Tribbles Homolog 3 Expression Research Articles

TRIB3 mediates the expression of Wnt5a and activation of nuclear factor-κB in Porphyromonas endodontalis lipopolysaccharide-treated osteoblasts.

Porphyromonas endodontalis lipopolysaccharide (LPS) is considered to be correlated with the progression of bone resorption in periodontal and periapical diseases. Wnt5a has recently been implicated in inflammatory processes, but its role is unclear as a P. endodontalis LPS-induced mediator in osteoblasts. Tribbles homolog 3 (TRIB3) encodes a pseudokinase and has been linked to inflammation in certain situations. Here, we found that P. endodontalis LPS induced Wnt5a expression in a dose- and time-dependent manner and it also upregulated translocation, phosphorylation and transcriptional activity of nuclear factor-κB (NF-κB) in MC3T3-E1 cells. Bay 11-7082 blocked the translocation of NF-κB and Wnt5a expression induced by P. endodontalis LPS. Chromatin immunoprecipitation assay further established that induction of Wnt5a by P. endodontalis LPS was mediated through the NF-κB p65 subunit. Additionally, P. endodontalis LPS increased expression of TRIB3 in osteoblasts after 10 h simulated time. Overexpression of TRIB3 enhanced NF-κB phosphorylation and Wnt5a induction, whereas knockdown of TRIB3 inhibited NF-κB phosphorylation and Wnt5a expression in P. endodontalis LPS-stimulated osteoblasts. These results suggest that P. endodontalis LPS has the ability to promote the expression of Wnt5a in mouse osteoblasts, and this induction is mainly mediated by NF-κB pathway. TRIB3 seems to modulate the sustained expression of Wnt5a in osteoblasts stimulated by P. endodontalis LPS, as well as regulating NF-κB phosphorylation.

Hepatic Phosphoserine Aminotransferase 1 Regulates Insulin Sensitivity in Mice via Tribbles Homolog 3.

Phosphoserine aminotransferase 1 (PSAT1) is an enzyme participating in serine synthesis. A role of PSAT1 in the regulation of insulin sensitivity, however, is unknown. In this study, we showed that hepatic PSAT1 expression and liver serine levels are reduced in genetically engineered leptin receptor-deficient (db/db) mice and high-fat diet (HFD)-induced diabetic mice. Additionally, overexpression of PSAT1 by adenovirus expressing PSAT1 improved insulin signaling and insulin sensitivity in vitro and in vivo under normal conditions. Opposite effects were observed when PSAT1 was knocked down by adenovirus expressing small hairpin RNA specific for PSAT1 (Ad-shPSAT1). Importantly, overexpression of PSAT1 also significantly ameliorated insulin resistance in diabetic mice. In addition, PSAT1 inhibited the expression of hepatic tribbles homolog 3 (TRB3) in vitro and in vivo, and adenoviruses expressing small hairpin RNA against TRB3-mediated inhibition of TRB3 reversed the attenuated insulin sensitivity in Ad-shPSAT1 mice. Interestingly, we found that serine mediates PSAT1 regulation of TRB3 expression and insulin signaling in vitro. These results identify a novel function for hepatic PSAT1 in regulating insulin sensitivity and provide important insights in targeting PSAT1 for treating insulin resistance and type 2 diabetes. Our results also suggest that nonessential amino acid serine may play an important role in regulating insulin sensitivity.

