Country：Japan

Country：Japan

Country：Japan

Authorship：Lead author,　Corresponding author   Language：English   Publishing type：Research paper (scientific journal)  

Transglutaminases (TGs) are a protein family that catalyzes isopeptide bond formation between glutamine and lysine residues of various proteins. There are eight TG isozymes in humans, and each is involved in diverse biological phenomena due to their characteristic distribution. Abnormal activity of TG1 and TG2, which are major TG isozymes, is believed to cause various diseases, such as ichthyosis and celiac disease. To elucidate TGs' mechanisms of action and develop new therapeutic strategies, it is essential to develop bioprobes that can specifically examine the activity of each TG isozyme, which has not been sufficiently studied. We previously have identified several substrate peptide sequences containing Gln residues for each isozyme and developed a method to detect isozyme-specific activities by incorporating a labeled substrate peptide into lysine residues of proteins. We prepared the fluorescein isothiocyanate (FITC)-labeled Gln substrate peptide (FITC-K5 and FITC-T26) and Rhodamine B-labeled Lys substrate peptide (RhoB-Kpep). Each TG reaction specifically cross-linked these probe pairs, and the proximity of FITC and Rhodamine B significantly decreased the fluorescence intensity of FITC depending on the concentration and reaction time of each TG. In this study, we developed a peptide-based biosensor that quickly and easily measures TG isozyme-specific activity. This probe is expected to be helpful in elucidating TG's physiological and pathological functions and in developing compounds that modulate TG activity.

DOI： 10.1007/s00726-023-03272-7

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PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：Wiley  

Formation of the human skin epidermis can be reproduced by a three-dimensional (3D) keratinocyte culture system, in which air-exposure is inevitable upon initiation of differentiation. In the continuous submerged culture without air-exposure, even with a differentiation-compatible medium, several keratinocyte-specific proteins were not induced resulting in the formation of aberrant epidermal layers. To clarify the mechanism by which air-exposure promotes keratinocyte differentiation, we performed a comparative analysis on biological properties between submerged and air-liquid interphase culture systems. By transcriptomic analysis, hypoxia-inducible factor (HIF)-related genes appeared to significantly change in these cultured cells. In submerged culture, the transcriptional activity of HIF on its canonical response element was enhanced, while air-exposure treatment drastically reduced the transcriptional activity despite the high HIF protein level. Regulating HIF activity through reagents and genetic manipulation revealed that the reduced but retained HIF-transcriptional activity was essentially involved in differentiation. Furthermore, we showed, for the first time, that artificial supplementation of oxygen in the submerged culture system could restore keratinocyte differentiation as observed in the air-exposed culture. Thus, we mechanistically evaluated how HIF regulates the air-exposure-dependent differentiation of keratinocytes in a 3D culture system.

DOI： 10.1111/febs.16707

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PubMed

Other Link： https://onlinelibrary.wiley.com/doi/full-xml/10.1111/febs.16707

Language：English   Publishing type：Research paper (scientific journal)  

