T cell lines and are conserved among different serosubtypes of meningococcal porin A. This mass tag-assisted approach provides a useful methodology for rapid identification of MHC class II presented bacterial CD4(+) T cell epitopes relevant for vaccine development.
authors.affiliation: Laboratory of Analytical Chemistry, National Institute of Public Health and the Environment, Bilthoven, The Netherlands.
authors.forename: Hugo D
authors.initials: HD
authors.lastname: Meiring
authors.forename: Betsy
authors.initials: B
authors.lastname: Kuipers
authors.forename: Jacqueline A M
authors.initials: JA
authors.lastname: van Gaans-van den Brink
authors.forename: Martien C M
authors.initials: MC
authors.lastname: Poelen
authors.forename: Hans
authors.initials: H
authors.lastname: Timmermans
authors.forename: Gino
authors.initials: G
authors.lastname: Baart
authors.forename: Humphrey
authors.initials: H
authors.lastname: Brugghe
authors.forename: Joost
authors.initials: J
authors.lastname: van Schie
authors.forename: Claire J P
authors.initials: CJ
authors.lastname: Boog
authors.forename: Ad P J M
authors.initials: AP
authors.lastname: de Jong
authors.forename: Cécile A C M
authors.initials: CA
authors.lastname: van Els
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D017951:Antigen Presentation / Q000276:immunology*; D001425:Bacterial Outer Membrane Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism* / Q000382:microbiology; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D018984:Epitopes, T-Lymphocyte / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015798:HLA-DR1 Antigen / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006720:Homozygote; D006801:Humans; D016056:Immunodominant Epitopes / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D022401:Meningococcal Vaccines / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D008969:Molecular Sequence Data; D009345:Neisseria meningitidis / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D009584:Nitrogen / Q000378:metabolism; D009587:Nitrogen Isotopes / Q000378:metabolism; D010446:Peptide Fragments / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D018272:Porins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D001425:Bacterial Outer Membrane Proteins; D018984:Epitopes, T-Lymphocyte; D015798:HLA-DR1 Antigen; D016056:Immunodominant Epitopes; D022401:Meningococcal Vaccines; D009587:Nitrogen Isotopes; D010446:Peptide Fragments; D018272:Porins; C057699:porin protein, Neisseria; D009584:Nitrogen