Palmitate Induces TRB3 Expression and Promotes Apoptosis in Human Liver Cells

Background/Aims: Parenteral nutrition-associated liver disease (PNALD) is a major complication for patients who require long-term parenteral nutrition. Treatment options for PNALD are limited and its pathogenesis is poorly understood. Tribbles homolog 3 (TRB3) is a pseudokinase that modulates many signal transduction cascades and may be involved in the pathogenesis of PNALD. The aim of this study was to examine the role of TRB3 in palmitate-induced endoplasmic reticulum (ER) stress, in the human liver cell line L02. Methods: L02 cells were treated with palmitate, and its effect on cell viability, mitochondrial membrane potential, apoptosis and TRB3 expression were assessed. The role of TRB3 was also studied using transient overexpression of TRB3 in L02 cells, as well as its interaction with Akt signaling. Results: We found that palmitate induced ER stress and apoptosis in L02 cells. Palmitate-associated ER stress was accompanied by a significant induction of TRB3 expression at the mRNA and protein level. Overexpression of TRB3 potentiated the deleterious effects of palmitate, which was associated with decreased levels of phospho-Akt. Conclusions: TRB3 is an important mediator of palmitate-induced apoptosis in human liver cells, suggesting that it may also be involved in the molecular mechanism underlying PNALD.

Tribbles 3 regulates the fibrosis cytokine TGF- β 1 through ERK1/2-MAPK signaling pathway in diabetic nephropathy.

To reveal the expression and possible role of tribbles homolog 3 (TRB3) in the incidence of type 2 diabetic nephropathy, we used immunohistochemistry, real-time quantitative PCR, western blot analysis, and enzyme-linked immunosorbent assay (ELISA) to study the expression of TRB3, extracellular signal-regulated kinase 1/2 mitogen-activated protein kinase (ERK1/2 MAPK), transforming growth factor β1 (TGF-β1), and collagen type IV in kidneys of db/db diabetic mice and in murine renal mesangial cells stimulated with high glucose. The expression of TRB3, TGF-β1, and collagen type IV was increased in kidneys of db/db diabetic mice. TGF-β1 and collagen type IV regulated by high glucose through ERK1/2 MAPK were downregulated by silencing TRB3 in renal mesangial cells. TRB3 may be involved in diabetic nephropathy by regulating the fibrosis cytokine TGF-β1 and collagen type IV through the ERK1/2 MAPK signaling pathway.

TRIB3 Mediates Glucose-Induced Insulin Resistance via a Mechanism That Requires the Hexosamine Biosynthetic Pathway

In the current study, we investigated the role of tribbles homolog 3 (TRIB3) in glucose-induced insulin resistance and whether the induction of TRIB3 by glucose is dependent on the nutrient-sensing hexosamine biosynthetic pathway (HBP) known to mediate glucose toxicity in diabetes. In diabetic rats, TRIB3 expression in skeletal muscle was increased after 10 days of hyperglycemia, and glycemia and muscle TRIB3 were both restored toward normal by insulin therapy. In L6 myocytes, the induction of TRIB3 by high glucose or glucosamine was reversible upon removal of these substrates. To assess the role of HBP in the induction of TRIB3, we demonstrated that the ability of high glucose to augment TRIB3 expression was prevented by azaserine, an inhibitor of glutamine: fructose-6-phosphate amidotransferase (GFAT), which is the rate-limiting enzyme in the HBP pathway. TRIB3 expression was also substantially stimulated by glucosamine, which bypasses GFAT, accompanied by a decrease in the insulin-stimulated glucose transport rate, and neither response was affected by azaserine. Further, knockdown of TRIB3 inhibited, and TRIB3 overexpression enhanced, the ability of both high glucose and glucosamine to induce insulin resistance. These data provide the mechanistic link between the HBP flux and insulin resistance and point to TRIB3 as a novel target for treatment of glucose-induced insulin resistance.

Knockdown of TRB3 induces apoptosis in human lung adenocarcinoma cells through regulation of Notch 1 expression

The upregulation of tribbles homolog 3 (TRB3), a pseudokinase in mammals, has been observed in several types of malignant cancer, including thyroid, ovarian, liver and colorectal cancer. However, the pathological role and the regulatory mechanism of TRB3 in cancer remain unknown. In the current study, we demonstrated that the expression of TRB3 was upregulated in non-small cell lung cancer (NSCLC), correlating with tumor metastasis, disease recurrence and poor survival in patients. Knocking down TRB3 in aggressive lung cancer cell lines was demonstrated to significantly inhibit their malignant behaviors, including invitro invasion and cell proliferation, as well as invivo metastasis and tumor growth. The correlation between TRB3 and Notch 1 expression revealed that Notch 1 was downregulated by the knockdown of TRB3 in the lung adenocarcinoma cell lines. These results have provided insights into the correlation between TRB3 expression and lung cancer progression, and thus may have potential for the prognosis and therapy of lung cancer.