Long-term peritoneal dialysis (PD) is often associated with peritoneal dysfunction leading to withdrawal from PD. The characteristic pathologic features of peritoneal dysfunction are widely attributed to peritoneal fibrosis and angiogenesis. The detailed mechanisms remain unclear, and treatment targets in clinical settings have yet to be identified. We investigated transglutaminase 2 (TG2) as a possible novel therapeutic target for peritoneal injury. TG2 and fibrosis, inflammation, and angiogenesis were investigated in a chlorhexidine gluconate (CG)-induced model of peritoneal inflammation and fibrosis, representing a noninfectious model of PD-related peritonitis. Transforming growth factor (TGF)-β type I receptor (TGFβR-I) inhibitor and TG2-knockout mice were used for TGF-β and TG2 inhibition studies, respectively. Double immunostaining was performed to identify cells expressing TG2 and endothelial-mesenchymal transition (EndMT). In the rat CG model of peritoneal fibrosis, in situ TG2 activity and protein expression increased during the development of peritoneal fibrosis, as well as increases in peritoneal thickness and numbers of blood vessels and macrophages. TGFβR-I inhibitor suppressed TG2 activity and protein expression, as well as peritoneal fibrosis and angiogenesis. TGF-β1 expression, peritoneal fibrosis, and angiogenesis were suppressed in TG2-knockout mice. TG2 activity was detected by α-smooth muscle actin-positive myofibroblasts, CD31-positive endothelial cells, and ED-1-positive macrophages. CD31-positive endothelial cells in the CG model were α-smooth muscle actin-positive, vimentin-positive, and vascular endothelial-cadherin-negative, suggesting EndMT. In the CG model, EndMT was suppressed in TG2-knockout mice. TG2 was involved in the interactive regulation of TGF-β. As inhibition of TG2 reduced peritoneal fibrosis, angiogenesis, and inflammation associated with TGF-β and vascular endothelial growth factor-A suppression, TG2 may provide a new therapeutic target for ameliorating peritoneal injuries in PD.

DOI： 10.1016/j.labinv.2022.100050

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

At the final stages of blood coagulation, fibrinogen is processed into insoluble fibrin by thrombin resulting in fibril-like structure formation. Via further cross-linking reactions between the fibrin gamma subunit by the catalytic action of blood transglutaminase (Factor XIII), this molecule gains further physical stability. Meanwhile, since fibrinogen is expressed in various cells and tissues, this molecule can exhibit other functions apart from its role in blood coagulation. To create a system studying on aberrant coagulation and investigate the physiological functions, using a model fish medaka (Oryzias latipes), we established gene-deficient mutants of fibrinogen gamma subunit protein in parallel with its biochemical analysis, such as tissue distribution pattern and substrate properties. By genetic deletion via genome editing, two distinct mutants displayed retardation of blood coagulation. The mutants showed lower hematocrit with aberrant erythrocyte maturation, which indicates that fibrin deficiency caused severe anemia, and also appeared as a model for investigation of the fibrin function.

DOI： 10.1093/jb/mvac065

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PubMed

Authorship：Lead author,　Corresponding author   Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

During fetal development, the barrier function of the fetal skin is developed under specific conditions for epidermis formation. In keratinocyte differentiation, the well-orchestrated production and modification of various structural proteins are induced. We assessed the epidermal barrier function in different fetal stages by evaluating the enzymatic activity of cross-linking proteins, transglutaminases, and the permeation of fluorescence dye in the stained epidermal sections. During days 15.5-17.5 in gestation, the enzymatic activities in the epidermis appeared to increase significantly; meanwhile, dye permeation was substantially decreased, suggesting the formation of a protective barrier. For the fetal epidermis formation in the earlier stage, unclarified stimulating factors in the amniotic fluid (AF) are possible to promote barrier function by stimulating keratinocyte differentiation. Thus, we performed proteomic spectrometric (MS) analysis on the components in the AF at different fetal stages. Also, we investigated the promotive ability of the components using a cultured keratinocyte differentiation system. According to the MS analysis, the AF components appeared to exhibit stage-specific variations, where possible unique functions have been identified. We also found that adding the AF from each stage to the medium for cultured keratinocytes specifically enhanced the levels of the differentiation markers. These results provide information on the possible role of AF that contains regulatory factors on keratinocyte differentiation.

