Constitutive and Inflammatory Immunopeptidome of Pancreatic beta-Cells

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Dudek, NL; Tan, CT; Gorasia, DG; Croft, NP; Illing, PT; Purcell, AWDate
2012-11-01Source Title
DiabetesPublisher
AMER DIABETES ASSOCUniversity of Melbourne Author/s
Dudek, Nadine; Gorasia, Dhana; CROFT, NATHAN; PURCELL, ANTHONY; TAN, CHOR; ILLING, PATRICIAAffiliation
Academic Services and RegistrarMelbourne Dental School
Biochemistry and Molecular Biology
Microbiology and Immunology
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Journal ArticleCitations
Dudek, N. L., Tan, C. T., Gorasia, D. G., Croft, N. P., Illing, P. T. & Purcell, A. W. (2012). Constitutive and Inflammatory Immunopeptidome of Pancreatic beta-Cells. DIABETES, 61 (11), pp.3018-3025. https://doi.org/10.2337/db11-1333.Access Status
Open AccessAbstract
Type 1 diabetes is characterized by the autoimmune destruction of pancreatic β-cells. Recognition of major histocompatibility complex (MHC)-bound peptides is critical for both the initiation and progression of disease. In this study, MHC peptide complexes were purified from NIT-1 β-cells, interferon-γ (IFN-γ)-treated NIT-1 cells, splenic and thymic tissue of 12-week-old NOD mice, and peptides identified by mass spectrometry. In addition to global liquid chromatography-tandem mass spectrometry analysis, the targeted approach of multiple-reaction monitoring was used to quantitate the immunodominant K(d)-restricted T-cell epitope islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)₂₀₆₋₂₁₄. We identified >2,000 MHC-bound peptides; 1,100 of these presented by β-cells grown under normal conditions or after exposure to IFN-γ. These include sequences from a number of known autoantigens. Quantitation of IGRP₂₀₆₋₂₁₄ revealed low-level presentation by K(d) (~25 complexes/cell) on NIT-1 cells after IFN-γ treatment compared with the simultaneous presentation of the endogenously processed K(d)-restricted peptide Janus kinase-1₃₅₅₋₃₆₃ (~15,000 copies/cell). We have successfully sequenced peptides from NIT-1 β-cells under basal and inflammatory conditions. We have shown the feasibility of quantitating disease-associated peptides and provide the first direct demonstration of the disparity between presentation of a known autoantigenic epitope and a common endogenously presented peptide.
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