Regulation of TRIB3 mRNA and Protein in Breast Cancer

Tribbles homolog 3 (TRIB3) is a scaffold protein activated under hypoxic conditions and involved in several cell survival and proliferation pathways. Recently, we reported opposite associations of TRIB3 mRNA and protein with breast cancer prognosis. In this study, we investigated this discrepancy between TRIB3 mRNA and protein in human breast cancer. We provide several lines of evidence demonstrating that TRIB3 is a stabile protein which levels are not controlled by rapid protein breakdown. Interestingly, we were able to show that during anoxia TRIB3 mRNA translation was profoundly inhibited. Hypoxia induced micro RNA 24 was not responsible for the translational repression of TRIB3. Furthermore miRNA-24 expression levels in breast cancer patient specimens showed no correlation with TRIB3 mRNA or TRIB3 protein levels, or with prognosis. Thus, the expression of miRNA-24 does not explain the difference between mRNA and protein expression of TRIB3 in this cohort of breast cancer patients. In conclusion, TRIB3 protein is a stable protein which levels are predominantly regulated by translational control of TRIB3 mRNA transcript.

Tribbles homolog 3 denotes a poor prognosis in breast cancer and is involved in hypoxia response

IntroductionHypoxia in solid tumors is associated with treatment resistance, resulting in poor prognosis. Tribbles homolog 3 (TRIB3) is induced during hypoxia and is involved in multiple cellular pathways involved in cell survival. Here, we investigated the role of TRIB3 in breast cancer.MethodsTRIB3 mRNA expression was measured in breast tumor tissue from 247 patients and correlated with clinicopathological parameters and clinical outcome. Furthermore, we studied TRIB3 expression regulation in cell lines, xenografts tissues and human breast cancer material using Reverse transcriptase, quantitative polymerase chain reaction (RT-qPCR) and immunohistochemical staining. Finally, the effect of small interfering RNA (siRNA) mediated TRIB3 knockdown on hypoxia tolerance was assessed.ResultsBreast cancer patients with low, intermediate or high TRIB3 expression exhibited a mean disease free survival (DFS) of 80 (95% confidence interval [CI] = 74 to 86), 74 (CI = 67 to 81), and 63 (CI = 55 to 71) months respectively (P = .002, Mantel-Cox log-rank). The prognostic value of TRIB3 was limited to those patients that had received radiotherapy as part of their primary treatment (n = 179, P = .005) and remained statistically significant after correction for other clinicopathological parameters (DFS, Hazard Ratio = 1.90, CI = 1.17 to 3.08, P = .009). In breast cell lines TRIB3 expression was induced by hypoxia, nutrient starvation, and endoplasmic reticulum stress in an hypoxia inducible factor 1 (HIF-1) independent manner. TRIB3 induction after hypoxia did not increase with decreasing oxygen levels. In breast tumor xenografts and human breast cancer tissues TRIB3 co-localized with the hypoxic cell marker pimonidazole. The induction of TRIB3 by hypoxia was shown to be regulated via the PERK/ATF4/CHOP pathway of the unfolded protein response and knockdown of TRIB3 resulted in a dose-dependent increase in hypoxia sensitivity.ConclusionsTRIB3 is independently associated with poor prognosis of breast cancer patients, possibly through its association with tumor cell hypoxia.

