Transglutaminase In Cells Research Articles

Circadian clock gene disruption in white blood cells of patients with celiac disease

Clock gene disruption has been reported in inflammatory and autoimmune diseases. Specifically, it has been shown that clock gene expression is down-regulated in intestinal tissue and peripheral blood mononuclear cells of patients with inflammatory bowel disease (IBD). We aimed to determine the systemic expression of the circadian clock genes in newly diagnosed untreated, young patients with celiac disease (CeD). We prospectively enrolled patients younger than 20 years old who underwent diagnostic endoscopic procedures either for CeD diagnosis or due to other gastrointestinal complaints, at the pediatric and adult gastroenterology units, the Tel Aviv Sourasky Medical Center from 8/2016–8/2022. Demographic data, anthropometric parameters, and endoscopic macroscopic and microscopic findings were obtained. Blood samples were obtained to determine tissue transglutaminase (tTG) and core clock gene (CLOCK, BMAL1, PER1, PER2, CRY1, CRY2) expression in white blood cells (WBC). Thirty individuals were analyzed (18 with newly diagnosed CeD and 12 controls). Expression of the clock genes CLOCK, BMAL1, CRY2, PER1 and PER2 was significantly reduced in CeD patients compared to controls, while CRY1 did not differ between the groups. In conclusion, newly diagnosed, untreated, young patients with CeD have reduced clock gene expression in WBC compared to controls. These results suggest that, in CeD, the inflammatory response is associated with systemic disruption of clock gene expression, as is manifested in other inflammatory and autoimmune diseases. ClinicalTrials.gov identifierNCT03662646.

Constitutive Differential Features of Type 2 Transglutaminase in Cells Derived from Celiac Patients and from Healthy Subjects.

Type 2 transglutaminase (TG2) is a ubiquitous enzyme able to modify gliadin peptides introduced into the organism through the diet. By means of its catalytic activity, TG2 seems to have an important pathogenetic role in celiac disease (CD), an inflammatory intestinal disease caused by the ingestion of gluten-containing cereals. A strong autoimmune response to TG2 characterizes CD development. Anti-TG2 antibodies specifically derange the uptake of the α-gliadin peptide 31–43 by control, but not by celiac dermal fibroblasts, underlying some different constitutive features regarding TG2 in healthy and celiac subjects. Our aim was to investigate whether these differences depended on a different TG2 subcellular distribution and whether peptide 31–43 differentially regulated TG2 expression and activity in cells of the two groups of subjects. We found that TG2 was more abundantly associated with membranes of celiac fibroblasts than of control cells, in particular with the early endosomal and autophagic compartments. We also found that peptide 31–43 differentially affected TG2 expression and activity in the two groups of cells, activating TG2 more in control than in celiac cells and inducing TG2 expression in celiac cells, but not in control ones. The different TG2 subcellular localization and the different way the peptide 31–43 modulates TG2 activity and availability into control and CD cells suggested that TG2 is involved in the definition of a constitutive CD cellular phenotype, thus having an important and still undefined role in CD pathogenesis.

Intracellular Enzyme-Triggered Assembly of Amino Acid-Modified Gold Nanoparticles for Accurate Cancer Therapy with Multimode.

Multiple amino acid (glutamine and lysine)-modified gold nanoparticles a with pH-switchable zwitterionic surface were fabricated through coordination bonds using ferrous iron (Fe2+) as bridge ions, which are able to spontaneously and selectively assemble in tumor cells for accurate tumor therapy combining enzyme-triggered photothermal therapy and H2O2-dependent catalytic medicine. These gold nanoparticles showed electric neutrality at pH 7.4 (hematological system) to prevent endocytosis of normal cells, which could be positively charged at pH 6.8 (tumor microenvironment) to promote the endocytosis of tumor cells to these nanoparticles, performing great tumor selectivity. After cell uptake, the specific enzyme (transglutaminase) in tumor cells would catalyze the polymerization of glutamine and lysine to cause the intracellular assembly of these gold nanoparticles, resulting in an excellent photothermal property for accurate tumor therapy. Moreover, the Fe2+ ion could decompose excess hydrogen peroxide (H2O2) in tumor cells via the Fenton reaction, resulting in a large amount of hydroxyl radicals (·OH). These radicals would also cause tumor cell damage. This synergetic therapy associating with high tumor selectivity generated an 8-fold in vitro cytotoxicity against tumor cells compared with normal cells under 48 h incubation with 10 min NIR irradiation. Moreover, in vivo data from tumor-bearing nude mice models showed that tumors can be completely inhibited and gradually eliminated after multimode treatment combining catalytic medicine and photothermal therapy for 3 weeks. This system takes advantage of three tumor microenvironment conditions (low pH, enzyme, and H2O2) to trigger the therapeutic actions, which is a promising platform for cancer therapy that achieved prolonged circulation time in the blood system, selective cellular uptake, and accurate tumor therapy in multiple models.

