Team:BioIQS-Barcelona/Celiac disease

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Project | Celiac Disease

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Understanding Celiac Disease

Celiac disease, also known as gluten-sensitive enteropathy, is the most common intestinal disorder of western populations. The disease pathology is characterized by an autoimmune enteropathy caused by an abnormal immune response to dietary gluten peptides that occurs in genetically susceptible individuals1. The overall prevalence of celiac disease in the United States in average-risk groups, according to several studies, is 0.7% to 1.0%.

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Symptoms

The ingestion of immunogenic gluten-derived peptides leads to inflammation and intestinal mucosal damage, which may result in a spectrum of:

  • gastrointestinal symptom
  • nutritional abnormalities
  • systemic complications
  • anaemia
  • osteoporosis
  • secondary autoimmunity
  • malignancy

What cells and molecules are involved?

The primary event of a gluten-induced immune response requires epitope-bearing oligopeptides to have access to lamina propria inside intestinal epithelial layer. Such oligopeptides are efficiently hydrolyzed by peptidases located in the brush border membrane of the differentiated enterocytes before they can be transported across the epithelium.

Once in the lamina propria, a deamination step performed by tissue transglutaminase (tTG) is necessary for these peptides to be presented by mature dendritic cells to T cell receptors by using the heterodimer encoded by HLA-DQ2 or DQ8 genes A and B and thus, triggering an immune response.

How does the HLA interact with gliadin peptides?

The molecular interaction between a gluten-derived peptide (gliadin) and specific HLA haplotypes has been broadly discussed. It has been exposed that the HLA association in celiac disease can be explained by a superior ability of DQ2/8 haplotypoes to bind the biased repertoire of proline-rich gluten peptides that have survived gastrointestinal digestion and that have been deamidated by tissue transglutaminase.

The gliadin–DQ* complex retains critical hydrogen bonds between the MHC and the peptide backbone despite the presence of many proline residues in the peptide that are unable to participate in amide-mediated hydrogen bonds.

Gliadin peptides

Gliadin is the water-soluble component of gluten, while glutenin is insoluble. There are three main types of gliadin (α, γ, and ω) that can trigger the immune response of celiac disease. Different gliadins are involved in the disease process in a different manner, some fragments being ‘toxic’ and others ‘immunogenic’.

A gliadin peptide is defined as ‘toxic’ if it is able to induce mucosal damage either when added in culture to duodenal mucosal biopsy, or when administered in vivo on proximal and distal intestine, whereas a fragment is defined ‘immunogenic’ if it is able to specifically stimulate HLA-DQ2- or HLA-DQ8-restricted T cell lines derived from jejunal mucosa or peripheral blood.

As mentioned, a common feature among gluten epitopes is the presence of multiple proline and glutamine residues, which make the peptides exceptionally impregnable by gastric, pancreatic and intestinal digestive proteases.

Moreover, tTG can hydrolyse glutamine to glutamic acid either at a lower pH or when no acceptor proteins are available, a process leading to an enhanced immunogenicity of gluten peptides.

HLA-DQ2 and DQ8 structures enhance affinity for deamidated gliadin peptides

HLA-DQ2 and -DQ8 haplotypes have greater risk of developing celiac disease. This is because single-nucleotide polymorphisms (SNP) on HLA-DQ α- and β-chains results on greater affinity for deamidated gliadin peptides. This deamidated peptides show negatively charged glutamic acid residues that are preferred in positions 4, 6, and 7 of the antigen-binding site's positively charged residues of HLA, allowing the formation of salt bridges.

In order to easily evaluate how these SNP on HLA-DQ α- and β-chains lead to a greater affinity for gliadin peptides, multiple alignment of HLA-DQ protein sequences has been performed and curated for DQ2 and DQ8 haplotypes. HLA-DQA and -DQB protein genotypes alignment was performed with protein sequences from IPD-IMGT/HLA (Directory hosted at the European Bioinformatics Institute) version 3.33.0 using MAFFT default alignment method.

Previous studies (Kim C-Y et al. 2004) have characterized the molecular interactions using PDB crystal structure (1S9V) of HLA-DQ2 complexed with deamidated gliadin peptide (LQPFPQPELPY). Such study proposed an interaction profile of 13 hydrogen bonds between the main-chain atoms of αI-gliadin and HLA-DQ2.

  • Ten interactions are mediated by conserved MHC residues (Arg-α52–COP2, His-β81–COP1, Asn-β82–NHP2, Asn-β82–COP2, Asn-α62–H2O–COP4, Asn-α11–H2O–COP4, Asn-α62–NHP6, Asn-α69–COP7, Trp-β61–H2O–COP8, Asn-α69–NHP9).
  • Three interactions are mediated by polymorphic MHC residues (Tyr-α9–NHP4, Tyr-α22–H2O–COP4, Lys-β71–H2O–COP5).