DOI： 10.1016/j.abb.2021.109003

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PubMed

Authorship：Lead author,　Corresponding author   Language：English   Publishing type：Research paper (scientific journal)  

Transglutaminase 2 (TG2) is a ubiquitously expressed enzyme catalyzing the crosslinking between Gln and Lys residues and involved in various pathophysiological events. Besides this crosslinking activity, TG2 functions as a deamidase, GTPase, isopeptidase, adapter/scaffold, protein disulfide isomerase, and kinase. It also plays a role in the regulation of hypusination and serotonylation. Through these activities, TG2 is involved in cell growth, differentiation, cell death, inflammation, tissue repair, and fibrosis. Depending on the cell type and stimulus, TG2 changes its subcellular localization and biological activity, leading to cell death or survival. In normal unstressed cells, intracellular TG2 exhibits a GTP-bound closed conformation, exerting prosurvival functions. However, upon cell stimulation with Ca2+ or other factors, TG2 adopts a Ca2+-bound open conformation, demonstrating a transamidase activity involved in cell death or survival. These functional discrepancies of TG2 open form might be caused by its multifunctional nature, the existence of splicing variants, the cell type and stimulus, and the genetic backgrounds and variations of the mouse models used. TG2 is also involved in the phagocytosis of dead cells by macrophages and in fibrosis during tissue repair. Here, we summarize and discuss the multifunctional and controversial roles of TG2, focusing on cell death/survival and fibrosis.

DOI： 10.3390/cells10071842

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PubMed

Language：Japanese   Publisher：(一社)日本腎臓学会  

Language：Japanese   Publisher：(一社)日本腎臓学会  

Language：English  

DOI： 10.1093/bbb/zbaa098.

Language：English   Publishing type：Research paper (scientific journal)  

At the last stage of the blood coagulation cascade, thrombin plays a central role in the processing of fibrinogen for the polymerization and in the additional activation of Factor XIII for the stable cross-linking of fibrin. In addition, thrombin carries out possible multiple roles via processing or interaction with various functional proteins. Several studies conducted in order to elucidate additional physiological significance are ongoing. To clarify further significance of thrombin and to establish an associated disease model, we characterized the orthologue gene for medaka (Oryzias latipes), a research model fish. Tissue distribution of medaka prothrombin has been immunotechnically analyzed. Furthermore, thrombin-deficient medaka mutants were viably established by utilizing a genome-editing method. The established gene-deficient mutants exhibited retarded blood coagulation even in the heterozygous fish. Taking advantage of their ease of handling, this specific model is useful for further investigation in medical research areas on human coagulation diseases.

DOI： 10.1093/bbb/zbaa098

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PubMed

Authorship：Lead author,　Corresponding author   Language：Japanese   Publishing type：Research paper (scientific journal)  

The transglutaminase (TGase) family consists of eight isozymes that catalyze Ca2+-dependent crosslink formation between glutamine and lysine residues of proteins. In the pathogenesis of various chronic diseases, among the TGase isozymes, TG2 in particular is upregulated and contributes to a critical role in fibrosis development and progression via the stabilization of extracellular matrix proteins and activation of TGF-β. Although TG2 has been considered a key enzyme in fibrosis, the causative role of TG2 and involvement of other isozymes remain unclear. We have recently developed a comprehensive analysis method targeting the isozyme-specific substrates of TGase in liver and kidney fibrosis. In this review article, we introduce a previously developed method for determining the activity and tissue distribution of TGase and for the detecting and identification of TGase substrates in an isozyme-specific manner. Using our comprehensive analysis method, we newly characterized the overlapping profile data regarding potential substrates of TG1 and TG2 that have been identified in liver and kidney fibrosis to date. Our results obtained by comparing the specificity and similarity of potential TGase substrates between different tissue fibrosis models provide a deeper understanding regarding the specific and common pathways in disease pathogenesis and progression.

DOI： 10.1016/j.ab.2020.113629

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PubMed

Language：Japanese   Publishing type：Research paper (scientific journal)  

By genome analysis, seven homologous genes (orthologues) of human transglutaminases (TGases) have been identified in medaka fish (Oryzias latipes), some of which clearly corresponded to the Factor XIII, TG1, and TG2. The enzymatically active-recombinant proteins for these medaka TGases has been successfully produced in bacteria or baculovirus-infected insect cell systems. Specific antibodies have been prepared and used in immunohistochemical analyses to reveal tissue distribution. Furthermore, gene-deficient medaka mutants for the genes encoding Factor XIII and TG1 have been established with analysis of their phenotypes. Retarded cross-linking of fibrin and higher sensitivity to osmolality are observed in which each gene had been knock-out. In this review, we summarize these biochemical features and the phenotypes of these gene-deficient fish.