Modulation of histamine H<sub>4</sub> receptor signaling by transglutaminase and histamine transporter in mast cells

Modulation of histamine H<sub>4</sub> receptor signaling by transglutaminase and histamine transporter in mast cells

Protective Effects of Triphala on Dermal Fibroblasts and Human Keratinocytes.

Human skin is body’s vital organ constantly exposed to abiotic oxidative stress. This can have deleterious effects on skin such as darkening, skin damage, and aging. Plant-derived products having skin-protective effects are well-known traditionally. Triphala, a formulation of three fruit products, is one of the most important rasayana drugs used in Ayurveda. Several skin care products based on Triphala are available that claim its protective effects on facial skin. However, the skin protective effects of Triphala extract (TE) and its mechanistic action on skin cells have not been elucidated in vitro. Gallic acid, ellagic acid, and chebulinic acid were deduced by LC-MS as the major constituents of TE. The identified key compounds were docked with skin-related proteins to predict their binding affinity. The IC50 values for TE on human dermal fibroblasts (HDF) and human keratinocytes (HaCaT) were 204.90 ± 7.6 and 239.13 ± 4.3 μg/mL respectively. The antioxidant capacity of TE was 481.33 ± 1.5 mM Trolox equivalents in HaCaT cells. Triphala extract inhibited hydrogen peroxide (H2O2) induced RBC haemolysis (IC50 64.95 μg/mL), nitric oxide production by 48.62 ± 2.2%, and showed high reducing power activity. TE also rescued HDF from H2O2-induced damage; inhibited H2O2 induced cellular senescence and protected HDF from DNA damage. TE increased collagen-I, involucrin and filaggrin synthesis by 70.72 ± 2.3%, 67.61 ± 2.1% and 51.91 ± 3.5% in HDF or HaCaT cells respectively. TE also exhibited anti-tyrosinase and melanin inhibition properties in a dose-dependent manner. TE increased the mRNA expression of collagen-I, elastin, superoxide dismutase (SOD-2), aquaporin-3 (AQP-3), filaggrin, involucrin, transglutaminase in HDF or HaCaT cells, and decreased the mRNA levels of tyrosinase in B16F10 cells. Thus, Triphala exhibits protective benefits on skin cells in vitro and can be used as a potential ingredient in skin care formulations.

Tissue Transglutaminase Expression and Activity in Human Ligamentum Flavum Cells Derived From Thoracic Ossification of Ligamentum Flavum

The study was undertaken to compare the expression and activity of tissue transglutaminase (TG2) in human ligamentum flavum cells derived from ossification of the ligamentum flavum (OLF) and non-OLF patients. To determine whether TG2 is involved in the pathologic process of OLF. OLF is a disease characterized by heterotopic formation of new bone in the flavum ligament. Recently, TG2 is proved to directly promote skeletal matrix mineralization and play an important role in the ossification. TG2 activity is vital to the differentiation of osteoblasts and the formation of mineralization. But whether TG2 is involved in the pathologic process of OLF is unknown. We investigated the relations between TG2 expression and OLF. OLF and non-OLF cells were cultured and osteocalcin, bone morphogenetic protein-2(BMP-2) and TG2 mRNA expressions were assayed by reverse transcription polymerase chain reaction. Meanwhile, alkaline phosphatase activity and calcified nodules were compared between OLF and non-OLF cells. To detect TG2 expression, Western blot and immunohistochemical analysis were carried out, and TG2 activity was compared between OLF and non-OLF cells. Our experiments demonstrated that OLF cells showed osteoblast-like activity and increased mRNA expression of BMP-2. More interesting, compared with non-OLF cells, OLF cells showed elevated expression levels of TG2 mRNA and protein, as well as enzyme activity. TG2 expression and enzyme activity are upregulated in the OLF cells and TG2 may be involved in the pathologic process of OLF.

