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The mammalian target of rapamycin (mTOR) pathway is abnormally activated in lung cancer. However, the anti-lung cancer effect of mTOR inhibitors as monotherapy is modest. Here, we identified that ginsenoside Rh2, an active component of Panax ginseng C. A. Mey., enhanced the anti-cancer effect of the mTOR inhibitor everolimus both in vitro and in vivo. Moreover, ginsenoside Rh2 alleviated the hepatic fat accumulation caused by everolimus in xenograft nude mice models. The combination of everolimus and ginsenoside Rh2 (labeled Eve-Rh2) induced caspase-independent cell death and cytoplasmic vacuolation in lung cancer cells, indicating that Eve-Rh2 prevented tumor progression by triggering paraptosis. Eve-Rh2 up-regulated the expression of c-MYC in cancer cells as well as tumor tissues. The increased c-MYC mediated the accumulation of tribbles homolog 3 (TRIB3)/P62+ aggresomes and consequently triggered paraptosis, bypassing the classical c-MYC/MAX pathway. Our study offers a potential effective and safe strategy for the treatment of lung cancer. Moreover, we have identified a new mechanism of TRIB3/P62+ aggresomes-triggered paraptosis and revealed a unique function of c-MYC.

Colorectal cancer (CRC) is the third most common cancer and the second leading cause of death due to cancer in the world. Therefore, the identification of novel druggable targets is urgently needed. Tribbles proteins belong to a pseudokinase family, previously recognized in CRC as oncogenes and potential therapeutic targets. Here, we analyzed the expression of TRIB1, TRIB2, and TRIB3 simultaneously in 33 data sets from CRC based on available GEO profiles. We show that all three Tribbles genes are overrepresented in CRC cell lines and primary tumors, though depending on specific features of the CRC samples. Higher expression of TRIB2 in the tumor microenvironment and TRIB3 overexpression in an early stage of CRC development, unveil a potential and unexplored role for these proteins in the context of CRC. Differential Tribbles expression was also explored in diverse cellular experimental conditions where either genetic or pharmacological approaches were used, providing novel hints for future research. This comprehensive bioinformatic analysis provides new insights into Tribbles gene expression and transcript regulation in CRC. info:eu-repo/semantics/publishedVersion

Pulmonary fibrosis (PF) is a chronic, progressive, fatal interstitial lung disease with limited available therapeutic strategies. We recently reported that the protein kinase glycogen synthase kinase-3β (GSK-3β) interacts with and inactivates the ubiquitin-editing enzyme A20 to suppress the degradation of the transcription factor CCAAT/enhancer-binding protein beta (C/EBPβ) in alveolar macrophages (AMs), resulting in a profibrotic phenotype of AMs and promoting the development of PF. Here, we showed that chronic lung injury upregulated the stress response protein tribbles homolog 3 (TRIB3), which interacted with GSK-3β and stabilized GSK-3β from ubiquitination and degradation. Elevated GSK-3β expression phosphorylated A20 to inhibit its ubiquitin-editing activity, causing the accumulation of C/EBPβ and the production of several profibrotic factors in AMs and promoting PF development. Activated C/EBPβ, in turn, increased the transcription of TRIB3 and GSK-3β, thereby establishing a positive feedback loop in AMs. The knockdown of TRIB3 expression or the pharmacologic disruption of the TRIB3‒GSK-3β interaction was an effective PF treatment. Our study reveals an intact profibrotic axis of TRIB3‒GSK-3β‒A20‒C/EBPβ in AMs, which represents a target that may provide a promising treatment strategy for PF.

Background Granulosa cells (GCs) in cumulus oophorus highly express follicle stimulating hormone receptor (FSHR), which is the most important mediator of both estradiol synthesis and oocyte maturation. Obese women have elevated free fatty acids (FFAs) levels in their follicular fluids and decreased FSHR expression in GCs, which is related to an altered protein kinase B/glycogen synthase kinase 3β (Akt/GSK3β) signaling pathway. Such FFA increases accompany 3-fold rises in pseudokinase 3 (TRIB3) expression and reduce the Akt phosphorylation status in both the human liver and in insulinoma cell lines. Therefore, in a high FFA environment, we determined if TRIB3 mediates regulation of FSHR via the Akt/GSK3β signaling pathway in human GCs. Methods GCs from women undergoing in vitro fertilization were collected and designated as high and low FFAs cohorts based on their follicular fluid FFA content. GCs with low FFA levels and a human granulosa-like tumor (KGN) cell line were exposed to palmitic acid (PA), which is a dominate FFA follicular fluid constituent. The effects were assessed of this substitution on the Akt/GSK3β signaling pathway activity as well as the expressions of TRIB3 and FSHR at both the gene and protein levels by qPCR, Western blot and immunofluorescence staining analyses. Meanwhile, the individual effects of TRIB3 knockdown in KGN cells and p-AKT inhibitors were compared to determine the mechanisms of FFA-induced FSHR downregulation. Results The average FSH dose consuming per oocyte (FSH dose/oocyte) was elevated and Top embryo quality ratio was decreased in women with high levels of FFAs in their follicular fluid. In these women, the GC TRIB3 and ATF4 protein expression levels were upregulated which was accompanied by FSHR downregulation. Such upregulation was confirmed based on corresponding increases in their gene expression levels. On the other hand, the levels of p-Akt decreased while p-GSK3β increased in the GCs. Moreover, TRIB3 knockdown reversed declines in FSHR expression and estradiol (E2) production in KGN cells treated with PA, which also resulted in increased p-Akt levels and declines in the p-GSK3β level. In contrast, treatment of TRIB3-knockdown cells with an inhibitor of p-Akt (Ser473) resulted in rises in the levels of both p-GSK3β as well as FSHR expression whereas E2 synthesis fell. Conclusions During exposure to a high FFA content, TRIB3 can reduce FSHR expression through stimulation of the Akt/GSK3β pathway in human GCs. This response may contribute to inducing oocyte maturation.

Tumor‐associated macrophages (TAMs), one of the most common cell components in the tumor microenvironment, have been reported as key contributors to cancer‐related inflammation and enhanced metastatic progression of tumors. To explore the underlying mechanism of TAM‐induced tumor progression, TAMs were isolated from colorectal cancer patients, and the functional interaction with colorectal cancer cells was analyzed. Our study found that coculture of TAMs contributed to a glycolytic state in colorectal cancer, which promoted the stem‐like phenotypes and invasion of tumor cells. TAMs produced the cytokine transforming growth factor‐β to support hypoxia‐inducible factor 1α (HIF1α) expression, thereby upregulating Tribbles pseudokinase 3 (TRIB3) in tumor cells. Elevated expression of TRIB3 resulted in activation of the β‐catenin/Wnt signaling pathway, which eventually enhanced the stem‐like phenotypes and cell invasion in colorectal cancer. Our findings provided evidence that TAMs promoted colorectal cancer progression in a HIF1α/TRIB3‐dependent manner, and blockade of HIF1α signals efficiently improved the outcome of chemotherapy, describing an innovative approach for colorectal cancer treatment., Our findings provided evidence that tumor‐associated macrophages promoted colorectal progression in a HIF1α/TRIB3‐dependent manner, and blockade of HIF1α signals efficiently improved the outcome of chemotherapy, describing an innovative approach for colorectal cancer treatment.