DOI： 10.1016/j.ab.2020.113610

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PubMed

Language：Japanese   Publishing type：Research paper (scientific journal)  

Transglutaminases (TGases) are an enzyme family that catalyzes protein cross-linking essential for several biological functions. In the previous studies, we characterized the orthologues of the mammalian TGase family in medaka (Oryzias latipes), an established fish model. Among the human isozymes, tissue-type transglutaminase (TG2) has multiple functions that are involved in several biological phenomena. In this study, we established medaka mutants deficient for the orthologue of human TG2 (OlTGT) using the CRISPR/Cas9 and TALEN systems. Although apparent morphological changes in the phenotype were not observed, movement retardation was found in the mutant fish when evaluated by a tank diving test. Furthermore, comparative immunohistochemistry analysis using in this fish model revealed that OlTGT was expressed at the periventricular layer of the optic tectum. Our findings provide novel insight for the relationship between tissue-type TGase and the nervous system and the associated behavior.

DOI： 10.1093/jb/mvaa038

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PubMed

Language：Japanese   Publishing type：Research paper (scientific journal)  

The skin epidermis functions as a barrier to various external stresses. In the outermost layer, the terminally differentiated keratinocytes result in cornification with tough structure by formation of cornified envelope beneath the plasma membrane. To complete the formation of the cornified envelope, several structural proteins are cross-linked via the catalytic action of transglutaminases (TG1, TG3, TG5, and TG6). The expression and activation of these enzymes are regulated in a tightly coordinated manner during keratinocyte differentiation. We here show the system detecting the activity of the TGases using specific glutamine-donor substrate peptides in a three-dimensional culture system of keratinocytes. In this review, we summarize the roles of the epidermal enzymes and introduce a detection method that will provide a system for evaluating the skin barrier function.

DOI： 10.1016/j.ab.2020.113606

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PubMed

Language：Japanese   Publisher：(一社)日本腎臓学会  

Language：Japanese   Publisher：(公社)日本生化学会  

Language：Japanese   Publisher：(公社)日本生化学会  

Language：Japanese   Publishing type：Research paper (scientific journal)  

During skin formation, particularly during differentiation of keratinocytes, unique post-translational modifications play a role in forming a proteinaceous supermolecule called the cornified envelope (CE), which is necessary for barrier function. Transglutaminases (TGs) are essential enzymes involved in the cross-linking of various keratinocyte structural proteins to complete CE formation. The TG family consists of eight isozymes, with two members, TG1 and TG3, located mainly in the epidermis. In an in vitro three-dimensional (3D) culture system, reconstruction of the epidermis allows cornification of the terminally differentiated keratinocytes. In this study, using isozyme-specific substrate peptides that enable detection of TG activity, we investigated the expression and the activation pattern of each isozyme during differentiation in this culture system. In the differentiating cells, the protein levels, enzymatic activities, as well as localization of TG1 and TG3 exhibited distinct patterns. Specific knockdown of these enzymes by siRNA revealed less cornification, suggesting that each TG contributes to the epidermal formation. In conclusion, we demonstrate the efficiency of the 3D system for studying differentiation-dependent expression and activity of distinct TGs by specific substrate peptides. ENZYME: Transglutaminase, EC2.3.2.13.

