Effects of the regulatory ligands calcium and GTP on the thermal stability of tissue transglutaminase

Cervellati, Carlo; Montin, Katy; Squerzanti, Monica; Mischiati, Carlo; Ferrari, Carlo; Spinozzi, Francesco; Mariani, Paolo; Amenitsch, Heinz; Bergamini, Carlo M.; Lanzara, Vincenzo

doi:10.1007/s00726-011-0963-6

Cited by 3 publications

(2 citation statements)

References 40 publications

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“…It therefore remains unclear how this extracellular TG2 is activated and why, even if on the surface of macrophages is present much TG2, only part of it is active. This partial activation does not fit with the well-known calciumdependent mechanism, so deeply investigated on the purified enzyme (Cervellati et al 2012). In the model with the soluble enzyme, the interaction of calcium with its binding sites on the closed TG2 structure destabilizes the binding of GTP and gives rise to a massive activation of the enzyme with exposure of the active site (Bergamini et al 2011).…”

Section: Discussionmentioning

confidence: 95%

Involvement of cell surface TG2 in the aggregation of K562 cells triggered by gluten

et al. 2016

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Gluten-induced aggregation of K562 cells represents an in vitro model reproducing the early steps occurring in the small bowel of celiac patients exposed to gliadin. Despite the clear involvement of TG2 in the activation of the antigen-presenting cells, it is not yet clear in which compartment it occurs. Herein we study the calcium-dependent aggregation of these cells, using either cell-permeable or cell-impermeable TG2 inhibitors. Gluten induces efficient aggregation when calcium is absent in the extracellular environment, while TG2 inhibitors do not restore the full aggregating potential of gluten in the presence of calcium. These findings suggest that TG2 activity is not essential in the cellular aggregation mechanism. We demonstrate that gluten contacts the cells and provokes their aggregation through a mechanism involving the A-gliadin peptide 31-43. This peptide also activates the cell surface associated extracellular TG2 in the absence of calcium. Using a bioinformatics approach, we identify the possible docking sites of this peptide on the open and closed TG2 structures. Peptide docks with the closed TG2 structure near to the GTP/GDP site, by establishing molecular interactions with the same amino acids involved in stabilization of GTP binding. We suggest that it may occur through the displacement of GTP, switching the TG2 structure from the closed to the active open conformation. Furthermore, docking analysis shows peptide binding with the β-sandwich domain of the closed TG2 structure, suggesting that this region could be responsible for the different aggregating effects of gluten shown in the presence or absence of calcium. We deduce from these data a possible mechanism of action by which gluten makes contact with the cell surface, which could have possible implications in the celiac disease onset.

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Section: Discussionmentioning

confidence: 95%

Involvement of cell surface TG2 in the aggregation of K562 cells triggered by gluten

et al. 2016

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“…The activity was measured with a filter paper assay that employed radioactive putrescine and dimethylcasein as the amine and protein acceptor substrates, as in ref. [25], at saturating (5 mM) and at sub-saturating (0.5 mM) concentrations of free calcium. In this last instance, we also included parallel assays in the presence of 0.2 mM Ca.GTP in order to assess the differential sensitivity to the effects of ligands [26], as discussed further on.…”

Section: Methodsmentioning

confidence: 99%

Changes in Protein Expression in Two Cholangiocarcinoma Cell Lines Undergoing Formation of Multicellular Tumor Spheroids In Vitro

et al. 2015

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Epithelial-to-Mesenchymal Transition (EMT) is relevant in malignant growth and frequently correlates with worsening disease progression due to its implications in metastases and resistance to therapeutic interventions. Although EMT is known to occur in several types of solid tumors, the information concerning tumors arising from the epithelia of the bile tract is still limited. In order to approach the problem of EMT in cholangiocarcinoma, we decided to investigate the changes in protein expression occurring in two cell lines under conditions leading to growth as adherent monolayers or to formation of multicellular tumor spheroids (MCTS), which are considered culture models that better mimic the growth characteristics of in-vivo solid tumors. In our system, changes in phenotypes occur with only a decrease in transmembrane E-cadherin and vimentin expression, minor changes in the transglutaminase protein/activity but with significant differences in the proteome profiles, with declining and increasing expression in 6 and in 16 proteins identified by mass spectrometry. The arising protein patterns were analyzed based on canonical pathways and network analysis. These results suggest that significant metabolic rearrangements occur during the conversion of cholangiocarcinomas cells to the MCTS phenotype, which most likely affect the carbohydrate metabolism, protein folding, cytoskeletal activity, and tissue sensitivity to oxygen.

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Transglutaminase 2 and phospholipase A2 interactions in the inflammatory response in human Thp-1 monocytes

et al. 2013

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Several experimental approaches have demonstrated that transglutaminase 2 (TG2) increased activity is involved in monocyte activation and inflammatory response. Preliminary results also demonstrate a TG-mediated post-translational modification of phospholipase A2 (PLA2), which catalyzes the release of arachidonic acid from its lipid storage sites. The control of PLA2-mediated production of eicosanoids has been found to be of great benefit for inflammatory disease treatment. However, the identification of the mechanisms of PLA2 activation is a very complex issue, because of the presence of multiple PLA2 forms. The aim of this study was to characterize the interactions between TG2 and sPLA2 in LPS-stimulated THP-1 cells, which were treated with TPA to induce early differentiated macrophage-type model. We demonstrated that increases in TG2 enzyme activity and protein expression may be considered an early event in monocyte/macrophage activation by LPS. Under these conditions, TG2 protein was co-immunoprecipitated with PLA2 by monoclonal antibody directed against the secretory form of the enzyme (sPLA2-V). Concomitantly, the PLA2 enzyme activity increased in TPA-treated cells exposed to LPS; these high levels of enzyme activity were significant reduced by R283, a site-specific inhibitor of TG2. Moreover, confocal laser scanning microscopy analysis of double-immunostained cytochemical specimens confirmed a co-localization of BAPA-labeled proteins and sPLA2-V in LPS-treated cells. These findings give evidence of a complex TG2/sPLA2-V, suggesting the possibility that sPLA2-V is a substrate for TG2. These results demonstrated that TG2 increases produced a sustained activation of PLA2 activity, suggesting a functional interaction between these enzymes in the regulation of inflammatory response.

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Effects of the regulatory ligands calcium and GTP on the thermal stability of tissue transglutaminase

Cited by 3 publications

References 40 publications

Involvement of cell surface TG2 in the aggregation of K562 cells triggered by gluten

Involvement of cell surface TG2 in the aggregation of K562 cells triggered by gluten

Changes in Protein Expression in Two Cholangiocarcinoma Cell Lines Undergoing Formation of Multicellular Tumor Spheroids In Vitro

Transglutaminase 2 and phospholipase A2 interactions in the inflammatory response in human Thp-1 monocytes

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