Long non-coding RNA (lncRNA) X-inactive specific transcript (Xist) is involved in apoptosis and inflammatory injury. This study aimed to assess the role of lncRNA Xist in sevoflurane-induced social and emotional impairment and neuronal apoptosis in neonatal mice and hippocampal neuronal cells. The performance in social and emotional tests and the expression levels of lncRNA Xist and microRNA (miR)-98-5p after sevoflurane exposure were measured. Moreover, the effects of suppression of lncRNA Xist on neuronal apoptosis and endoplasmic reticulum (ER) stress were determined. Subsequently, the association among lncRNA Xist, miR-98-5p, and ER degradation-enhancing α-mannosidase-like 1 protein (EDEM1) was explored. Our results showed that lncRNA Xist increased, miR-98-5p decreased, and social and emotional impairment appeared after sevoflurane exposure. Furthermore, suppression of lncRNA Xist improved sevoflurane-induced social and emotional impairment and reduced sevoflurane-induced neuronal apoptosis and ER stress in vivo and in vitro. Moreover, lncRNA Xist negatively regulated miR-98-5p expression, and it contributed to sevoflurane-induced neuronal apoptosis and ER stress by sponging miR-98-5p. Additionally, EDEM1 was identified as a target of miR-98-5p. Our findings revealed that the knockdown of lncRNA Xist ameliorates sevoflurane-induced social and emotional impairment through inhibiting neuronal apoptosis and ER stress by targeting the miR-98-5p/EDEM1 axis., Graphical abstract, Chen and colleagues proved that lncRNA XIST knockdown can ameliorate sevoflurane-induced social and emotional impairment through inhibiting neuron apoptosis and endoplasmic reticulum stress by targeting the miR-98-5p/EDEM1 axis. The understanding of cell death mechanisms in anesthesia-caused social and emotional impairment is vital to improve interventions in patients and reduce neural dysfunctionality.

The current understanding of the molecular pathogenesis of diabetic retinopathy does not provide a mechanistic link between early molecular changes and the subsequent progression of the disease. In this study, we found that human diabetic retinas overexpressed TRIB3 and investigated the role of TRIB3 in diabetic retinal pathobiology in mice. We discovered that TRIB3 controlled major molecular events in early diabetic retinas via HIF1α-mediated regulation of retinal glucose flux, reprogramming cellular metabolism, and governing of inflammatory gene expression. These early molecular events further defined the development of neurovascular deficit observed in mice with diabetic retinopathy. TRIB3 ablation in the streptozotocin-induced mouse model led to significant retinal ganglion cell survival and functional restoration accompanied by a dramatic reduction in pericyte loss and acellular capillary formation. Under hypoxic conditions, TRIB3 contributed to advanced proliferative stages by significant upregulation of GFAP and VEGF expression, thus controlling gliosis and aberrant vascularization in oxygen-induced retinopathy mouse retinas. Overall, our data reveal that TRIB3 is a master regulator of diabetic retinal pathophysiology that may accelerate the onset and progression of diabetic retinopathy to proliferative stages in humans and present TRIB3 as a potentially novel therapeutic target for diabetic retinopathy.

C/EBP homologous protein (CHOP) has been identified as a suitable therapeutic target to combat atherosclerosis but the mechanism has not been fully studied. Here, we sought to define the role and underlying mechanism of CHOP in atherosclerosis. Mouse models of atherosclerosis in ApoE-/- mice were established by high-fat feeding, where miR-208 expression was determined. Then atherosclerotic plaque tissues were isolated from the model mice. Loss- and gain-function assays were performed on trypsinized vascular smooth muscle cells (VSMCs) to test the in vitro effect of CHOP in controlling the tribbles homologue 3 (TRIB3)/microRNA-208 (miR-208)/tissue inhibitor of metalloproteinases-3 (TIMP3) axis in atherosclerosis by determining cell proliferation and migration as well as blood lipid levels. Moreover, expression of α-smooth muscle actin (α-SMA) and type I collagen expression was determined using immunofluorescence staining to assess plaque stability in mice. miR-208 expression was elevated in atherosclerosis samples and miR-208 overexpression promoted proliferation and migration of VSMCs but diminished plaque stability in mice. TIMP3 was targeted by miR-208, which could be abrogated by upregulation of TIMP3. In addition, CHOP increased TRIB3 expression to upregulate miR-208 and to downregulate TIMP3, which potentiated VSMC proliferation and migration in vitro and in vivo. Taken together, inhibition of CHOP may inhibit the proliferation and migration of VSMCs as well as reduce the levels of TC, TG, and LDL-C but increase the level of HDL-C through the TRIB3/miR-208/TIMP3 axis, thereby inhibiting the progression of atherosclerosis.

UTX/KDM6A encodes a major histone H3 lysine 27 (H3K27) demethylase, and is frequently mutated in various types of human cancers. Although UTX appears to play a crucial role in oncogenesis, the mechanisms involved are still largely unknown. Here we show that a specific pharmacological inhibitor of H3K27 demethylases, GSK-J4, induces the expression of transcription activating factor 4 (ATF4) protein as well as the ATF4 target genes (e.g. PCK2, CHOP, REDD1, CHAC1 and TRIB3). ATF4 induction by GSK-J4 was due to neither transcriptional nor post-translational regulation. In support of this view, the ATF4 induction was almost exclusively dependent on the heme-regulated eIF2α kinase (HRI) in mouse embryonic fibroblasts (MEFs). Gene expression profiles with UTX disruption by CRISPR-Cas9 editing and the following stable re-expression of UTX showed that UTX specifically suppresses the expression of the ATF4 target genes, suggesting that UTX inhibition is at least partially responsible for the ATF4 induction. Apoptosis induction by GSK-J4 was partially and cell-type specifically correlated with the activation of ATF4-CHOP. These findings highlight that the anti-cancer drug candidate GSK-J4 strongly induces ATF4 and its target genes via HRI activation and raise a possibility that UTX might modulate cancer formation by regulating the HRI-ATF4 axis.

Made available in DSpace on 2021-06-25T11:10:56Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-01-01 COVID-19 is prevalent in the elderly. Old individuals are more likely to develop pneumonia and respiratory failure due to alveolar damage, suggesting that lung senescence may increase the susceptibility to SARS-CoV-2 infection and replication. Considering that human coronavirus (HCoVs; SARS-CoV-2 and SARS-CoV) require host cellular factors for infection and replication, we analyzed Genotype-Tissue Expression (GTEx) data to test whether lung aging is associated with transcriptional changes in human protein-coding genes that potentially interact with these viruses. We found decreased expression of the gene tribbles homolog 3 (TRIB3) during aging in male individuals, and its protein was predicted to interact with HCoVs nucleocapsid protein and RNA-dependent RNA polymerase. Using publicly available lung single-cell data, we found TRIB3 expressed mainly in alveolar epithelial cells that express SARS-CoV-2 receptor ACE2. Functional enrichment analysis of age-related genes, in common with SARS-CoV-induced perturbations, revealed genes associated with the mitotic cell cycle and surfactant metabolism. Given that TRIB3 was previously reported to decrease virus infection and replication, the decreased expression of TRIB3 in aged lungs may help explain why older male patients are related to more severe cases of the COVID-19. Thus, drugs that stimulate TRIB3 expression should be evaluated as a potential therapy for the disease. Department of Structural and Functional Biology Institute of Biosciences São Paulo State University (UNESP) Faculty of Medicine São Paulo State University UNESP Department of Chemical and Biological Sciences Institute of Biosciences São Paulo State University (UNESP) Department of Biochemistry and Tissue Biology Institute of Biology State University of Campinas (UNICAMP) Department of Structural and Functional Biology Institute of Biosciences São Paulo State University (UNESP) Faculty of Medicine São Paulo State University UNESP Department of Chemical and Biological Sciences Institute of Biosciences São Paulo State University (UNESP)