TRB3: an oxidant stress-induced pseudokinase with a potential to negatively modulate MCP-1 cytokine in diabetic nephropathy

TRB3: an oxidant stress-induced pseudokinase with a potential to negatively modulate MCP-1 cytokine in diabetic nephropathy

Mixed Lineage Kinase-3 Stabilizes and Functionally Cooperates with TRIBBLES-3 to Compromise Mitochondrial Integrity in Cytokine-induced Death of Pancreatic Beta Cells

Mixed lineage kinases (MLKs) have been implicated in cytokine signaling as well as in cell death pathways. Our studies show that MLK3 is activated in leukocyte-infiltrated islets of non-obese diabetic mice and that MLK3 activation compromises mitochondrial integrity and induces apoptosis of beta cells. Using an ex vivo model of islet-splenocyte co-culture, we show that MLK3 mediates its effects via the pseudokinase TRB3, a mammalian homolog of Drosophila Tribbles. TRB3 expression strongly coincided with conformational change and mitochondrial translocation of BAX. Mechanistically, MLK3 directly interacted with and stabilized TRB3, resulting in inhibition of Akt, a strong suppressor of BAX translocation and mitochondrial membrane permeabilization. Accordingly, attenuation of MLK3 or TRB3 expression each prevented cytokine-induced BAX conformational change and attenuated the progression to apoptosis. We conclude that MLKs compromise mitochondrial integrity and suppress cellular survival mechanisms via TRB3-dependent inhibition of Akt.

Human TRB3 is upregulated in stressed cells by the induction of translationally efficient mRNA containing a truncated 5′-UTR

Tribbles homolog 3 (TRB3) is a pseudokinase that has been implicated in the control of stress response, cell viability and metabolic processes, and has been linked to medical conditions, including insulin resistance, cardiovascular disease and diabetes. Therefore, the understanding of mechanisms that regulate TRB3 expression is of considerable importance. We have previously described the existence of several human (h) TRB3 mRNA isoforms that differ in their 5′-untranslated region (5′-UTR). In this study, we use a reverse transcription-quantitative polymerase chain reaction (RT-qPCR) system to characterize the expression levels of hTRB3 mRNA isoforms in HepG2 hepatoma cells cultured in regular medium or exposed to arsenite, and investigate the effect of hTRB3 5′-UTR variants on the efficiency of mRNA translation. The data indicate that of the hTRB3 mRNA splice variants, 1A is predominant (> 80% of molecules) in both the stressed and unstressed states, and that the remainder consists mainly of 1B4, with the variants 1B1, 1B2 and 1B3 together forming less than 1% of the population in either condition. In addition to the substantial transcriptional upregulation of all hTRB3 mRNA splice variants, the exposure of cells to arsenite results in a marked increase in the proportion of splice variant 1A molecules containing a truncated 5′-UTR. The shortened 1A 5′-UTR proved to be translationally more efficient than the untruncated 1A 5′-UTR, due to the lack of an inhibitory upstream open reading frame (uORF). Thus, increased transcription as well as altered usage of 5′-UTR variants contributes to the upregulation of hTRB3 protein synthesis in stressful conditions.

TRB3 suppresses adipocyte differentiation by negatively regulating PPARγ transcriptional activity

In the course of an effort to identify the regulators for peroxisome proliferator-activated receptor gamma (PPARgamma)-dependent perilipin gene expression, we found that tribbles homolog 3 (TRB3), containing a single kinase domain without enzymatic activity, downregulates PPARgamma transcriptional activities by protein-protein interaction. We examined the role that TRB3 plays in adipocyte differentiation in 3T3-L1 cells. TRB3 gene and protein expression was increased during adipocyte differentiation concomitantly with an increase in the mRNA levels of CCAAT/enhancer binding protein homologous protein. The physical interaction between TRB3 and PPARgamma was also verified in 3T3-L1 adipocytes. Forced TRB3 expression in 3T3-L1 cells decreased the mRNA levels of PPARgamma-target genes and intracellular triglyceride levels, whereas knockdown of TRB3 expression by RNA interference increased them. TRB3 also inhibits PPARgamma-dependent adipocyte differentiation in lentivirus-mediated PPARgamma-expressing 3T3-L1 cells. These results provide evidence that TRB3 acts as a potent negative regulator of PPARgamma, a master regulator of adipocyte differentiation, and tightly controls adipogenesis.