Constitutive and inflammatory immunopeptidome of pancreatic beta-cells

Nadine L Dudek, Chor Teck Tan, Dhana G Gorasia, Nathan P Croft, Patricia T Illing, Anthony W Purcell

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Type 1 diabetes is characterized by the autoimmune destruction of pancreatic ?-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 ?-cells, interferon-? (IFN-?)-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 Kd-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 ?-cells grown under normal conditions or after exposure to IFN-?. 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-? treatment compared with the simultaneous presentation of the endogenously processed Kd-restricted peptide Janus kinase-1355-363 ( 15,000 copies/cell). We have successfully sequenced peptides from NIT-1 ?-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. ? 2012 by the American Diabetes Association.
Original languageEnglish
Pages (from-to)3018 - 3025
Number of pages8
JournalDiabetes
Volume61
Issue number11
DOIs
Publication statusPublished - 2012
Externally publishedYes

Cite this

Dudek, Nadine L ; Tan, Chor Teck ; Gorasia, Dhana G ; Croft, Nathan P ; Illing, Patricia T ; Purcell, Anthony W. / Constitutive and inflammatory immunopeptidome of pancreatic beta-cells. In: Diabetes. 2012 ; Vol. 61, No. 11. pp. 3018 - 3025.
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Constitutive and inflammatory immunopeptidome of pancreatic beta-cells. / Dudek, Nadine L; Tan, Chor Teck; Gorasia, Dhana G; Croft, Nathan P; Illing, Patricia T; Purcell, Anthony W.

In: Diabetes, Vol. 61, No. 11, 2012, p. 3018 - 3025.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Dudek, Nadine L

AU - Tan, Chor Teck

AU - Gorasia, Dhana G

AU - Croft, Nathan P

AU - Illing, Patricia T

AU - Purcell, Anthony W

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AB - Type 1 diabetes is characterized by the autoimmune destruction of pancreatic ?-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 ?-cells, interferon-? (IFN-?)-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 Kd-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 ?-cells grown under normal conditions or after exposure to IFN-?. 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-? treatment compared with the simultaneous presentation of the endogenously processed Kd-restricted peptide Janus kinase-1355-363 ( 15,000 copies/cell). We have successfully sequenced peptides from NIT-1 ?-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. ? 2012 by the American Diabetes Association.

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