Type 2 Transglutaminase in Neurodegenerative Diseases: The Mitochondrial Connection

"Tissue" or type 2 Transglutaminase (TG2) is a peculiar multifunctional enzyme able to catalyse Ca(2+)-dependent post-translational modification of proteins, by establishing covalent bonds between peptide-bound glutamine residues and either lysine residues or mono- and poly-amines. In addition, it may act also as a G protein in transmembrane signalling, as a kinase, as a protein disulphide isomerase and as a cell surface adhesion mediator. The vast array of biochemical functions exerted by TG2 characterises and distinguishes it from all the other members of the transglutaminase family. Multiple lines of evidence suggest an involvement of the enzyme in neurodegenerative diseases, such as Huntington's (HD) and Parkinson (PD), and that its inhibition, either via drug treatments or genetic approaches, might be beneficial for the treatment of these syndromes. This review will exploit the recent developments in the comprehension of the role played by type 2 transglutaminase in eukaryotic cells, focusing on the role exerted by TG2 on mitochondrial physiology and on the regulation of cell death pathways at the basis of neurodegenerative diseases.

Tissue Transglutaminase Inhibits Autophagy in Pancreatic Cancer Cells

Elevated expression of tissue transglutaminase (TG2) in cancer cells has been implicated in the development of drug resistance and metastatic phenotypes. However, the role and the mechanisms that regulate TG2 expression remain elusive. Here, we provide evidence that protein kinase Cdelta (PKCdelta) regulates TG2 expression, which in turn inhibits autophagy, a type II programmed cell death, in pancreatic cancer cells that are frequently insensitive to standard chemotherapeutic agents. Rottlerin, a PKCdelta-specific inhibitor, and PKCdelta small interfering RNA (siRNA) down-regulated the expression of TG2 mRNA and protein and induced growth inhibition without inducing apoptosis in pancreatic cancer cells. Inhibition of PKCdelta by rottlerin or knockdown of TG2 protein by a TG2-specific siRNA resulted in a marked increase in autophagy shown by presence of autophagic vacuoles in the cytoplasm, formation of the acidic vesicular organelles, membrane association of microtubule-associated protein 1 light chain 3 (LC3) with autophagosomes, and a marked induction of LC3-II protein, important hallmarks of autophagy, and by electron microscopy. Furthermore, inhibition of TG2 by rottlerin or by the siRNA led to accumulation of green fluorescent protein (GFP)-LC3-II in autophagosomes in pancreatic cancer cells transfected with GFP-LC3 (GFP-ATG8) expression vector. Knockdown of Beclin-1, a specific autophagy-promoting protein and the product of Becn1 (ATG6), inhibited rottlerin-induced and TG2 siRNA-induced autophagy, indicating that Beclin-1 is required for this process. These results revealed that PKCdelta plays a critical role in the expression of TG2, which in turn regulates autophagy. In conclusion, these results suggest a novel mechanism of regulation of TG2 and TG2-mediated autophagy in pancreatic cancer cells.

Cystamine inhibits transglutaminase and caspase-3 cleavage in glutamate-exposed astroglial cells.

