Immunologic Research

, Volume 55, Issue 1–3, pp 91–99 | Cite as

Mapping I-Ag7 restricted epitopes in murine G6PC2

  • Tao Yang
  • Anita C. Hohenstein
  • Catherine E. Lee
  • John C. Hutton
  • Howard W. Davidson
Immunology in Colorado

Abstract

G6PC2, also known as islet-specific glucose 6-phosphatase catalytic subunit-related protein (IGRP), is a major target of autoreactive CD8+ T cells in both diabetic human subjects and the non-obese diabetic (NOD) mouse. However, in contrast to the abundant literature regarding the CD8+ response to this antigen, much less is known about the potential involvement of IGRP-reactive CD4+ T cells in diabetogenesis. The single previous study that examined this question in NOD mice was based upon a candidate epitope approach and identified three I-Ag7-restricted epitopes that each elicited spontaneous responses in these animals. However, given the known inaccuracies of MHC class II epitope prediction algorithms, we hypothesized that additional specificities might also be targeted. To address this issue, we immunized NOD mice with membranes from insect cells overexpressing full-length recombinant mouse IGRP and measured recall responses of purified CD4+ T cells using a library of overlapping peptides encompassing the entire 355-aa primary sequence. Nine peptides representing 8 epitopes gave recall responses, only 1 of which corresponded to any of the previously reported sequences. In each case proliferation was blocked by a monoclonal antibody to I-Ag7, but not the appropriate isotype control. Consistent with a role in diabetogenesis, proliferative responses to 4 of the 9 peptides (3 epitopes) were also detected in CD4+ T cells purified from the pancreatic draining lymph nodes of pre-diabetic female animals, but not from peripheral lymph nodes or spleens of the same animals. Intriguingly, one of the newly identified spontaneously reactive epitopes (P8 [IGRP55–72]) is highly conserved between mice and man, suggesting that it might also be a target of HLA-DQ8-restricted T cells in diabetic human subjects, an hypothesis that we are currently testing.

Keywords

Autoimmune diabetes NOD IGRP Epitope 

Notes

Acknowledgments

This work was supported by NIH R01 DK052068 (to JCH and HWD), the University of Colorado Health Sciences Center Diabetes and Endocrinology Research Center (P30 DK57516), and American Diabetes Association research grant 1-04-RA-44 (to JCH). Tao Yang gratefully acknowledges support from an American Diabetes Association mentored post-doctoral fellowship (7-04-MN-19).

Supplementary material

12026_2012_8368_MOESM1_ESM.docx (19 kb)
Supplementary material 1 (DOCX 18 kb)

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Tao Yang
    • 1
    • 3
  • Anita C. Hohenstein
    • 1
  • Catherine E. Lee
    • 1
  • John C. Hutton
    • 1
  • Howard W. Davidson
    • 1
    • 2
  1. 1.Barbara Davis Center for Childhood DiabetesUniversity of Colorado Anschutz Medical CampusAuroraUSA
  2. 2.Integrated Department of ImmunologyUniversity of Colorado Anschutz Medical CampusAuroraUSA
  3. 3.Department of EndocrinologyThe First Affiliated Hospital of Nanjing Medical UniversityNanjingChina

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