Yutong Xing,1,2,* Peng Luo,2,* Rui Hu,1 Duanduan Wang,1 Gang Zhou,2 Jie Jiang3 1Department of Cardiothoracic Surgery, The Fifth Hospital of Xiamen, Xiamen, People’s Republic of China; 2Department of Cardiothoracic Surgery, The First Affiliated Hospital of Jiamusi University, Jiamusi, People’s Republic of China; 3Department of Thoracic Surgery, The First Affiliated Hospital of Xiamen University, Xiamen, People’s Republic of China*These authors contributed equally to this workCorrespondence: Yutong Xing Department of Cardiothoracic SurgeryThe Fifth Hospital of Xiamen, Xiamen, Fujian 361101, People’s Republic of ChinaEmail xingyt2018@163.comBackground: The pseudokinase Tribbles 3 (TRIB3) is involved in many cellular processes and various cancers. In recent years, the importance of metabolic transformation in the maintenance of malignant tumors has become increasingly prominent. Abnormal metabolism of cancer cells is considered a hallmark of cancer. However, the exact role and molecular mechanism of TRIB3 in lung adenocarcinoma (LUAD) cell reprogramming is largely unknown.Methods: The oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) of cells were examined with a Seahorse XF Extracellular Flux Analyzer. In vitro and in vivo RT-qPCR, Western blotting, and functional assays were performed to explore the functional roles of TRIB3 in LUAD.Results: In the present study, we demonstrated that TRIB3 is remarkably upregulated in LUAD cell lines as well as tissues. TRIB3 knockdown significantly inhibited LUAD cell growth and suppressed LUAD cell invasion, while TRIB3 overexpression conferred the opposite effects. Moreover, silencing TRIB3 suppressed the tumorigenesis and metastatic ability of LUAD cells. Mechanistically, we demonstrated that silencing TRIB3 significantly impaired aerobic glycolysis ability in LUAD cells. Furthermore, our data indicated that TRIB3 knockdown decreased hypoxia-inducible factor (HIF)1α levels and targeted the glycolytic genes regulated by HIF1α.Conclusion: Together, our findings revealed a previously unappreciated function of TRIB3 in cancer cell metabolism and tumor progression, illustrating that TRIB3 could be considered a valuable therapeutic target for LUAD patients.Keywords: TRIB3, LUAD, aerobic glycolysis, HIF1&alpha

The transcription factor MYC is deregulated in almost all human cancers, especially in aggressive lymphomas, through chromosomal translocation, amplification, and transcription hyperactivation. Here, we report that high expression of tribbles homologue 3 (TRIB3) positively correlates with elevated MYC expression in lymphoma specimens; TRIB3 deletion attenuates the initiation and progression of MYC-driven lymphoma by reducing MYC expression. Mechanistically, TRIB3 interacts with MYC to suppress E3 ubiquitin ligase UBE3B-mediated MYC ubiquitination and degradation, which enhances MYC transcriptional activity, causing high proliferation and self-renewal of lymphoma cells. Use of a peptide to disturb the TRIB3-MYC interaction together with doxorubicin reduces the tumor burden in MycEμ mice and patient-derived xenografts. The pathophysiological relevance of UBE3B, TRIB3 and MYC is further demonstrated in human lymphoma. Our study highlights a key mechanism for controlling MYC expression and a potential therapeutic option for treating lymphomas with high TRIB3-MYC expression., c-MYC is often deregulated in human cancers including lymphomas. Here, the authors show that a member of the pseudokinase family, tribbles homologue 3 (TRIB3), interacts with c-MYC to suppress c-MYC ubiquitination and degradation, leading to increased proliferation and self-renewal of lymphoma cells.

The pseudokinase Tribble 3 (TRIB3) is known as a regulator in cellular responses to a variety of stresses, such as glucose insufficiency and endoplasmic reticulum (ER) stress. TRIB3 is upregulated in various cancer tissues and is closely connected to the poor prognosis of patients. However, the underlying regulation and function of TRIB3 in glioblastoma (GBM) is still largely unknown. In this study, the upregulation of TRIB3 was confirmed both in primary specimens from GBM patients and in vitro with GBM cell lines. Overexpression of specific TRIB3 transcripts promoted cell growth and migration in vitro, while knockdown of TRIB3 expression exerted a repressive effect on these cellular processes. The growth-promoting effect of TRIB3 was also demonstrated in a xenograft mouse model. Mechanistic studies further revealed that TRIB3 was able to suppress autophagic flux and that this suppression was responsible for TRIB3 silencing-induced proliferation and migration of GBM cells. These findings indicate that the suppression of autophagic flux by TRIB3 drives the invasion and proliferation of GBM cells, thus suggesting that TRIB3 is a potential novel therapeutic target for the treatment of glioma.

Background Tribbles pseudokinase 3 (TRIB3) protein is a pseudokinase which plays an important role in cellular stress, metabolism, and tumor progression. However, the expression and function of TRIB3 in ovarian cancer is unknown. Methods TRIB3 expression was detected by immunohistochemistry in the ovarian tissue samples. Following down-regulation of TRIB3 by siRNA, multiple aspects of ovarian cancer cells were detected by the MTT assay, flow cytometry, scratch test and Transwell. Additionally, changes in related molecules and the MEK/ERK pathway were detected by western blotting. Finally, many bioinformatic methods, websites and databases, such as gene set enrichment analysis (GSEA), DVAID, Genemania, TISIDB and cBioPortal were used to study the TRIB3. Results The expression level of TRIB3 was higher in ovarian epithelial malignant tumors as compared to other groups. Patients with a high expression level of TRIB3 had significantly shorter survival times,which was consistent with the results of analysis of the KM-plot database. Down-regulation of TRIB3 expression significantly inhibited the proliferation, invasion, and migration capabilities of ovarian cancer cells, and induced apoptosis and cell cycle arrest. Following TRIB3 siRNA transfection, expression levels of relative proteins were found to be decreased. Additionally, analysis in DAVID website and GSEA revealed that TRIB3 expression was associated with multiple biological processes. Protein phosphorylation levels of MEK and ERK also decreased following TRIB3-siRNA transfection. The Genemania website was used to analyze the proteins that interact with TRIB3. Analysis of TRIB3 in the TISIDB database and cBioPortal website showed that ovarian cancer patients with high levels of mutation in TRIB3 had poor prognosis, and that the expression of TRIB3 was related to immunomodulation. Conclusions The TRIB3 was highly expressed and promoting the malignant behavior of ovarian cancer cells by activating the MEK-ERK signaling pathway. It was also found to be associated with genetic variations and immune modulators.