Although the precise role of transglutaminase in cell death is unknown, several findings demonstrate that tissue transglutaminase selectively accumulates in cells undergoing apoptosis both in vivo and in vitro. Calcium-dependent transglutaminase reactions are also implicated in several neurodegenerative diseases, including alterations in the release of excitatory amino acids. One prevalent theme in cell damage induced by excitotoxic stimuli in different regions of the CNS is that apoptosis may be executed by intracellular caspase proteases. Furthermore, the presence of functional ion channel-gated receptors in glial cells suggests that also astrocytes can be susceptible to glutamate's toxic effects. In this study, we demonstrated that prolonged exposure to glutamate (100 microM) of cultured astrocytes caused an increase in the expression of tissue transglutaminase (tTG). This effect was prevented by preincubation with GYKI 52466, an antagonist of AMPA/KA receptors. Glutamate exposure also promoted an increase in caspase-3 compared with control cultures. Confocal laser microscopy analysis demonstrated the presence of activated caspase-3 in the cytoplasm as well as in the nucleus. The inhibition of TG-catalyzed reactions by cystamine (1 mM) blocked the activation pathway of caspase-3, with an evident reduction of enzyme cleavage. These results suggest that glutamate increased both TG and caspase-3 in astroglial cells early in the excitotoxin-induced events.

“Tissue” transglutaminase in AIDS

Apoptosis or programmed cell death (PCD) is an active process of cellular self-destruction, essential for embryonic development and maintenance of homeostasis of multicellular organisms. Programmed cell death induction can serve as a defence mechanism of the host against intracellular microbes. Virus infections trigger host cell apoptosis, which can either limit virus production or contribute directly to viral pathogenesis. Several independent laboratories have identified “tissue” transglutaminase (tTG) as a potentially important player of the cell death program(s). This gene is specifically expressed in cells dying during mammalian development as well as in those undergoing apoptosis in various patho-physiological and experimental settings [Eur. J. Cell Biol. 56 (1991) 170; Piacentini, M., Davies, P.J.A., Fesus, L., 1994. Tissue transglutaminase in cells undergoing apoptosis. In: Tomei, L.D., Cope, F.O. (Eds.), Apoptosis II: The molecular basis of apoptosis in disease. Cold Spring Harbor Lab. Press, pp. 143–165.]. This chapter reviews recent studies concerning the expression and the possible role of “tissue” transglutaminase (tTG) in apoptotic cells; particular emphasis is given to its expression in the cell death pathways associated with HIV infection in the immune system. We propose here that the induction of the tTG gene in cells of the immune system, as well as the detection of the isodipeptide ε(γ-glutamyl)lysine in plasma, are useful markers of apoptosis and might make it possible to monitor disease progression in HIV-infected individuals.

Thrombin upregulates tissue transglutaminase in endothelial cells: a potential role for tissue transglutaminase in stability of atherosclerotic plaque.

Atherosclerosis is characterized by thickening of the vessel wall, smooth muscle cell proliferation, macrophage infiltration, and deposition of a fibrin network. Transglutaminases are a family of enzymes catalyzing the formation of stable covalent cross-links between proteins. Here, we show that tissue transglutaminase (tTG) synthesis by human umbilical vein endothelial cells is upregulated by thrombin, the serine protease that causes fibrin formation and many cellular inflammatory effects. Thrombin upregulated tTG 2-fold at the mRNA and protein level. Cellular cross-linking activity was increased to an even greater extent; antibody to tTG neutralized the increased activity. The effect on tTG expression required active thrombin and was mediated mainly through protease-activated receptor-1, a thrombin receptor. Increased tTG antigen and activity were evident in human umbilical vein endothelial cells and extracellular matrix in situ. Thrombin treatment also led to a cellular redistribution of tTG. Normal vessel wall stained positively for tTG in the smooth muscle cells and in the subendothelium. The intensity of staining increased in vessel walls with plaque, where there was a striking increase in tTG in the smooth muscle cells immediately below the plaque. These studies indicate a role for tTG in the stabilization of atherosclerotic plaques and suggest that its local expression can be controlled by thrombin.

Retinoic acid-induced tissue transglutaminase and apoptosis in vascular smooth muscle cells.