DOI： 10.1111/febs.14832

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

The glomerulus primarily comprises mesangial cells, glomerular microvascular endothelial cells, and podocytes. IgA nephropathy is the most common primary glomerulonephritis worldwide and has a risk of progression to end-stage renal disease. IgA nephropathy is characterized by predominant IgA deposition in the glomerular mesangial area, where TG2 is significantly enhanced. Therefore, identification of glomerular TG2 substrates is the first step in elucidating the role of TG2 as a crosslinking enzyme during disease progression. To clarify potential glomerular TG2 substrates, and to establish a procedure for substrate identification, we attempted to identify those molecules using normal mouse glomeruli. Extracts from mouse glomerular and non-glomerular fractions were treated with our established biotin-labeled substrate peptide, which specifically crosslinks to the lysine-donor substrates depending on TG2 activity. Peptide-incorporated proteins were then purified using avidin resin and identified via mass spectrometry. In parallel, we performed the identification using corresponding samples from TG2 knockout mice. Consequently, potential TG2 substrates were separately identified in glomerular and non-glomerular fractions. They were mainly identified as novel TG2 substrates and partly include the well-known substrates. These results potentially provide novel insights into the mechanism underlying IgA nephropathy and may help elucidate the physiological functions of TG2.

DOI： 10.1016/j.abb.2018.10.001

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Web of Science

Language：English   Publishing type：Research paper (scientific journal)  

Most antiaging factors or life span extenders are associated with calorie restriction (CR). Very few of these factors function independently of, or additively with, CR. In this study, we focused on tschimganine, a compound that was reported to extend chronological life span (CLS). Although tschimganine led to the extension of CLS, it also inhibited yeast cell growth. We acquired a Schizosaccharomyces pombe mutant with a tolerance for tschimganine due to the gene crm1. The resulting Crm1 protein appears to export the stress-activated protein kinase Sty1 from the nucleus to the cytosol even under stressful conditions. Furthermore, we synthesized two derivative compounds of tschimganine, α-hibitakanine and β-hibitakanine; these derivatives did not inhibit cell growth, as seen with tschimganine. α-hibitakanine extended the CLS, not only in S. pombe but also in Saccharomyces cerevisiae, indicating the possibility that life span regulation by tschimganine derivative may be conserved across various yeast species. We found that the longevity induced by tschimganine was dependent on the Sty1 pathway. Based on our results, we propose that tschimganine and its derivatives extend CLS by activating the Sty1 pathway in fission yeast, and CR extends CLS via two distinct pathways, one Sty1-dependent and the other Sty1-independent. These findings provide the potential for creating an additive life span extension effect when combined with CR, as well as a better understanding of the mechanism of CLS.

DOI： 10.1111/gtc.12604

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

The generation, maturation and remodelling of the extracellular matrix (ECM) are essential for the formation of alveoli during lung development. Alveoli formation is disturbed in preterm infants that develop bronchopulmonary dysplasia (BPD), where collagen fibres are malformed, and perturbations to lung ECM structures may underlie BPD pathogenesis. Malformed ECM structures might result from abnormal protein cross-linking, in part attributable to the increased expression and activity of transglutaminase 2 (TGM2) that have been noted in affected patient lungs, as well as in hyperoxia-based BPD animal models. The objective of the present study was to assess whether TGM2 plays a causal role in normal and aberrant lung alveolarization. Targeted deletion of Tgm2 in C57BL/6J mice increased septal thickness and reduced gas-exchange surface area in otherwise normally developing lungs. During aberrant lung alveolarization that occurred under hyperoxic conditions, collagen structures in Tgm2-/- mice were partially protected from the impact of hyperoxia, where normal dihydroxylysinonorleucine and hydroxylysylpiridinoline collagen cross-link abundance was restored; however, the lung alveolar architecture remained abnormal. Inhibition of transglutaminases (including TGM2) with cysteamine appreciably reduced transglutaminase activity in vivo, as assessed by Nε -(γ-l-glutamyl)-l-lysine abundance and TGM catalytic activity, and restored normal dihydroxylysinonorleucine and hydroxylysylpiridinoline collagen cross-link abundance under pathological conditions. Furthermore, a moderate improvement in alveoli size and gas-exchange surface density was noted in cysteamine-treated mouse lungs in which BPD was modelled. These data indicate that TGM2 plays a role in normal lung alveolarization, and contributes to the formation of aberrant ECM structures during disordered lung alveolarization.