Objective To explore the biological function and molecular mechanism of Sp2 in hepatocellular carcinoma (HCC). Methods Tissue microarray immunohistochemistry and western blot were used to study the expression of Sp2 in hepatocellular tissue and adjacent non‐neoplastic tissues (ANT). In HCC cell lines, the role of Sp2 was determined by in vitro experiments such as CCK8, clone formation test, Transwell assay, wound‐healing assay, and flow cytometry apoptotic analysis, and its possible mechanism was analyzed. Results Compared with ANT, Sp2 expression in HCC tissues was significantly up‐regulated, which was strongly associated with stage of tumor and poor prognosis of patients. TCGA database were further confirmed these results. Besides, functional studies had shown that Sp2 knockdown not only leads to a decrease in cell proliferation and an increase in cell apoptosis but also inhibits the cells' abilities of migration and invasion. Sp2 silencing could inhibit the expression of TRIB3 protein and down‐regulate the endoplasmic reticulum stress (ERS) level of HCC. Conclusion Sp2 may play a part in promoting cancer by regulating TRIB3 protein, which may be a factor of prognostic and a potential new therapeutic target for HCC., We found that Sp2 expression was significantly up‐regulated in HCC tissues, which was closely related to tumor staging and poor prognosis of patients. Functional studies had shown that Sp2 knockdown not only leads to a decrease in cell proliferation and an increase in cell apoptosis, but also inhibits migration and invasion of hepatoma cells. Sp2 may play a role in promoting cancer by regulating TRIB3 protein, which may be a prognostic factor and a potential new therapeutic target for HCC.

Radioresistance becomes the major obstacle to reduce tumor recurrence and improve prognosis in the treatment of esophageal squamous cell carcinoma (ESCC). Thus new strategies for radioresistant ESCC are urgently needed. Herein, we reported that tribbles pseudokinase 3 (TRIB3) serves as a key regulator of radioresistance in ESCC. TRIB3 is overexpressed in ESCC tissues and cell lines. High expression of TRIB3 significantly correlates with poor radiotherapy response and prognosis in ESCC patients. Upregulation of TRIB3 in ESCC cells conferred radioresistance in vitro and in vivo by interacting with TAZ thus impeding β-TrCP-mediated TAZ ubiquitination and degradation. Conversely, silencing TRIB3 sensitized ESCC cells to ionizing radiation. More importantly, TRIB3 was significantly correlated with TAZ activation in ESCC biopsies, and patients with high expression of both TRIB3 and TAZ suffered the worst radiotherapy response and survival. Our study uncovers the critical mechanism of ESCC resistance to radiotherapy, and provides a new pharmacological opportunity for developing a mechanism-based strategy to eliminate radioresistant ESCC in clinical practice.

Under the differentiation induction therapy with all-trans retinoic acid and arsenic trioxide, nearly 95% of typical acute promyelocyte leukemia (APL), which is characterized by the presence of PML-RARA, patients can be cured. Though its good prognosis, if left untreated, the natural survival duration of typical APL patients is only 1 month, but some exceptional cases also exist. Occasionally, we have observed the entire natural clinical course of one extremely indolent APL patient, who developed from pre-APL stage (

© 2021 by the authors., [Background]: Tribbles pseudokinase 3 (TRIB3) has been proposed to both promote and restrict cancer generation and progression. However, the precise mechanisms that determine this dual role of TRIB3 in cancer remain to be understood. In this study we aimed to investigate the role of TRIB3 in luminal breast cancer, the most frequent subtype of this malignancy. [Methods]: We genetically manipulated TRIB3 expression in a panel of luminal breast cancer cell lines and analyzed its impact on cell proliferation, and the phosphorylation, levels, or subcellular localization of TRIB3 and other protein regulators of key signaling pathways in luminal breast cancer. We also analyzed TRIB3 protein expression in samples from luminal breast cancer patients and performed bioinfor-matic analyses in public datasets. Results: TRIB3 enhanced the proliferation and AKT phosphoryla-tion in luminal A (HER2-) but decreased them in luminal B (HER2+) breast cancer cell lines. TRIB3 negatively regulated the stability of HER2 in luminal B breast cancer cell lines. TRIB3 expression was associated with increased disease-free survival and a better response to therapy in luminal breast cancer patients. [Conclusions]: Our findings support the exploration of TRIB3 as a potential biomarker and therapeutic target in luminal breast cancer., This work was funded by the Instituto de Salud Carlos III (ISCIII) and confounded by the European Regional Development Fund (ERDF), “A way to make Europe”, grant number PI18/00442 integrated into the State Plan for R & D + I 2017-2020; by the European Commission through the Horizon 2020 European Training Networks program, grant number H2020-MSCA-ITN308 2016 721532; and by the Breast Cancer Now (formerly Breast Cancer Campaign) foundation, grant number 2012NovSP033. Alba Orea-Soufi was supported by a predoctoral FPU contract from Spanish Government. Work in SC-L group is supported by a MINECO/FEDER research grant RTI2018-094130-B-100.

Doublecortin-like kinase 1 (DCLK1) promotes tumour proliferation in human colorectal cancer (CRC). To elucidate the mechanism and clinical relevance of this association, we performed expression analysis using commercially available colon carcinoma cell lines (SW480, HCT116, CaCO2, SW48 and SKCO1) and immunohistochemical analysis of 200 resected CRC samples for correlation with clinical features. DCLK1 showed a high level of expression, especially in SW480 and HCT116 cells. Silencing DCLK1 expression using short hairpin DCLK1 (shDCLK1) RNA inhibited the growth and invasion capacities of these cell lines, which showed signs of entering into the mesenchymal–epithelial transition (MET). We found evidence of a strong correlation of DCLK1 expression with that of Tribbles homolog 3 (TRIB3), and silencing TRIB3 also led to the MET phenotype in these cells. In the clinical samples, compared with samples showing low expression of DCLK1, high expression was associated with poor prognosis in terms of overall and recurrence-free survival (P < 0.0001). The results of univariate and multivariate analysis suggested that high expression of DCLK1 in clinical colon cancer samples was tied to poor prognosis, cancer invasion depth and lymph node metastasis. DCLK1 expression correlates with malignant grade of colon cancer and offers a potential treatment target.

Although dyslipidemia commonly occurs in patients with acute promyelocytic leukemia (APL) in response to anti-APL therapy, the underlying mechanism and the lipid statuses of patients with newly diagnosed APL remain to be addressed. Methods: We conducted a retrospective study to investigate the lipid profiles of APL patients. PML-RARα transgenic mice and APL cells-transplanted mice were used to assess the effects of APL cells on the blood/liver lipid levels. Subsequently, gene set enrichment analysis, western blot and dual luciferase reporter assay were performed to examine the role and mechanism of PML-RARα and TRIB3 in lipid metabolism regulation in APL patients at pretreatment and after induction therapy. Results: APL patients exhibited a higher prevalence of dyslipidemia before anti-APL therapy based on a retrospective study. Furthermore, APL cells caused secretion of triglycerides, cholesterol, and PCSK9 from hepatocytes and degradation of low-density lipoprotein receptors in hepatocytes, which elevated the lipid levels in APL cell-transplanted mice and Pml-Rarα transgenic mice. Mechanistically, pseudokinase TRIB3 interacted with PML-RARα to inhibit PPARγ activity by interfering with the interaction of PPARγ and RXR and promoting PPARγ degradation. Thus, reduced PPARγ activity in APL cells decreased leptin but increased resistin expression, causing lipid metabolism disorder in hepatocytes and subsequent dyslipidemia in mice. Although arsenic/ATRA therapy degraded PML-RARα and restored PPARγ expression, it exacerbated dyslipidemia in APL patients. The elevated TRIB3 expression in response to arsenic/ATRA therapy suppressed PPARγ activity by disrupting the PPARγ/RXR dimer, which resulted in dyslipidemia in APL patients undergoing therapy. Indeed, the PPAR activator not only enhanced the anti-APL effects of arsenic/ATRA by suppressing TRIB3 expression but also reduced therapy-induced dyslipidemia in APL patients. Conclusion: Our work reveals the critical role of the PML-RARα/PPARγ/TRIB3 axis in the development of dyslipidemia in APL patients, potentially conferring a rationale for combining ATRA/arsenic with the PPAR activator for APL treatment.