Retinoids exert antiproliferative and prodifferentiating effects in vascular smooth muscle cells (SMCs) and reduce neointimal mass in balloon-injured blood vessels. The mechanisms through which retinoids carry out these effects are unknown but likely involve retinoid receptor-mediated changes in gene expression. Here we report the cloning, chromosomal mapping, and biological activity of the retinoid-response gene rat tissue transglutaminase (tTG). Northern blotting studies showed that tTG is rapidly and dose-dependently induced in a protein synthesis-independent manner after stimulation with the natural retinoid all-trans retinoic acid (atRA). The induction of tTG was selective for atRA and its stereoisomers 9-cis and 13-cis RA, because little or no elevation in mRNA expression was observed with a panel of growth factors. Western blotting and immunofluorescence confocal microscopy showed an accumulation of cytosolic tTG protein after atRA stimulation. Radiolabeled cross-linking studies revealed a corresponding elevation in in vitro tTG activity. The increase in tTG activity was reduced in the presence of 2 distinct inhibitors of tTG (monodansylcadaverine and cystamine). atRA-induced tTG mRNA and protein expression were followed by a significant elevation in SMC apoptosis. Such retinoid-induced programmed cell death could be partially inhibited with each tTG inhibitor and was completely blocked when both inhibitors were used simultaneously. These results establish a role for atRA in the sequential stimulation of tTG and apoptosis in cultured SMCs. atRA-mediated apoptosis in SMCs seems to require the participation of active tTG, suggesting a potential mechanistic link between this retinoid-inducible gene and programmed cell death.

Differences in transglutaminase mRNA after polyamine depletion in two cell lines.

Polyamines serve as natural substrates for the transglutaminase that catalyzes covalent cross-linking of proteins and is involved in cellular adhesion and proliferation. This study tests the hypothesis that intracellular polyamines play a role in the regulation of transglutaminase expression in rat small intestinal crypt cells (IEC-6 cell line) and human colon carcinoma cells (Caco-2 cell line). Treatment with alpha-difluoromethylornithine (DFMO; a specific inhibitor of polyamine synthesis) significantly depleted the cellular polyamines putrescine, spermidine, and spermine in both cell lines. In IEC-6 cells, polyamine depletion was associated with a decrease in the levels of transglutaminase mRNA. In Caco-2 cells, however, polyamine depletion significantly increased the levels of transglutaminase mRNA and enzyme activity. In both cell lines, ornithine decarboxylase mRNA levels increased and protooncogene c-myc mRNA decreased in the presence of DFMO. Addition of polyamines to cells treated with DFMO reversed the effect of DFMO on the levels of mRNA for these genes in both lines. There was no significant change in the stability of transglutaminase mRNA between control and DFMO-treated IEC-6 cells. In contrast, the half-life of mRNA for transglutaminase in Caco-2 cells was dramatically increased after polyamine depletion. Spermidine, when given together with DFMO, completely prevented increased half-life of transglutaminase mRNA in Caco-2 cells. These results indicate that 1) expression of transglutaminase requires polyamines in IEC-6 cells but is inhibited by these agents in Caco-2 cells, 2) polyamines modulate transglutaminase expression at the level of mRNA through different pathways in these two cell lines, and 3) posttranscriptional regulation plays a major role in the induction of transglutaminase mRNA in polyamine-deficient Caco-2 cells.

Identification of Cytoplasmic Actin as an Abundant Glutaminyl Substrate for Tissue Transglutaminase in HL-60 and U937 Cells Undergoing Apoptosis

A lysine derivative, 3-[Nalpha[Nepsilon-[2', 4'-dinitrophenyl]-amino-n-hexanoyl-L-lysylamido]-propane-1-ol, a novel amine substrate of transglutaminases, was synthesized and delivered into intact HL-60 and U937 human leukemia cells to probe the function of the intracellular enzyme. The novel substrate compound was covalently incorporated into intracellular proteins in these cells expressing high levels of tissue transglutaminase and undergoing apoptosis following the induction of their differentiation with dimethyl sulfoxide and retinoic acid. Immunoaffinity purification and microsequencing of labeled proteins identified cytoplasmic actin as the main endogenous glutaminyl substrate in these cells. As shown by confocal image analysis, cells revealed distinct labeling of the microfilament meshwork structures by the novel compound as the result of the intracellular action of transglutaminase.

