Isozyme-specific comprehensive characterization of transglutaminase-crosslinked substrates in kidney fibrosis

Abstract

Chronic kidney disease is characterized by prolonged decline in renal function, excessive accumulation of ECM, and progressive tissue fibrosis. Transglutaminase (TG) is a crosslinking enzyme that catalyzes the formation of covalent bonds between glutamine and lysine residues, and is involved in the induction of renal fibrosis via the stabilization of ECM and the activation of TGF-β1. Despite the accumulating evidences indicating that TG2 is a key enzyme in fibrosis, genetic knockout of TG2 reduced by only 50% the elevated protein crosslinking and fibrous protein in renal fibrosis model, whereas treatment with TG inhibitor almost completely reduced these levels. Here, we also clarified the distributions of TG isozymes and their in situ activities and identified the isozyme-specific crosslinked substrates for both TG1 and TG2 in fibrotic kidney. We found that TG1 activity was markedly enhanced in renal tubular epithelium and interstitial areas, whereas TG2 activity increased only in the extracellular space. In total, 47 and 67 possible candidates were identified as TG1 and TG2 substrates, respectively, only in fibrotic kidney. Among them, several possible substrates related to renal disease and fibrosis were identified. These findings provide novel insights into the mechanisms of renal fibrosis through the targeting of isozyme-specific TG substrates.

Amelioration of paraquat-induced pulmonary fibrosis in mice by regulating miR-140-5p expression with the fibrogenic inhibitor Xuebijing

“…47 TG2 also enhances fibrillar fibronectin production and maturation in an enzyme-independent manner. [48][49][50] TG2 is upregulated in PQ-induced lung fibrosis mouse models and promotes fibrosis by increasing MMP levels and Smad2, Smad3, TGF-β1, and β-catenin phosphorylation. However, XBJ treatment downregulated TG2 expression; decreased MMP; phosphorylated Smad2, Smad3, TGF-β1, and β-catenin levels; and ameliorated pulmonary fibrosis (Figure 8).…”

Spotlight on the Transglutaminase 2-Heparan Sulfate Interaction

“…Intracellular TG2 is inhibited by guanine and adenine nucleotides and low calcium level (0.1 μmol·L −1 ); increases in intracellular calcium (0.5–1.5 mmol·L −1 ) or the concentration of calcium in the extracellular environment activate TG2 transamidation, as recently reviewed [5]. Substrates of TG2 transamidation have been collected in the TRANSDAB database () [6] (up to 2010), but further interactors and numerous substrates have been revealed in recent proteomic studies [7,8,9]. Based on the conformational heparin binding site of TG2 [10], discussed in this review (Section 6), heparan sulfate (HS) elements of proteoglycans may represent a further level of control of TG2 transamidation.…”

Biocatalysis by Transglutaminases: A Review of Biotechnological Applications

“…Hitomi’s group has developed important probes for the identification of specific substrates of TG family members, with applications in liver and kidney disease [ 84 , 89 ]. The mechanism of externalisation of TG2 from cells to reach the ECM is an unconventional pathway that has fascinated many research groups.…”

“…Research in this field is ongoing, as a better knowledge of TGs specific substrates would further clarify their role in both physiology and disease. Furthermore, multiple TGs transamidation substrates have been exploited in a variety of applications, such as assay systems for in situ visualisation of TG activity [ 109 ], identification of endogenous targets of TGs in cells and tissues [ 84 , 89 , 110 ], and TGs-mediated bio-conjugation of proteins. Hence, a strong interest in finding novel and more specific TGs substrates is still alive in this area of research.…”