Objective The aim of this study was to examine the effect of TRIB3 on proliferation, apoptosis, and migration of endometrial cancer (EC) cells and explore the relationship between TRIB3 and AKT signaling pathway in EC progression. Methods Immunohistochemical analysis was performed to measure the expression level of TRIB3 in normal endometrium tissues and EC tissues. Overexpression and shRNA knockdown techniques were applied by transfecting EC cells (ISK and AN3CA), and the effect of TRIB3 on EC cell biological behaviors was evaluated. Cell Counting Kit-8 and colony formation assays were utilized to investigate EC cell proliferation ability, and flow cytometry was performed to assess the apoptosis of EC cells. Moreover, the migration and invasion of EC cells were detected by transwell assay, and the levels of MMP-2 and MMP-9 were measured by ELISA. Additionally, Western blot analysis was carried out to determine the levels of AKT and p-AKT. Results The expression level of TRIB3 was higher in EC than normal endometrium tissues, and its overexpression promoted apoptosis and suppressed proliferation of EC cells. Furthermore, TRIB3 retarded the migration and invasion of EC cells and decreased the levels of MMP-2 and MMP-9. Conversely, TRIB3 inhibition enhanced the expression levels of MMP-2 and MMP-9, and proliferation and migration of EC cells but suppressed their apoptosis. Similarly, TRIB3 overexpression reduced while its knockdown increased the level of p-AKT. Conclusion TRIB3 inhibited proliferation and migration and promoted apoptosis of EC cells probably through regulating AKT signaling pathway.

Background Integrin αvβ3 has been proposed as crucial determinant for tumor sustained progression and a molecular marker for the estimation of tumor angiogenesis. Our study suggested that integrin αvβ3 could efficiently promote lung cancer cell proliferation and stem-like phenotypes in a tribbles homolog 3 (TRIB3) dependent manner. Result Integrin αvβ3 could mediate the activation of FAK/AKT pro-survival signaling pathway. Meanwhile, activated TRIB3 interacted with AKT to upregulated FOXO1 and SOX2 expression, resulting in sustained tumor progression in lung cancer. Our further analysis revealed that TRIB3 was significantly upregulated in lung tumor tissues and correlated with the poor outcome in clinical patients, indicating the potential role of TRIB3 in diagnostic and prognostic estimation for patients with lung cancer. Conclusion Our study showed here for the first time that integrin αvβ3 promote lung cancer development by activating the FAK/AKT/SOX2 axis in a TRIB3 dependent signaling pathway, and interrupting TRIB3/AKT interaction significantly improved the outcome of chemotherapy in tumor-bearing mice, representing a promising therapeutic strategy in lung cancer.

Eccentric exercise training accompanied by a low-fat diet can prevent insulin resistance (IR) and is currently an effective method for the treatment of IR induced by high-fat diet (HFD)-associated obesity. However, the molecular mechanisms underlying this improvement of IR in adipose tissue are still not completely clear. In this study, 5–6-week-old male mice were randomly divided into a standard control diet (SCD) group (SC, n = 12) and a HFD group (HF, n = 72). After 12 weeks, 12 mice in each group were randomly sacrificed. The remaining mice in the HF group were randomly submitted to one of the following experimental protocols for 8 weeks: obesity-HFD-sedentary (OHF-Sed, n = 14), obesity-HFD-exercise (OHF-Ex, n = 16), obesity-SCD-sedentary (OSC-Sed, n = 14), and obesity-SCD-exercise (OSC-Ex, n = 16). All obese mice in the exercise group were subjected to downhill running. Half of the mice in each group received an insulin injection (0.75 U/kg) before sample collection. Epididymal fat was removed and weighed. Adipocyte size and inflammatory cell infiltration were observed by H&E staining. Both basal and insulin-stimulated GLUT4 fluorescence and protein contents were detected by immunofluorescence and Western blot. Levels of IL-1β and IL-10 were detected by ELISA. Protein contents of iNOS, Arg-1, TRIB3, p-AKT, and AKT were determined by Western blot. CD86 and CD206 fluorescence were determined by immunofluorescence. The results showed that a HFD for 12 weeks induced IR accompanied by adipose tissue macrophages M1 polarization (increased iNOS protein content and CD86 fluorescence) and TRIB3-AKT activation. Downhill running accompanied by a low-fat diet attenuated IR (p < 0.01), reduced inflammation levels (increased IL-10 protein content and decreased IL-1β protein content), inhibited adipose tissue macrophages M1 polarization (decreased iNOS protein content and CD86 fluorescence) and promoted M2 polarization (increased Arg-1 protein content and CD206 fluorescence), and suppressed TRIB3-AKT signaling. We concluded that downhill running accompanied by dietary fat regulation attenuates HFD-related IR in mice, which may be associated with reduced TRIB3-AKT signaling and activated M2 macrophages in adipose tissue.

Interleukin (IL-6) is a pleotropic cytokine with both tumor-promoting and -inhibitory effects on breast cancer growth. However, the mechanisms governing the outcome of IL-6 on cancer progression remain to be clarified. Our study unraveled a novel long noncoding RNA (lncRNA) AU021063 downstream of IL-6 signaling. We found that IL-6 induced the expression of AU021063 predominantly in breast cancer compared to other cancer types. Mechanistically, IL-6 induced AT-rich interactive domain 5a (Arid5a) expression, which promotes the transcription of AU021063. In turn, AU021063 promotes breast cancer metastasis through stabilizing tribbles homolog 3 (Trib3) and activating Mek/Erk signaling pathway. Genetic ablation of either Arid5a, AU021063 or Trib3 abolished breast cancer metastasis in vitro and in vivo. Overall, our study highlights the importance of IL-6-Arid5a-AU021063 axis in regulating breast cancer invasiveness and metastasis, which may provide potential novel therapeutics for breast cancer.

© 2021 by the authors., The Tribbles pseudokinases family consists of TRIB1, TRIB2, TRIB3 and STK40 and, although evolutionarily conserved, they have distinctive characteristics. Tribbles members are expressed in a context and cell compartment-dependent manner. For example, TRIB1 and TRIB2 have potent oncogenic activities in vertebrate cells. Since the identification of Tribbles proteins as modulators of multiple signalling pathways, recent studies have linked their expression with several pathologies, including cancer. Tribbles proteins act as protein adaptors involved in the ubiquitin-proteasome degradation system, as they bridge the gap between substrates and E3 ligases. Between TRIB family members, TRIB2 is the most ancestral member of the family. TRIB2 is involved in protein homeostasis regulation of C/EBPα, β-catenin and TCF4. On the other hand, TRIB2 interacts with MAPKK, AKT and NFkB proteins, involved in cell survival, proliferation and immune response. Here, we review the characteristic features of TRIB2 structure and signalling and its role in many cancer subtypes with an emphasis on TRIB2 function in therapy resistance in melanoma, leukemia and glioblastoma. The strong evidence between TRIB2 expression and chemoresistance provides an attractive opportunity for targeting TRIB2., This work was supported by the Spanish Ministry of Science, Innovation and Universities through Grant RTI2018-094629-B-I00 to W.L.