Putative hydrogen-bonding network in the DQ2–αI-gliadin complex (shown as red dashes). αI-gliadin is shown in yellow (C, yellow; N, blue; O, red). Backbone structure of HLA-DQ α- and β-chains are shown in green and blue ribbon plots, respectively, and side chains engaged in hydrogen bonding are shown in gray (C, gray; N, blue; O, red). Extracted from Kim, C.-Y., Quarsten, H., Bergseng, E., Khosla, C., & Sollid, L. M. in Proceedings of the National Academy of Sciences 2004; 101(12): 4175–4179.

Single-nucleotide polymorphisms on HLA-DQ residues lead to gliadin binding

α-52

Hidden Markov Models for HLA-DQ-α chain for residues 50-53. For total HMM, all described HLA-DQA protein sequences were used. For DQ2- and DQ8-positive, total MSA was curated as follows: DQ2-positive contains HLA-DQA1*0501 or *0505 sequences and DQ8-positive HLA-DQA1*03. DQ2 presents a deletion on R53.

Notice that, for non-celiac-related haplotypes, DQ-α chain presents a glycine on residue 52. On the other hand, celiac-associated haplotypes have an arginine in residue 52, a positive charged residue that allows a salt bridge to be formed with negatively charged glutamic acid residue of gliadin peptides. Thus, the enhanced HLA-DQ heterodimer affinity for gliadin peptide can be explained for HLA-DQ2 and -DQ8 haplotypes. Notice that DQ2-positive presents a deletion on residue α-53 which is responsible of shifting the short α-helix towards the long α-helical stretch of α-chain.

α-22

Hidden Markov Models for HLA-DQ-α chain for residues 20-23. For total HMM, all described HLA-DQA protein sequences were used. For DQ2- and DQ8-positive, total MSA was curated as follows: DQ2-positive contains HLA-DQA1*0501 or *0505 sequences and DQ8-positive HLA-DQA1*03.

Through a network of water-mediated hydrogen bonds, the phenolic OH of Tyr-22 bonds to the carbonyl oxygen of the peptide residue in position 4 and presumably contributes to selective presentation of αI-gliadin. Interestingly, not celiac-associated HLA presents a Phe residue at the corresponding position.

β-71

Hidden Markov Models for HLA-DQ-α chain for residues 69-72. For total HMM, all described HLA-DQA protein sequences were used. For DQ2- and DQ8-positive, total MSA was curated as follows: DQ2-positive contains HLA-DQA1*0501 or *0505 sequences and DQ8-positive HLA-DQA1*03.

Particularly striking is a unique positive electrostatic region between the P4 and P6/P7 pockets in DQ2 caused by Lysβ71. DQ8 contains a threonine at β71 and has an overall neutral electrostatic potential in this region. Consistent with the structural differences, there is no data available indicating that the αI-gliadin epitope is recognized in the context of DQ8 in celiac disease patients. Notice that, for celiac-associated genotypes, residue 70 is an Arg that can form a salt bridge. On the other hand, for non-celiac-associated genotypes, residue 70 is an uncharged Gly.

These mentioned positions are particularly interesting since the P6 Glu of αI-gliadin, which is formed by tTG-catalyzed deamidation, participates in an extensive hydrogen-bonding network. Although the DQ2-specific residue, Lys-β71, plays a key role in stabilizing this residue, the x-ray structure reveals a surprisingly complex network of non-covalent interactions. If Gln were present at this position instead of Glu (i.e., the non-deamidated native gliadin peptide), this intricate hydrogen-bonding network would be disrupted. Also, the Gln carbonyl oxygen is a less attractive hydrogen-bond acceptor than the negatively charged Glu carboxylate oxygen.

In vitro binding assay shows that the deamidated αI-gliadin has a 25-fold higher affinity compared with the non-deamidated counterpart, which may be partially due to having a more potent hydrogen-bond acceptor that can form additional protein- to-protein hydrogen bonds, resulting in a more stable gliadin–DQ2 complex.

Hydrogen-bonding network in the epitope-binding site of DQ2. αI-gliadin is shown in yellow (C, yellow; N, blue; O, red), and HLA-DQ2 α- and β-chains are shown in gray (C, gray; N, blue; O, red). Gray spheres represent water molecules. Atom-to-atom distances are given in Å. Extracted from Kim, C.-Y., Quarsten, H., Bergseng, E., Khosla, C., & Sollid, L. M. in Proceedings of the National Academy of Sciences 2004; 101(12): 4175–4179.