DOI： 10.1111/febs.14596

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Hepatocellular carcinoma (HCC) is a highly lethal cancer that has a high rate of recurrence, in part because of cancer stem cell (CSC)-dependent field cancerization. Acyclic retinoid (ACR) is a synthetic vitamin A-like compound capable of preventing the recurrence of HCC. Here, we performed a genome-wide transcriptome screen and showed that ACR selectively suppressed the expression of MYCN, a member of the MYC family of basic helix-loop-helix-zipper transcription factors, in HCC cell cultures, animal models, and liver biopsies obtained from HCC patients. MYCN expression in human HCC was correlated positively with both CSC and Wnt/β-catenin signaling markers but negatively with mature hepatocyte markers. Functional analysis showed repressed cell-cycle progression, proliferation, and colony formation, activated caspase-8, and induced cell death in HCC cells following silencing of MYCN expression. High-content single-cell imaging analysis and flow cytometric analysis identified a MYCN+ CSC subpopulation in the heterogeneous HCC cell cultures and showed that these cells were selectively killed by ACR. Particularly, EpCAM+ cells isolated using a cell-sorting system showed increased MYCN expression and sensitivity to ACR compared with EpCAM- cells. In a long-term (>10 y) follow-up study of 102 patients with HCC, MYCN was expressed at higher levels in the HCC tumor region than in nontumor regions, and there was a positive correlation between MYCN expression and recurrence of de novo HCC but not metastatic HCC after curative treatment. In summary, these results suggest that MYCN serves as a prognostic biomarker and therapeutic target of ACR for liver CSCs in de novo HCC.

DOI： 10.1073/pnas.1802279115

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PubMed

Language：Japanese   Publisher：(一社)日本腎臓学会  

Authorship：Lead author,　Corresponding author   Language：Japanese  

Authorship：Lead author   Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Transglutaminase (TG) is an essential enzyme to catalyze cross-linking reactions of epidermal proteins. Recently, we biochemically characterized human skin TG orthologues for medaka (Oryzias latipes), a model fish. By genome editing, gene-modified fishes for the two orthologues were obtained, both of which lack the ordinal enzymes. These fish appeared to exhibit higher susceptibility to osmolality at the period of larvae.

DOI： 10.1080/09168451.2018.1453294

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

The acute phase protein orosomucoid-1 (Orm1) is mainly expressed by hepatocytes (HPCs) under stress conditions. However, its specific function is not fully understood. Here, we report a role of Orm1 as an executer of HPC proliferation. Increases in serum levels of Orm1 were observed in patients after surgical resection for liver cancer and in mice undergone partial hepatectomy (PH). Transcriptome study showed that Orm1 became the most abundant in HPCs isolated from regenerating mouse liver tissues after PH. Both in vitro and in vivo siRNA-induced knockdown of Orm1 suppressed proliferation of mouse regenerating HPCs and human hepatic cells. Microarray analysis in regenerating mouse livers revealed that the signaling pathways controlling chromatin replication, especially the minichromosome maintenance protein complex genes were uniformly down-regulated following Orm1 knockdown. These data suggest that Orm1 is induced in response to hepatic injury and executes liver regeneration by activating cell cycle progression in HPCs.