BACKGROUND: Tribbles pseudokinase 3 (TRIB3) is a member of the tribbles-related family, which has been determined in various cancers, including renal cell carcinoma, acute promyelocytic leukemia, colorectal cancer, endometrial cancer, and glioma. However, its role in retinoblastoma (RB) has not yet been explored. METHODS: The expression level of TRIB3 was detected in RB tissues and cell lines using qRT-PCR. The effects of TRIB3 on cell proliferation and invasion capacities were analyzed with MTT, crystal violet, and transwell assays. Western blot and rescue assays were conducted to explore the underlying mechanism. RESULTS: This study found that TRIB3 was upregulated in human RB tissues compared to adjacent normal tissues both at the mRNA and protein levels. Overexpression of TRIB3 significantly promoted cell proliferation and invasion of RB cells, while TRIB3 knockdown inhibited these processes. Moreover, the mechanism deciphering experiments showed that TRIB3 overexpression can increase AKT and mTOR phosphorylation. Conversely, TRIB3 knockdown decreased the phosphorylation of AKT and mTOR. Additionally, MK2206, a potent AKT inhibitor, blocked the promotive effects of TRIB3 in RB cells. CONCLUSION: This study demonstrated that TRIB3 acts as an oncogene and plays a crucial role in the proliferation and invasion of RB cells via regulating the AKT/mTOR signaling pathway. Therefore, TRIB3 may serve as a potential target in the diagnosis and/or treatment of RB.

BackgroundBladder cancer is a common malignant tumor characterized by high mortality and high management costs; however, it lacks useful molecular prognostic markers. Tribbles pseudokinase 3 (TRIB3) is a pseudokinase that participates in cell tumor progression and metabolism and whose function in bladder cancer is not precisely known.Main MethodsWe downloaded transcriptome data and clinical data of bladder cancer from associated databases and extracted the expression matrix of TRIB3 for multiple bioinformatics analysis. RT-PCR detected the expression of TRIB3 in bladder cancer cells. After knockdown of TRIB3 with siRNA, we investigated TRIB3 function using CCK8, Cell Cycle and Transwell assays.Key FindingsKaplan–Meier analysis of TRIB3 in the four cohorts showed that high expression of TRIB3 correlated with poor outcome. Expression of TRIB3 positively correlated with stage and grade and down-regulation of TRIB3 expression significantly inhibited proliferation, migration and cell cycle of bladder cancer cells.SignificanceTRIB3 is a potential prognostic marker and therapeutic target. It can be used to individualize the treatment of bladder cancer.

Various retinal degenerative disorders manifest in alterations of the AKT/mTOR axis. Despite this, consensus on the therapeutic targeting of mTOR in degenerating retinas has not yet been achieved. Therefore, we investigated the role of AKT/mTOR signaling in rd16 retinas, in which we restored the AKT/mTOR axis by genetic ablation of pseudokinase TRB3, known to inhibit phosphorylation of AKT and mTOR. First, we found that TRB3 ablation resulted in preservation of photoreceptor function in degenerating retinas. Then, we learned that the mTOR downstream cellular pathways involved in the homeostasis of photoreceptors were also reprogrammed in rd16 TRB3−/− retinas. Thus, the level of inactivated translational repressor p-4E-BP1 was significantly increased in these mice along with the restoration of translational rate. Moreover, in rd16 mice manifesting decline in p-mTOR at P15, we found elevated expression of Beclin-1 and ATG5 autophagy genes. Thus, these mice showed impaired autophagy flux measured as an increase in LC3 conversion and p62 accumulation. In addition, the RFP-EGFP-LC3 transgene expression in rd16 retinas resulted in statistically fewer numbers of red puncta in photoreceptors, suggesting impaired late autophagic vacuoles. In contrast, TRIB3 ablation in these mice resulted in improved autophagy flux. The restoration of translation rate and the boost in autophagosome formation occurred concomitantly with an increase in total Ub and rhodopsin protein levels and the elevation of E3 ligase Parkin1. We propose that TRB3 may retard retinal degeneration and be a promising therapeutic target to treat various retinal degenerative disorders.

Glioma is the most common primary malignant brain tumor with poor survival and limited therapeutic options. The non-psychoactive phytocannabinoid cannabidiol (CBD) has been shown to be effective against glioma; however, the molecular target and mechanism of action of CBD in glioma are poorly understood. Here we investigated the molecular mechanisms underlying the antitumor effect of CBD in preclinical models of human glioma. Our results showed that CBD induced autophagic rather than apoptotic cell death in glioma cells. We also showed that CBD induced mitochondrial dysfunction and lethal mitophagy arrest, leading to autophagic cell death. Mechanistically, calcium flux induced by CBD through TRPV4 (transient receptor potential cation channel subfamily V member 4) activation played a key role in mitophagy initiation. We further confirmed TRPV4 levels correlated with both tumor grade and poor survival in glioma patients. Transcriptome analysis and other results demonstrated that ER stress and the ATF4-DDIT3-TRIB3-AKT-MTOR axis downstream of TRPV4 were involved in CBD-induced mitophagy in glioma cells. Lastly, CBD and temozolomide combination therapy in patient-derived neurosphere cultures and mouse orthotopic models showed significant synergistic effect in both controlling tumor size and improving survival. Altogether, these findings showed for the first time that the antitumor effect of CBD in glioma is caused by lethal mitophagy and identified TRPV4 as a molecular target and potential biomarker of CBD in glioma. Given the low toxicity and high tolerability of CBD, we therefore propose CBD should be tested clinically for glioma, both alone and in combination with temozolomide. Abbreviations: 4-PBA: 4-phenylbutyrate; AKT: AKT serine/threonine kinase; ATF4: activating transcription factor 4; Baf-A1: bafilomycin A(1); CANX: calnexin; CASP3: caspase 3; CAT: catalase; CBD: cannabidiol; CQ: chloroquine; DDIT3: DNA damage inducible transcript 3; ER: endoplasmic reticulum; GBM: glioblastoma multiforme; GFP: green fluorescent protein; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; PARP1: poly(ADP-ribose) polymerase; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; SLC8A1: solute carrier family 8 member A1; SQSTM1: sequestosome 1; TCGA: The cancer genome atlas; TEM: transmission electron microscopy; TMZ: temozolomide; TRIB3: tribbles pseudokinase 3; TRPC: transient receptor potential cation channel subfamily C; TRPV4: transient receptor potential cation channel subfamily V member 4

SALL4, a transcriptional factor involved in embryonic stem cell self-renewal and pluripotency, is overexpressed in gastric cancer (GC). However, the association of SALL4 with the survival of GC patients is not well-understood, and the role of SALL4 in cancer progression is still unknown. In the present study, a total of 1,815 GC patients who underwent radical resection at Peking Cancer Hospital were included consecutively from 2015 to 2018, confirming the prognostic value of SALL4 and validating by data from TCGA and GEO. The protein and mRNA expression levels of SALL4 were evaluated by immunohistochemistry and qPCR, respectively. Besides, GSEA and WGCNA were applied to explore the SALL4-related cancer-promoting signaling pathways and gene modules. Our results showed that overexpression of SALL4 was observed in 16.7% of GC patients. SALL4 positivity was associated with male, older age, mixed-type histology, late stages, lymphatic metastasis, vascular invasion, non-cardia location, high AFP level, and no EBV infection background. SALL4 could be served as a marker for prognostic prediction in GC, and SALL4-positive GC was significantly associated with shortened survival. Further, the bioinformatic analysis indicated that the Wnt/β-catenin signaling pathway was activated in SALL4-high cases compared with SALL4-low cases. Expression of SALL4 was also positively correlated with the expression of multiple co-expressed genes, such as TRIB3, which plays an important role in activating the Wnt/β-catenin pathway. Our findings indicate that SALL4 is associated with clinicopathological features related to cancer progression in GC and its function in the Wnt/β-catenin pathway.

