Transglutaminase-2 and Celiac Disease

Celiac disease is an autoimmune disease that has become increasingly prevalent in recent years.   It is an immune reaction that occurs with gluten, which is a protein found in wheat, barley, and malt.  This reaction causes inflammation, which damages the small intestine.  You may be thinking that wheat, malt, and barley are in everything!  So, what do people who suffer from celiac eat?  As someone who suffers from celiac, I can tell you the transition to a gluten free diet wasn’t easy, but luckily there are a lot of good, gluten free alternatives that have been developed.  Even though celiac can be helped with dietary accommodation, it is still important to try to understand what is causes the autoimmune response in order to potentially develop a treatment for celiac disease someday.  Unfortunately, it is not well understood how the autoreactive B cells become activated in celiac, but it is known that celiac disease is associated with autoantibodies produced that are specific for the transglutaminase-2 (TG2) enzyme.  In the paper Transglutaminase 2-Specific Autoantibodies in Celiac Disease Target Clustered, N-Terminal Epitopes Not Displayed on the Surface of Cells, the authors Iversen et al. investigated the mechanism that controls the formation of the TG2 autoantibodies.

Transglutaminase-2 is an enzyme that is involved in the deamination of glutamine residues, during them into glutamic acid.  This process increases their affinity for the HLA molecules associated with the disease, thus increasing the reactivity of gluten.  The anti-TG2 autoantibodies are an important marker of celiac disease, but it is unknown how these autoantibodies are contributing the disease.  In the lab’s previous research, they have identified various monoclonal (target 1 particular antigen) TG2-reactive antibodies (mAbs), and they have found that the antibodies target distinct (but close or overlapping) regions.  They define these regions as epitopes 1-4 throughout the paper. 

           

In order to determine the specificity of the mAbs, they stained the small intestinal tissue sections with immunofluorescent dye in either wild-type or mice deficient in TG2.  There was only fluorescence (indicating mAb binding) in the tissues containing TG2, which suggests that the mAbs are highly specific to TG2.  They also attempted to determine whether the mAbs would react with other members of the transglutaminase family, TG3 and TG6, which have also been associated with celiac disease.  They did not find any reactivity, which indicated that autoantibodies against other transglutaminases in celiac disease are created in dependently than the anti-TG2 autoantibodies.

TG2 has both an open and closed conformation.  It was thought that the epitopes on TG2 targeted by the autoantibodies were conformational, it would be expected that the mAbs would differ in binding strength to the two conformations.  Using an ELISA assay, the authors used the natural conformational regulators of TG2, Ca²⁺ (open conformation) and GTP (closed conformation) to measure the binding strength of the mAbs in each of the conformations.  The binding affinity of the mAbs was increased by Ca²⁺, but decreased by GTP.  This suggests that in celiac disease the open conformation of TG2 is the one that is targeted by the autoantibodies.

In a recent study done by Simon-Vecsei et al. , they identified an epitope targeted by celiac disease serum autoantibodies.  The authors wanted to know if their mAbs would react with the newly discovered epitope.  They constructed a triple mutation in the epitope and measured the binding strength with the mAbs by ELISA.  These mutations resulted in loss in reactivity for the mAbs in the epitope 2 group and the epitope 3 group.  This suggests that while this epitope is one of the major targets of autoantibodies in celiac disease, it is not the only one since the mAbs in the epitope 1 and 4 groups do not interact with it. 

TG2 is found both in the cytosol and on the cell surface.  The authors wanted to determine how the mAbs interact with TG2 in the different locations.  When TG2 is expressed on the cell surface, the mAbs did not react with it.  This indicates that the epitopes are hidden when TG2 is bound to the surface.  When they tested TG2 inside cells, they found that all the mAbs did react with TG2.  They also tested the cytosolic TG2 for binding with polyclonal sera (antibodies from the blood) of a celiac patient and found that antibodies also recognize TG2. 

The results from their experiments supported their original model for autoreactive B cell activation.  They believe that the autoantibodies bound to TG2 are bound by the BCR on B cells.  This allows B cells to uptake TG2-gluten complexes, which they process and present as gluten peptides for CD4+ T cells.  The epitopes of TG2 that they identified overlap in the N-terminal end of the protein.  These epitopes are hidden when TG2 is bound to the cell surface, so they hypothesize that the B cells recognizing this region escape negative selection in this way.  While recognizing these epitopes is a good start to developing a treatment for celiac disease, there is still a long way to go.  It is clear that there are many different epitopes in the N-terminal region of TG2 that are recognized by the autoantibodies, and some of the autoantibodies present may not have even been identified in this study.  Also, the anti-TG2 autoantibodies do not cross react with other transglutaminases that cause celiac disease, so that is an additional suite of autoanitibodies and their respective epitopes that have yet to be identified.  Obviously, there are other autoimmune diseases out there that may deserve more attention, and I know that having a gluten free diet isn’t the worst thing that could ever happen to others or myself.  However, I think that celiac disease is really interesting, and maybe understanding this disease will help us understand other autoimmune disease.

Main paper:  Rasmus Iversen, Roberto Di Niro, Jorunn Stamnaes, Knut E. A. Lundin, Patrick C. Wilson and Ludvig M. Sollid. Transglutaminase 2−Specific Autoantibodies in Celiac Disease Target Clustered, N-Terminal Epitopes Not Displayed on the Surface of Cells. J Immunol 2013; 190:5981-5991.

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