Vitamin A Stimulation of Insulin Secretion

Retinol or retinoic acid is required for insulin release. Retinoids increase transglutaminase activity, and transglutaminase has been implicated in islet insulin release. To examine whether transglutaminase could mediate effects of retinoids on insulin secretion, we measured (i) transglutaminase activity in islets from rats deficient in vitamin A or repleted with retinol or retinoic acid, (ii) transglutaminase activity in RINm5F and INS-1 insulin-secreting cells cultured in retinol or retinoic acid, (iii) mRNA for transglutaminase in RINm5F and INS-1 cells, and (iv) insulin secretion from INS-1 cells in response to retinoic acid. Islets from rats repleted with retinol or retinoic acid showed more than twice the transglutaminase activity of islets from vitamin A deficient rats. Retinoic acid increased RINm5F cells and INS-1 cell transglutaminase activity. Retinol did not increase transglutaminase activity. Transglutaminase mRNA was detected in INS-1 cells but not in RINm5F cells. Retinoic acid increased insulin secretion from INS-1 cells as observed previously in RINm5F cells. In conclusion, retinoic acid increases transglutaminase activity in both rat islets and two insulin-secreting from INS-1 cells. Transglutaminase is a candidate for mediating retinoid-induced changes in insulin secretion.

Colocalization of Tissue Transglutaminase and Stress Fibers in Human Vascular Smooth Muscle Cells and Human Umbilical Vein Endothelial Cells

The subcellular distribution of tissue transglutaminase in human umbilical vein endothelial cells and human arterial and venous smooth muscle cells was examined. Double-immunofluorescence staining of smooth muscle cells and endothelial cells with anti-transglutaminase antisera and rhodamine-tagged phalloidin revealed codistribution of transglutaminase with the stress fibers, with endothelial cells also containing a cytoplasmic pool. This pattern of distribution was confirmed by confocal microscopy. Immunoprecipitation experiments demonstrated that transglutaminase co-immunoprecipitated with myosin in high-molecular-weight complexes, but not with actin, suggesting that the association of transglutaminase with the stress fibers was due to its cross-linking to myosin. About 97% of endothelial cell transglutaminase activity was present in the cytosolic fraction and 3% in the particulate fraction. The detergent-insoluble fraction was practically devoid of activity as measured by the putrescine assay, but was active as evidenced by the covalent cross-linking of 125I-fibronectin. Western blotting with a polyclonal rabbit antiserum raised against human erythrocyte transglutaminase detected high levels of enzyme in endothelial cell cytosol and both detergent-soluble and detergent-insoluble membrane fractions. In contrast, smooth muscle cells contained much less cytosolic transglutaminase, as determined either functionally or antigenically. Furthermore, within the particulate fraction of the smooth muscle cells, most of the enzyme was located in the detergent-insoluble fraction, as assessed by Western blot analysis. Retinoic acid increased the levels of enzyme in the cytosol of all cell types and the increases were correlated with increases in mRNA. Thus, tissue transglutaminase is present in various particulate fractions of vascular smooth muscle cells and endothelial cells and may be present in this cellular fraction by virtue of autocross-linking of the enzyme itself to stress fiber-associated myosin.

Interaction of liposome-associated all-trans-retinoic acid with squamous carcinoma cells.

Because of their antiproliferative and differentiation-inducing properties, retinoids have been used clinically as therapeutic and chemopreventive agents against squamous-cell carcinomas (SCC). As is the case for many therapeutic agents, however, the administration of retinoids is associated with toxic effects. Because encapsulation of certain drugs in lipid vesicles (liposomes) has been shown to result in reduced toxic effects, we studied the in vitro interaction of liposome-encapsulated all-trans-retinoic acid (L-ATRA) with a SCC line (MDA 886Ln) and its multicellular tumor spheroid (MTS) model. Various L-ATRA formulations were tested for incorporation of retinoic acid, toxic effects against human red blood cells, uptake and retention by tumor cells, and antiproliferative effects against SCC. Of the different formulations tested, L-ATRA containing diphosphatidyl palmitoylcholine (DPPC) and stearylamine (SA; 9:1, w/w) showed optimal drug incorporation, high stability, and minimal toxicity toward red blood cells and was highly efficacious in delivering ATRA and, thus, in inhibiting the growth of MDA 886Ln and its MTS model. DPPC: SA L-ATRA inhibited the expression of the enzyme keratinocyte transglutaminase in epidermal cells as effectively as did the free drug. These results suggest that liposomes can serve as an effective carrier system for the delivery of retinoids to SCC.

