An overview of the first 50 years of transglutaminase research

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Free and protein-conjugated polyamines in mouse epidermal cells. Effect of high calcium and retinoic acid.

We have investigated polyamine metabolism in primary cultures of mouse epidermal cells. These cells, which grow at low Ca2+ levels as a monolayer with characteristics of basal cells, terminally differentiate when the extracellular Ca2+ level is raised above 1 mM. The cellular levels of free polyamines were measured, and, after incubation of cell cultures with [3H]putrescine, the distribution of label in both acid-soluble and acid-insoluble cellular components was examined. Free polyamine levels were reduced in cells induced to differentiate. Treatment with retinoic acid, which prevents differentiation and causes increased proliferation, resulted in an increase in free putrescine. Upon adjustment of the calcium concentration to a level that induces differentiation, the enzyme transglutaminase was activated, and a concomitant increase in the level of both protein-bound mono- and bis-gamma-glutamyl derivatives of putrescine and spermidine was observed. Isolation of a material of apparent molecular weight about 6000 which contains only mono-gamma-glutamylpolyamines and the finding of both mono- and bis-gamma-glutamylpolyamines in the protein fraction containing cornified cell envelopes provided the basis for speculation on polyamines in envelope formation. Our data suggest that polyamines play a role during epidermal cell differentiation through transglutaminase-mediated post-translational modification

Gradual Exposure to Salinity Improves Tolerance to Salt Stress in Rapeseed (Brassica napus L.)

Soil salinity is considered one of the most severe abiotic stresses in plants; plant acclimation to salinity could be a tool to improve salt tolerance even in a sensitive genotype. In this work we investigated the physiological mechanisms underneath the response to gradual and prolonged exposure to sodium chloride in cultivars of Brassica napus L. Fifteen days old seedlings of the cultivars Dynastie (salt tolerant) and SY Saveo (salt sensitive) were progressively exposed to increasing soil salinity conditions for 60 days. Salt exposed plants of both cultivars showed reductions of biomass, size and number of leaves. However, after 60 days the relative reduction in biomass was lower in sensitive cultivar as compared to tolerant ones. An increase of chlorophylls content was detected in both cultivars; the values of the quantum eciency of PSII photochemistry (FPSII) and those of the electron transport rate (ETR) indicated that the photochemical activity was only partially reduced by NaCl treatments in both cultivars. Ascorbate peroxidase (APX) activity was higher in treated samples with respect to the controls, indicating its activation following salt exposure, and confirming its involvement in salt stress response. A gradual exposure to salt could elicit dierent salt stress responses, thus preserving plant vitality and conferring a certain degree of tolerance, even though the genotype was salt sensitive at the seed germination stage. An improvement of salt tolerance in B. napus could be obtained by acclimation to saline conditions

The Role of Tissue Transglutaminase in Cancer Cell Initiation, Survival and Progression

Tissue transglutaminase (transglutaminase type 2; TG2) is the most ubiquitously expressed member of the transglutaminase family (EC 2.3.2.13) that catalyzes specific post-translational modifications of proteins through a calcium-dependent acyl-transfer reaction (transamidation). In addition, this enzyme displays multiple additional enzymatic activities, such as guanine nucleotide binding and hydrolysis, protein kinase, disulfide isomerase activities, and is involved in cell adhesion. Transglutaminase 2 has been reported as one of key enzymes that is involved in all stages of carcinogenesis; the molecular mechanisms of action and physiopathological effects depend on its expression or activities, cellular localization, and specific cancer model. Since it has been reported as both a potential tumor suppressor and a tumor-promoting factor, the role of this enzyme in cancer is still controversial. Indeed, TG2 overexpression has been frequently associated with cancer stem cells' survival, inflammation, metastatic spread, and drug resistance. On the other hand, the use of inducers of TG2 transamidating activity seems to inhibit tumor cell plasticity and invasion. This review covers the extensive and rapidly growing field of the role of TG2 in cancer stem cells survival and epithelial⁻mesenchymal transition, apoptosis and differentiation, and formation of aggressive metastatic phenotypes

Transglutaminase 2, a double face enzyme

During the years Amino Acids have dedicated much attention to the transglutaminase (Tgase) field and this special issue is therefore the fifth of a series. In a previous occasion, we celebrated the 50 years from transglutaminase discovery with the editorial “An overview of the first 50 years of transglutaminase research” (Beninati et al. 2009). Following ideally from that point, we have now chosen as general theme for this issue the multiple roles played by type 2 transglutaminase as a multi-face protein. Most people are accustomed to analyze events and facts by taking into account objects and their opposite to get the whole, as in the case of light and darkness, day and night, or black and white as within a chessboard. This is also the case for type 2 transglutaminase (Tgase2): to get the complete picture we must examine both sides of the problem, the protein with its opposite activities, the involvement in cells and tissues leading either to cell growth/differentiation or conversely to cell death/atrophy, and their implications in health protection and pathology. The aim in launching this special issue was to contribute to settle these topics and we introduce now these general concepts dividing ideally the path in the steps mentioned above

Spotlight on the transglutaminase 2 gene: A focus on genomic and transcriptional aspects