DOI： 10.1016/j.ebiom.2017.09.008

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PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY  

Web of Science

Language：English   Publishing type：Research paper (scientific journal)  

In the final process of blood coagulation, fibrin molecules are stabilized via a catalytic reaction by Factor XIIIA (FXIIIA), a member of the transglutaminase (TGase) family that catalyzes protein cross-linking reactions. In this study, we characterized the orthologue of this enzyme in medaka (Oryzias latipes), an established model fish in which a coagulation system is also preserved. The recombinant protein of this orthologue enzyme was produced in baculovirus-infected insect cells and used for analysis of its biochemical properties including activation by thrombin proteolysis and calcium dependence of the TGase enzymatic activity. Immunostaining and immunoblotting revealed that medaka FXIIIA is expressed in the kidney, bone, and esophagus in addition to blood cells. Furthermore, a gene-mutant fish was established using the CRISPR/Cas9 system. The loss of FXIIIA expression was validated in the mutants, and phenotypes, such as absence of fibrin cross-linking, were investigated in the established mutant fish.

DOI： 10.1111/febs.14153

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

In the FANTOM5 project, transcription initiation events across the human and mouse genomes were mapped at a single base-pair resolution and their frequencies were monitored by CAGE (Cap Analysis of Gene Expression) coupled with single-molecule sequencing. Approximately three thousands of samples, consisting of a variety of primary cells, tissues, cell lines, and time series samples during cell activation and development, were subjected to a uniform pipeline of CAGE data production. The analysis pipeline started by measuring RNA extracts to assess their quality, and continued to CAGE library production by using a robotic or a manual workflow, single molecule sequencing, and computational processing to generate frequencies of transcription initiation. Resulting data represents the consequence of transcriptional regulation in each analyzed state of mammalian cells. Non-overlapping peaks over the CAGE profiles, approximately 200,000 and 150,000 peaks for the human and mouse genomes, were identified and annotated to provide precise location of known promoters as well as novel ones, and to quantify their activities.

DOI： 10.1038/sdata.2017.112

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PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s00726-016-2322-0.

Authorship：Lead author,　Corresponding author   Language：English   Publishing type：Research paper (scientific journal)  

Transglutaminases (TGs) comprise a protein family in which the members catalyze the formation of isopeptide bonds between glutamine and lysine residues in various proteins. Expression studies on its three major members, FXIII, TG1, and TG2, have been performed in a relatively large number of mammalian tissues in comparison with those on the other isozymes. We previously identified a highly reactive substrate peptide, including glutamine, for each isozyme from a phage display library and developed a method for detecting isozyme-specific activities by incorporating a labeled substrate peptide into lysine residues of proteins. Here, we describe genetically encoded Förster resonance energy transfer (FRET)-based probes composed of each fluorescence protein (Cerulean and EVenus) fused with substrate peptides. The probe pairs, designated as Trac-MTG (His-CerΔ11-LQ/EV-K-His) containing linker and substrate peptide sequence for microbial TG (MTG), increased the EVenus:Cerulean fluorescence intensity ratio by more than 1.5-fold. Furthermore, we demonstrated that Trac-TG1 (His-CerΔ11-K5) and Trac-TG2 (His-CerΔ11-T26) containing substrate peptide sequence for mammalian TGs successfully detected the isozyme-specific activity of TG1 and TG2, respectively. In this study, we developed a rapid and convenient experimental system for measuring the isozyme-specific activity of TGs. The application of these probes for analyses in cells and tissues will be helpful for elucidating the physiological and pathological functions of TGs.

DOI： 10.1007/s00726-016-2322-0

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Transglutaminase is an enzyme family responsible for post-translational modification such as protein cross-linking and the attachment of primary amine and/or deamidation of glutamine-residue in proteins. Medaka (Oryzias latipes), a recently established model fish, has similar functional proteins to those characterized in mammals. Previously, we found the apparent orthologues that correspond to human transglutaminases in medaka. In this study, regarding the medaka orthologue of human tissue-type transglutaminase (OlTGT), recombinant protein was expressed in an active form in bacteria cultured at low temperature. Using the recombinant protein, we biochemically characterized the enzymatic activity and also obtained a monoclonal antibody that specifically recognized OlTGT. Immunochemical analysis revealed that OlTGT was not expressed ubiquitously, unlike its mammalian orthologue, but in primarily limited tissues such as the eye, brain, spinal cord, and gas gland.