The mechanism by which only some individuals infected with Mycobacterium tuberculosis develop necrotic granulomas with progressive disease while others form controlled granulomas that contain the infection remains poorly defined. Mice carrying the sst1-suscepible (sst1(S)) genotype develop necrotic inflammatory lung lesions, similar to human tuberculosis (TB) granulomas, which are linked to macrophage dysfunction, while their congenic counterpart (B6) mice do not. In this study we report that (a) sst1(S) macrophages developed aberrant, biphasic responses to TNF characterized by superinduction of stress and type I interferon pathways after prolonged TNF stimulation; (b) the late-stage TNF response was driven via a JNK/IFN-β/protein kinase R (PKR) circuit; and (c) induced the integrated stress response (ISR) via PKR-mediated eIF2α phosphorylation and the subsequent hyperinduction of ATF3 and ISR-target genes Chac1, Trib3, and Ddit4. The administration of ISRIB, a small-molecule inhibitor of the ISR, blocked the development of necrosis in lung granulomas of M. tuberculosis–infected sst1(S) mice and concomitantly reduced the bacterial burden. Hence, induction of the ISR and the locked-in state of escalating stress driven by the type I IFN pathway in sst1(S) macrophages play a causal role in the development of necrosis in TB granulomas. Interruption of the aberrant stress response with inhibitors such as ISRIB may offer novel host-directed therapy strategies.

© 2021 by the authors., The Tribbles family of pseudokinases controls a wide number of processes during cancer on-set and progression. However, the exact contribution of each of the three family members is still to be defined. Their function appears to be context-dependent as they can act as oncogenes or tumor suppressor genes. They act as scaffolds modulating the activity of several signaling pathways involved in different cellular processes. In this review, we discuss the state-of-knowledge for TRIB1, TRIB2 and TRIB3 in the development and progression of colorectal cancer. We take a perspective look at the role of Tribbles proteins as potential biomarkers and therapeutic targets. Specifically, we chronologically systematized all available articles since 2003 until 2020, for which Tribbles were associated with colorectal cancer human samples or cell lines. Herein, we discuss: (1) Tribbles amplification and overexpression; (2) the clinical significance of Tribbles overexpression; (3) upstream Tribbles gene and protein expression regulation; (4) Tribbles pharmacological modulation; (5) genetic modulation of Tribbles; and (6) downstream mechanisms regulated by Tribbles; establishing a comprehensive timeline, essential to better consolidate the current knowledge of Tribbles’ role in colorectal cancer., This work was supported by the Spanish Ministry of Science, Innovation and Universities through Grant RTI2018-094629-B-I00 to W.L. This work was supported by the European Commission project TRIBBLES–748585 and Fundação para a Ciência e a Tecnologia (FCT) (grant PTDC/MEDONC/4167/2020 “ENDURING”) to B.I.F.

Simple Summary Tribbles proteins possess the structure of protein kinases but function by forming protein complexes rather than phosphorylating substrates. Here we review the structure–function relationship of TRIB1 in cancers. Of the Tribbles proteins, TRIB1 is currently the most well characterised structurally. TRIB1 has different states that could potentially be targeted by small-molecule inhibitors and well-established relevance in acute myeloid leukaemia through degradation of transcription factors. Less is understood about the role of TRIB1 in solid tumours. Further research is required to fully realise the potential of TRIB1 as either a direct target of small-molecule drugs or a biomarker of treatment response across diverse cancer types. Abstract The Tribbles family of proteins—comprising TRIB1, TRIB2, TRIB3 and more distantly related STK40—play important, but distinct, roles in differentiation, development and oncogenesis. Of the four Tribbles proteins, TRIB1 has been most well characterised structurally and plays roles in diverse cancer types. The most well-understood role of TRIB1 is in acute myeloid leukaemia, where it can regulate C/EBP transcription factors and kinase pathways. Structure–function studies have uncovered conformational switching of TRIB1 from an inactive to an active state when it binds to C/EBPα. This conformational switching is centred on the active site of TRIB1, which appears to be accessible to small-molecule inhibitors in spite of its inability to bind ATP. Beyond myeloid neoplasms, TRIB1 plays diverse roles in signalling pathways with well-established roles in tumour progression. Thus, TRIB1 can affect both development and chemoresistance in leukaemia; glioma; and breast, lung and prostate cancers. The pervasive roles of TRIB1 and other Tribbles proteins across breast, prostate, lung and other cancer types, combined with small-molecule susceptibility shown by mechanistic studies, suggests an exciting potential for Tribbles as direct targets of small molecules or biomarkers to predict treatment response.

Endometrial cancer (EC) is the second most common gynecological malignancy worldwide. Tribbles pseudokinase 3 (TRIB3) is a scaffolding protein that regulates intracellular signal transduction, and its role in tumor development is controversial. Here, we investigated the biological function of TRIB3 in EC. We found that the messenger RNA (mRNA) expression level of TRIB3 was significantly and positively correlated with shorter overall survival of EC patients in The Cancer Genome Atlas database. The protein expression of TRIB3 was found to be significantly increased in EC cancer stem cells (CSCs) enriched by tumorsphere cultivation. Knockdown of TRIB3 in EC cells suppressed tumorsphere formation, the expression of cancer stemness genes, and the in vivo tumorigenesis. The expression of &beta, catenin at both the protein and the mRNA levels was downregulated upon TRIB3 silencing. TRIB3 was found to interact with E74 Like ETS transcription factor 4 (ELF4) in the nucleus and bound to ELF4 consensus sites within the catenin beta 1 (CTNNB1) promoter in EC cell lines. These data indicated that TRIB3 may regulate CTNNB1 transcription by enhancing the recruitment of ELF4 to the CTNNB1 promoter. In conclusion, our results suggest that TRIB3 plays an oncogenic role in EC and positively regulates the self-renewal and tumorigenicity of EC-CSCs. Targeting TRIB3 is considered as a potential therapeutic strategy in future EC therapy.

Background Altered lipid metabolism is involved in the development of many tumors. However, the role of dissimilar lipid metabolism in head and neck squamous cell carcinoma (HNSCC) is not fully established. Aims Here, we sought to determine the prognostic value of lipid metabolism‐related genes in HNSCC. Methods RNA‐seq data and clinical features of 545 HNSCC cases were obtained from The Cancer Genome Atlas database. A regulatory network of transcription factors‐lipid metabolism genes and a risk prognostic model of lipid metabolism‐related genes was developed using bioinformatics and Cox regression modeling. We used tumor immune estimation resource to analyze immune cell infiltration in patients with HNSCC based on the prognostic index (PI) of lipid metabolism‐related genes. Results A total of 136 differentially expressed lipid metabolism genes were identified. Of these, 23 are related to prognosis. In addition to predicting HNSCC prognosis, 11 lipid metabolism‐related genes (ARSI, CYP27B1, CYP2D6, DGKG, DHCR7, LPIN1, PHYH, PIP5K1B, PLA2G2D, RDH16, and TRIB3) also affect HNSCC clinical features (stage, gender, and pathological stage). The PI of lipid metabolism‐related genes embodied the state of HNSCC tumor immune microenvironment., Lipid metabolism‐related genes effectively predicts head and neck squamous cell carcinoma (HNSCC) prognosis and reflects the state of immune cell in HNSCC.