The type 2 isoenzyme is the most widely expressed transglutaminase in mammals displaying several intra- and extracellular activities depending on its location (protein modification, modulation of gene expression, membrane signalling and stabilization of cellular interactions with the extracellular matrix) in relation to cell death, survival and differentiation. In contrast with the appreciable knowledge about the regulation of the enzymatic activities, much less is known concerning its inducible expression, which is altered in inflammatory and neoplastic diseases. In this context, we first summarize the gene’s basic features including single-nucleotide polymorphism characterization, epigenetic DNA methylation and identification of regulatory regions and of transcription factor-binding sites at the gene promoter, which could concur to direct gene expression. Further aspects related to alternative splicing events and to ncRNAs (microRNAs and lncRNAs) are involved in the modulation of its expression. Notably, this important gene displays transcriptional variants relevant for the protein’s function with the occurrence of at least seven transcripts which support the synthesis of five isoforms with modified catalytic activities. The different expression of the TG2 (type 2 transglutaminase) variants might be useful for dictating the multiple biological features of the protein and their alterations in pathology, as well as from a therapeutic perspective

A protein produced by a monocytic human cell line can induce apoptosis on tumor cells

A serum-free medium conditioned by U937, a human cell line of monocyte/macrophage origin, was found capable of inducing apoptosis on exponentially growing U937 cells themselves (autocrine suicide). The apoptosis-inducing agent is a macromolecule and possibly a protein (SKT factor), with a relative molecular mass in the range of 18-25 kDa. All human tumor cell lines examined have been induced to apoptosis with high efficiency, whereas non transformed human lymphocytes and monocytes are insensitive to the apoptosis-inducing activity; moreover, partially differentiated U937 are not killed but induced to full maturation. These observations suggest that the SKT factor could possibly be a cytokine with a specific cytotoxic tropism, that resembles in many respects the cytokine tumor necrosis factor (TNF), even though no TNF is detectable in the conditioned medium

Spermidine delays eye lens opacification in vitro by suppressing transglutaminase-catalyzed crystallin cross-linking

A Ca(2+)-dependent TG activity, identified in the eye lens of several mammalian species, has long been implicated in cataract formation. The precise mechanism of the involvement of this enzyme in this process remains unclear. The purpose of this work was to investigate the modulatory effect of polyamines on TG activity during rabbit eye lens in vitro opacification. We observed, in an in vitro Ca(2+)-induced cataract model, a rapid decrease of the endogenous levels of SPD with the progression of opacification, paralleled by an increase of crystallin cross-linking by bis(γ-glutamyl)SPD. This pattern was reversed adding exogenous SPD to the incubation medium. Indeed, endogenous SPD levels were restored and cross-linking by bis(γ-glutamyl)SPD were drastically reduced. Surprisingly, under this experimental condition, the loss of transparency of lens was delayed. We found that exogenous SPD incubation led to a remarkable increase of mono(γ-glutamyl)SPD, likely responsible of the inhibition of cross-linking of lens crystallins and of the transparency persistence

A Central Contribution of TG2 Activity to the Antiproliferative and Pro-Apoptotic Effects of Caffeic Acid in K562 Cells of Human Chronic Myeloid Leukemia

Caffeic acid (CA) has shown antitumor activity in numerous solid and blood cancers. We have recently reported that CA is active in reducing proliferation and triggering apoptosis in both Imatinib-sensitive and resistant Chronic Myeloid Leukemia (CML) cells. Tissue transglutaminase type 2 (TG2) enzyme is involved in cell proliferation and apoptosis of numerous types of cancer. However, its activity has different effects depending on the type of tumor. This work investigated the possible involvement of TG2 activation in the triggering of CA-dependent anticancer effects on the K562 cell line, which was studied as a model of CML. CA-dependent changes in TG2 activity were compared with the effects on cell proliferation and apoptosis. The use of N-acetylcysteine (NAC), an antioxidant molecule, suggested that the antiproliferative effect of CA was due to the increase in reactive oxygen species (ROS). The use of a TG2 inhibitor showed that TG2 activity was responsible for the increase in ROS generated by CA and reduced both caspase activation and triggering of CA-dependent apoptosis. The knocking-down of TGM2 transcripts confirmed the crucial involvement of TG2 activation in CML cell death. In conclusion, the data reported, in addition to ascertaining the important role of TG2 activation in the antiproliferative and pro-apoptotic mechanism of CA allowed us to hypothesize a possible therapeutic utility of the molecules capable of triggering the activation pathways of TG2 in the treatment of CML

Organ Culture Model of Liver for the Study of Cancer Treatment for Hepatocellular Carcinoma

Abstract: The liver, the largest organ of the human body, is a multifunctional organ with various metabolic activities that plays a fundamental role in maintaining the body and in sustaining life. Although the liver has great regenerative capacity and recovery, the damage caused by chronic diseases such as cancer or viral infections can lead to permanent loss of liver function. Studies on the mechanism of liver disease, have focused on the selection of cell and tissue culture techniques, including strategies based on in vitro models. The organ culture is a promising tool for the study of liver diseases, because it can mimic the complex of the microenvironment in vivo using a three-dimensional model of human liver tissue. These models allow a better study of the specific functions of the liver. In this context, we have analyzed the development of a hepatocarcinoma, obtained by inoculating a murine hepatocarcinoma cell line, Hepa 1/A1s, in the liver of 10 mice of the strain C57BL / 6. After 20 days from the inoculation, the portion of liver invaded by the tumor was removed from the animals and cultured. A group of 5 liver explants were used as a control and other 5 explants were cultured for 4 weeks in a complete medium containing 10% Citozym, a food supplement with reported antioxidant properties. The cancer-invaded hepatic lobes, treated with Citozym, showed a clear reduction of the weight and the volume of the hepatic tumors, when compared with the control explants