Regulation of type I and type II transglutaminase in normal human bronchial epithelial and lung carcinoma cells.

In cultured, undifferentiated normal human bronchial epithelial (HBE) cells, transglutaminase activity was localized predominantly in the cytosolic fraction of cell lysates. Upon squamous differentiation, this cytosolic activity declined and was replaced by a 40-fold increase in the activity of particulate (membrane-associated) transglutaminase. Immunoblot analysis demonstrated that the cytosolic transglutaminase was Type II (tissue) transglutaminase and that squamous differentiation shifted gene expression to the Type I (epidermal) transglutaminase. Retinoic acid, an inhibitor of squamous cell differentiation, suppressed the increase in Type I transglutaminase. The decrease in Type II transglutaminase activity was unaffected by retinoic acid. Transforming growth factor-beta 1 (TGF-beta 1) enhanced Type II transglutaminase activity about 10-fold in the undifferentiated cells but did not increase Type I transglutaminase or cholesterol sulfate, two early markers of squamous differentiation. TGF-beta 2 was equivalent to TGF-beta 1 in inducing Type II transglutaminase and in inhibiting the growth of HBE cells. The differentiation-related and TGF-beta-induced changes in transglutaminase activity were reflected in the level of transglutaminase Type I and Type II protein and mRNA. Expression of transglutaminases in lung carcinoma cell lines was variable. No correlation was observed between the expression of Type I transglutaminase and the classification of the cells as squamous cell carcinoma. Several lung carcinoma cell lines exhibited high levels of Type II transglutaminase activity that were increased several-fold by TGF-beta 1 treatment. Retinoic acid was ineffective in altering transglutaminase expression in most cell lines but induced Type II transglutaminase in a time- and dose-dependent manner in NCI-HUT-460 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Retinoids induce tissue transglutaminase in NIH-3T3 cells

We report that all-trans and 13-cis-retinoic acid as well as the synthetic compound CH-55 enhance tissue transglutaminase activity as they increase NIH-3T3 cell adhesiveness. The 4-hydroxyphenylretinamide (4-HPR) with low activity in inducing attachment, lectin binding and growth inhibition also fails to induce tranglutaminase. Thyroxine (Thy), a compound with a response element common to RA, is inactive. The tumor promoter 12-tetradecanoylphorbol-13-acetate (TPA), which increases adhesiveness with different kinetics than RA, failed to enhance tranglutaminase. We conclude that retinoids with biological activity in inducing adhesion, inhibition of growth and increase of lectin binding, are also active in inducing transglutaminase activity.

?Tissue? transglutaminase is specifically expressed in neonatal rat liver cells undergoing apoptosis upon epidermal growth factor-stimulation

We recently reported that activation of "tissue" transglutaminase (EC 2.3.2.13; tTG) in liver cells undergoing apoptosis determines extensive cross-linking of cellular proteins resulting in the formation of SDS-insoluble shells in the so-called "apoptotic bodies". In attempt to obtain further insight into the role played by tTG in apoptosis of liver cells, we investigated its expression in primary cultures of neonatal rat liver cells stimulated with epidermal growth factor (EGF). EGF-treatment of neonatal rat liver cells induces first hyperplasia of hepatocytes, followed by involution characterized by a high incidence of apoptosis. The proliferative phase of hepatocytes is paralleled by a 10-fold increase in tTG mRNA level, which is followed, during the phase of involution, by sequential increases in enzyme activity and levels of SDS-insoluble apoptotic bodies. tTG immunostaining at both the light- and electron-microscopic levels shows that the most intensive reaction is present in globular structures showing the typical morphological appearance of mature apoptotic bodies. In early apoptotic stages, tTG protein is localized in the perinuclear region of the cell. Intense immunostaining is also found in the apoptotic bodies present inside phagosomes within the cytoplasm of neighboring cells. This evidence confirms and extends our previous findings, indicating that tTG induction and activation specifically takes place in cells undergoing apoptosis, suggesting a key role for the enzyme in the apoptotic program.