title: Sampling From the Proteome to the Human Leukocyte Antigen-DR (HLA-DR) Ligandome Proceeds Via High Specificity.
abstract: Comprehensive analysis of the complex nature of the Human Leukocyte Antigen (HLA) class II ligandome is of utmost importance to understand the basis for CD4(+)T cell mediated immunity and tolerance. Here, we implemented important improvements in the analysis of the repertoire of HLA-DR-presented peptides, using hybrid mass spectrometry-based peptide fragmentation techniques on a ligandome sample isolated from matured human monocyte-derived dendritic cells (DC). The reported data set constitutes nearly 14 thousand unique high-confident peptides,i.e.the largest single inventory of human DC derived HLA-DR ligands to date. From a technical viewpoint the most prominent finding is that no single peptide fragmentation technique could elucidate the majority of HLA-DR ligands, because of the wide range of physical chemical properties displayed by the HLA-DR ligandome. Our in-depth profiling allowed us to reveal a strikingly poor correlation between the source proteins identified in the HLA class II ligandome and the DC cellular proteome. Important selective sieving from the sampled proteome to the ligandome was evidenced by specificity in the sequences of the core regions both at their N- and C- termini, hence not only reflecting binding motifs but also dominant protease activity associated to the endolysosomal compartments. Moreover, we demonstrate that the HLA-DR ligandome reflects a surface representation of cell-compartments specific for biological events linked to the maturation of monocytes into antigen presenting cells. Our results present new perspectives into the complex nature of the HLA class II system and will aid future immunological studies in characterizing the full breadth of potential CD4(+)T cell epitopes relevant in health and disease.
authors.affiliation: From the ‡Institute for Translational Vaccinology, P.O. Box 450, 3720 AL Bilthoven, the Netherlands; §Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Science Faculty, Utrecht University, Padualaan 8, 3584 CH Utrecht, the Netherlands;
authors.forename: Geert P M
authors.initials: GP
authors.lastname: Mommen
authors.affiliation: §Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Science Faculty, Utrecht University, Padualaan 8, 3584 CH Utrecht, the Netherlands; ¶Netherlands Proteomics Centre, Padualaan 8, 3584 CH Utrecht, the Netherlands;
authors.forename: Fabio
authors.initials: F
authors.lastname: Marino
authors.affiliation: From the ‡Institute for Translational Vaccinology, P.O. Box 450, 3720 AL Bilthoven, the Netherlands;
authors.forename: Hugo D
authors.initials: HD
authors.lastname: Meiring
authors.affiliation: ‖Centre for Infectious Disease Control, National Institute for Public Health and the Environment, P.O. Box 1, 3720 AL Bilthoven, the Netherlands;
authors.forename: Martien C M
authors.initials: MC
authors.lastname: Poelen
authors.affiliation: ‖Centre for Infectious Disease Control, National Institute for Public Health and the Environment, P.O. Box 1, 3720 AL Bilthoven, the Netherlands;
authors.forename: Jacqueline A M
authors.initials: JA
authors.lastname: van Gaans-van den Brink
authors.affiliation: §Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Science Faculty, Utrecht University, Padualaan 8, 3584 CH Utrecht, the Netherlands; ¶Netherlands Proteomics Centre, Padualaan 8, 3584 CH Utrecht, the Netherlands; **Chemistry Research Laboratory, Department of Chemistry, University of Oxford, Mansfield Road, OX13TA, Oxford, United Kingdom; ‡‡Department of Biochemistry, University of Oxford, South Parks Road, OX1 3QU, Oxford, United Kingdom.
authors.forename: Shabaz
authors.initials: S
authors.lastname: Mohammed
authors.affiliation: §Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Science Faculty, Utrecht University, Padualaan 8, 3584 CH Utrecht, the Netherlands; ¶Netherlands Proteomics Centre, Padualaan 8, 3584 CH Utrecht, the Netherlands;
authors.forename: Albert J R
authors.initials: AJ
authors.lastname: Heck
authors.affiliation: ‖Centre for Infectious Disease Control, National Institute for Public Health and the Environment, P.O. Box 1, 3720 AL Bilthoven, the Netherlands; cecile.van.els@rivm.nl.
authors.forename: Cécile A C M
authors.initials: CA
authors.lastname: van Els
mesh_terms: D017951:Antigen Presentation; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000378:metabolism; D006684:HLA-DR Antigens / Q000378:metabolism*; D006801:Humans; D008024:Ligands; D013058:Mass Spectrometry; D010455:Peptides / Q000378:metabolism*; D020543:Proteome / Q000378:metabolism*
chemical_list: D006684:HLA-DR Antigens; D008024:Ligands; D010455:Peptides; D020543:Proteome