DOI： 10.1080/09168451.2016.1256757

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PubMed

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1016/j.bbrc.2016.07.051.

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DOI： 10.1038/cddis.2016.150.

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DOI： 10.1158/1940-6207.CAPR-15-0326.

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DOI： doi: 10.1371/journal.pone.0144194.

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DOI： doi: 10.1371/journal.pbio.1002315.

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DOI： doi: 10.1371/journal.pone.0144176.

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DOI： doi: 10.1038/cddis.2015.339.

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DOI： 10.1016/j.bbrc.2015.10.001.

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DOI： doi:10.1111/j.1440-1746.2011.07009.x

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1002/jcp.22833

Language：English   Publishing type：Research paper (scientific journal)  

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DOI： 10.1369/jhc.2010.957225

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：Japanese  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：Japanese  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Rapid communication, short report, research note, etc. (scientific journal)  

Language：Japanese   Publisher：(一社)日本腎臓学会  

J-GLOBAL

Language：English   Publishing type：Research paper, summary (international conference)   Publisher：WILEY  

Web of Science

J-GLOBAL

Event date： 2022.11

Presentation type：Oral presentation (invited, special)  

Event date： 2022.11 - 2022.12

Language：Japanese   Presentation type：Oral presentation (invited, special)  

Venue：オンライン開催  

Event date： 2022.6

Language：Japanese   Presentation type：Oral presentation (invited, special)  

Venue：ウインクあいち（愛知県産業労働センター）  

Event date： 2018.9

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2018.6

Language：English   Presentation type：Oral presentation (invited, special)  

Country：Japan  

Event date： 2017.12

Language：Japanese   Presentation type：Oral presentation (invited, special)  

Country：Japan  

Event date： 2014.10

Language：English   Presentation type：Oral presentation (invited, special)  

Country：Japan  

Event date： 2014.6 - 2014.7

Language：English   Presentation type：Oral presentation (general)  

Country：Italy  

Event date： 2012.3

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2012.3

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2011.12

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2011.11

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2011.10

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2011.10

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2011.10

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2011.9

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2011.6

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2011.6

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2010.12

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2010.11

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2010.10

Language：English   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2010.7

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2010.7

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2010.5

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2010.5

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2010.2

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2010.2

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2010.2

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2009.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Country：Japan  

Event date： 2009.10

Language：English   Presentation type：Oral presentation (invited, special)  

Country：Japan  

Event date： 2009.10

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2009.10

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2009.10

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2009.10

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2009.9

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2009.6

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Country：Japan  

Event date： 2009.1

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2008.12

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2008.11

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2008.10

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2008.9

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2008.5

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2008.2

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2007.12

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2007.12

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2007.11

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2007.10

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2007.9

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2007.9

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation (invited, special)  

Country：Japan  

Event date： 2006.6

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2006.6

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2006.5

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2005.10

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2005.10

Language：Japanese   Presentation type：Oral presentation (invited, special)  

Country：Japan  

Event date： 2005.5

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2004.12

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2004.10

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2004.10

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2004.9

Language：English   Presentation type：Poster presentation  

Country：Japan  

Event date： 2004.6

Language：Japanese   Presentation type：Oral presentation (invited, special)  

Country：Japan  

Event date： 2004.5

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2003.12

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Event date： 2003.11

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2003.10

Language：Japanese   Presentation type：Symposium, workshop panel (nominated)  

Country：Japan  

Event date： 2002.12

Language：Japanese   Presentation type：Poster presentation  

Country：Japan  

Language：English   Presentation type：Poster presentation  

Author：Other 

Author：Other 

Author：Myself 

Author：Other 

Author：Other 

Author：Other