Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 191 (4), 603-12

The Intestinal T Cell Response to Alpha-Gliadin in Adult Celiac Disease Is Focused on a Single Deamidated Glutamine Targeted by Tissue Transglutaminase

Affiliations

The Intestinal T Cell Response to Alpha-Gliadin in Adult Celiac Disease Is Focused on a Single Deamidated Glutamine Targeted by Tissue Transglutaminase

H Arentz-Hansen et al. J Exp Med.

Abstract

The great majority of patients that are intolerant of wheat gluten protein due to celiac disease (CD) are human histocompatibility leukocyte antigen (HLA)-DQ2(+), and the remaining few normally express HLA-DQ8. These two class II molecules are chiefly responsible for the presentation of gluten peptides to the gluten-specific T cells that are found only in the gut of CD patients but not of controls. Interestingly, tissue transglutaminase (tTG)-mediated deamidation of gliadin plays an important role in recognition of this food antigen by intestinal T cells. Here we have used recombinant antigens to demonstrate that the intestinal T cell response to alpha-gliadin in adult CD is focused on two immunodominant, DQ2-restricted peptides that overlap by a seven-residue fragment of gliadin. We show that tTG converts a glutamine residue within this fragment into glutamic acid and that this process is critical for T cell recognition. Gluten-specific T cell lines from 16 different adult patients all responded to one or both of these deamidated peptides, indicating that these epitopes are highly relevant to disease pathology. Binding studies showed that the deamidated peptides displayed an increased affinity for DQ2, a molecule known to preferentially bind peptides containing negatively charged residues. Interestingly, the modified glutamine is accommodated in different pockets of DQ2 for the different epitopes. These results suggest modifications of anchor residues that lead to an improved affinity for major histocompatibility complex (MHC), and altered conformation of the peptide-MHC complex may be a critical factor leading to T cell responses to gliadin and the oral intolerance of gluten found in CD.

Figures

Figure 1
Figure 1
Biochemical purification of a peptide fragment from α-9 recombinant stimulatory for TCC CD387 E34. A T cell–reactive superdex fraction of the tTG-treated α-9 recombinant chymotrypsin digest was separated by ion exchange chromatography (A). The absorbency at 216 nm is overlaid on a histogram depicting the proliferative response of TCC CD387 E34 to the resulting fractions. Fractions 18 and 19 were further separated by reverse-phase HPLC, each producing a small T cell–reactive peak in fraction 14 (fraction 19 shown in B). tTG treatment of synthetic peptide α-9(57–68) and separation by Mono-Q generates a new peak at fractions 17 and 18 that is recognized by TCC CD387 E34 (C).
Figure 2
Figure 2
Recognition of peptides α-9(57–68) and α-2(62–75) by two TCCs. Peptides α-9(57–68) (A) and α-2(62–75) (B) were tested in their native state (♦) or after treatment with human tTG (▪) or acid/heat (a/h; ▴) for their ability to induce proliferation in (A) TCC CD387 E34 and (B) TCC CD412 R5.32. Responses are given in cpm.
Figure 2
Figure 2
Recognition of peptides α-9(57–68) and α-2(62–75) by two TCCs. Peptides α-9(57–68) (A) and α-2(62–75) (B) were tested in their native state (♦) or after treatment with human tTG (▪) or acid/heat (a/h; ▴) for their ability to induce proliferation in (A) TCC CD387 E34 and (B) TCC CD412 R5.32. Responses are given in cpm.
Figure 3
Figure 3
Testing of synthetic glutamine to glutamic acid–substituted peptides for their ability to stimulate TCCs. TCC CD387 E34 (A) and TCC CD412 R5.32 (B) were tested for recognition of a set of singly substituted peptides that had the glutamine residues substituted for glutamic acid residues.
Figure 3
Figure 3
Testing of synthetic glutamine to glutamic acid–substituted peptides for their ability to stimulate TCCs. TCC CD387 E34 (A) and TCC CD412 R5.32 (B) were tested for recognition of a set of singly substituted peptides that had the glutamine residues substituted for glutamic acid residues.
Figure 4
Figure 4
Analysis of peptides α-9(57–68) and α-2(62–75) binding to DQ2. Acid/heat (a/h)-treated, glutamic acid–substituted, and truncated peptide variants of the (A) α-9(57–68) and (B) α-2(62–75) peptides were assessed for binding to DQ2 using a cell free competitive binding assay. The presumed binding frame for the two peptides is marked below the sequence. Broken bars indicate that peptide binds too poorly to DQ2 to be measured accurately in this assay.
Figure 4
Figure 4
Analysis of peptides α-9(57–68) and α-2(62–75) binding to DQ2. Acid/heat (a/h)-treated, glutamic acid–substituted, and truncated peptide variants of the (A) α-9(57–68) and (B) α-2(62–75) peptides were assessed for binding to DQ2 using a cell free competitive binding assay. The presumed binding frame for the two peptides is marked below the sequence. Broken bars indicate that peptide binds too poorly to DQ2 to be measured accurately in this assay.
Figure 5
Figure 5
Recognition of peptides α-9(57–68)E65 and α-2(62–75)E65 by polyclonal TCLs. 17 TCLs were tested for recognition of the synthetic α-9(57–68)E65 and α-2(62–75)E65 peptides at 5 μM. The y-axis crosses the x-axis at a stimulation index of 1, i.e., where no effect on T cell proliferation is observed.
Figure 6
Figure 6
Recognition of α-gliadin recombinants by a panel of polyclonal TCLs. Stimulation of polyclonal TCLs by α-9 (57–68) and α-2(62–75) peptides or tTG-treated α-recombinants that have been digested with pepsin or chymotrypsin. Peptides were tested at 5 μM. Recombinant α-gliadins and gliadin from the wheat variety Kadett were tested at 50 μg/ml. Data is shown for five representative TCLs out of eight that were tested.

Similar articles

See all similar articles

Cited by 169 articles

See all "Cited by" articles

References

    1. Sollid L.M., Markussen G., Ek J., Gjerde H., Vartdal F., Thorsby E. Evidence for a primary association of celiac disease to a particular HLA-DQ α/β heterodimer. J. Exp. Med. 1989;169:345–350. - PMC - PubMed
    1. Sollid L.M., Thorsby E. HLA susceptibility genes in celiac diseasegenetic mapping and role in pathogenesis. Gastroenterology. 1993;105:910–952. - PubMed
    1. Lundin K.E.A., Scott H., Hansen T., Paulsen G., Halstensen T.S., Fausa O., Thorsby E., Sollid L.M. Gliadin-specific, HLA-DQ(α1*0501,β1*0201) restricted T cells isolated from the small intestinal mucosa of celiac disease patients. J. Exp. Med. 1993;178:187–196. - PMC - PubMed
    1. Molberg Ø., Kett K., Scott H., Thorsby E., Sollid L.M., Lundin K.E.A. Gliadin-specific HLA DQ2-restricted T cells are commonly found in small intestinal biopsies from coeliac disease patients, but not from controls. Scand. J. Immunol. 1997;46:103–108. - PubMed
    1. Nilsen E.M., Lundin K.E.A., Krajci P., Scott H., Sollid L.M., Brandtzaeg P. Gluten specific, HLA-DQ restricted T cells from coeliac mucosa produce cytokines with Th1 or Th0 profile dominated by interferon-γ. Gut. 1995;37:766–776. - PMC - PubMed

Publication types

MeSH terms

Feedback