This study aimed to investigate the role of vascular insulin resistance (VIR) and Tribbles homolog 3 (TRIB3) in the pathogenesis of hypoxia-induced pulmonary hypertension (HPH). Rats were subjected to low air pressure and low oxygen intermittently for 4 weeks to induce HPH. The mean right ventricular pressure (mRVP), mean pulmonary arterial pressure (mPAP), and right ventricular index (RVI) were significantly increased in HPH rats. Pulmonary arteries from HPH rats showed VIR with reduced vasodilating effect of insulin. The protein levels of peroxisome proliferator-activated receptor gamma (PPARγ), phosphoinositide 3-kinase (PI3K), phosphorylations of Akt, and endothelial nitric oxide (NO) synthase (eNOS) were decreased, and TRIB3 and phosphorylated extracellular signal-regulated protein kinases (ERK1/2) were increased in pulmonary arteries of HPH rats. Early treatment of pioglitazone (PIO) partially reversed the development of HPH, improved insulin-induced vasodilation, and alleviated the imbalance of the insulin signaling. The overexpression of TRIB3 in rat pulmonary arterial endothelial cells (PAECs) reduced the levels of PPARγ, PI3K, phosphorylated Akt (p-Akt), and phosphorylated eNOS (p-eNOS) and increased p-ERK1/2 and the synthesis of endothelin-1 (ET-1), which were further intensified under hypoxic conditions. Moreover, TRIB3 knockdown caused significant improvement in Akt and eNOS phosphorylations and, otherwise, a reduction of ERK1/2 activation in PAECs after hypoxia. In conclusion, impaired insulin-induced pulmonary vasodilation and the imbalance of insulin-induced signaling mediated by TRIB3 upregulation in the endothelium contribute to the development of HPH. Early PIO treatment improves vascular insulin sensitivity that may help to limit the progression of hypoxic pulmonary hypertension.

Advances in the understanding of the Tribbles family of pseudokinases (TRIB1, TRIB2 and TRIB3) reveal these proteins as potentially valuable biomarkers of disease diagnosis, prognosis, prediction and clinical strategy. In their role as signalling mediators and scaffolding proteins, TRIBs lead to changes in protein stability and activity, which impact on diverse cellular processes such as proliferation, differentiation, cell cycle and cell death. We review the role of TRIB proteins as promising therapeutic targets, with an emphasis on their role in cancer, and as biomarkers, with potential application across diverse pathological processes.

Rapalogs have become standard-of-care in patients with metastatic breast, kidney, and neuroendocrine cancers. Nevertheless, tumor escape occurs after several months in most patients, highlighting the need to understand mechanisms of resistance. Using a panel of cancer cell lines, we show that rapalogs downregulate the putative protein kinase TRIB3 (tribbles pseudokinase 3). Blood samples of a small cohort of patients with cancer treated with rapalogs confirmed downregulation of TRIB3. Downregulation of TRIB3 was mediated by LRRFIP1 independently of mTOR and disrupted its interaction with the spliceosome, where it participated in rapalog-induced deregulation of RNA splicing. Conversely, overexpression of TRIB3 in a panel of cancer cell lines abolished the cytotoxic effects of rapalogs. These findings identify TRIB3 as a key component of the spliceosome, whose repression contributes significantly to the mechanism of resistance to rapalog therapy. Significance: Independent of mTOR signaling, rapalogs induce cytoxicity by dysregulating spliceosome function via repression of TRIB3, the loss of which may, in the long term, contribute to therapeutic resistance.

Mounting evidence supports that the malignant phenotypes of cancers are defined not only by the intrinsic activity of tumor cells but also by immune cells that are recruited and activated in tumor-related microenvironment. Here, we developed a diagnostic and prognostic model for colon cancer, based on expression profiles of immune-related genes and immune cell component. As a result, we found that B cell infiltration ratio, CD4+ T cells, as well as immune-related genes of TRIB3, CHGA, CASP7, LGALS4, LEP, NOX4, IL17A, and HSPD1 may be highly relevant with clinical outcome of colon cancer.

Tribbles pseudokinase 3 (TRIB3) is a member of the mammalian pseudokinase tribbles family and is involved in multiple biological processes. However, the role of TRIB3 in renal cell carcinoma (RCC) remains unclear. In this study, we aimed to elucidate the biological functions of TRIB3 in RCC and explore its underlying mechanisms. TRIB3 expression and its correlation with clinicopathological features was evaluated in 123 patients with RCC. A series of cytological experiments were performed to clarify the biological functions of TRIB3, and potential molecular regulatory mechanisms were explored using transcriptome sequencing. TRIB3 expression was significantly elevated in RCC tissues compared to that in paracancerous tissues, and high expression of TRIB3 was correlated with both advanced tumor stage and unfavorable prognosis. TRIB3 knockdown markedly inhibited RCC cell proliferation, migration and invasion. Furthermore, overexpression of TRIB3 promoted RCC cell proliferation, migration, invasion and xenograft tumor growth. Notably, TRIB3 expression was modulated by hypoxia-inducible factor-1α (HIF-1α), which enhanced cell viability and invasiveness via targeting the MAPK signaling pathway. This study reveals the potential oncogenic role of TRIB3 in RCC pathogenesis and illustrates the mechanisms underlying TRIB3-mediated tumor progression, providing new insight into the development of TRIB3 as a tumor biomarker and therapeutic target.

COVID-19 is prevalent in the elderly. Old individuals are more likely to develop pneumonia and respiratory failure due to alveolar damage, suggesting that lung senescence may increase the susceptibility to SARS-CoV-2 infection and replication. Considering that human coronavirus (HCoVs; SARS-CoV-2 and SARS-CoV) require host cellular factors for infection and replication, we analyzed Genotype-Tissue Expression (GTEx) data to test whether lung aging is associated with transcriptional changes in human protein-coding genes that potentially interact with these viruses. We found decreased expression of the gene tribbles homolog 3 (TRIB3) during aging in male individuals, and its protein was predicted to interact with HCoVs nucleocapsid protein and RNA-dependent RNA polymerase. Using publicly available lung single-cell data, we found TRIB3 expressed mainly in alveolar epithelial cells that express SARS-CoV-2 receptor ACE2. Functional enrichment analysis of age-related genes, in common with SARS-CoV-induced perturbations, revealed genes associated with the mitotic cell cycle and surfactant metabolism. Given that TRIB3 was previously reported to decrease virus infection and replication, the decreased expression of TRIB3 in aged lungs may help explain why older male patients are related to more severe cases of the COVID-19. Thus, drugs that stimulate TRIB3 expression should be evaluated as a potential therapy for the disease.

Objectives Hypericin is a polycyclic aromatic naphthodianthrone that occurs naturally. It is also an active ingredient in some species of the genus Hypericum. Emerging evidence suggests that hypericin has attracted great attention as a potential anticancer drug and exhibits remarkable antiproliferative effect upon irradiation on various tumour cells. This paper aims to summarise the anticancer effect and molecular mechanisms modulated by hypericin-medicated photodynamic therapy and its potential role in the cancer treatment. Key findings Hypericin-medicated photodynamic therapy could inhibit the proliferation of various tumour cells including bladder, colon, breast, cervical, glioma, leukaemia, hepatic, melanoma, lymphoma and lung cancers. The effect is primarily mediated by p38 mitogen-activated protein kinase (MAPK), JNK, PI3K, CCAAT-enhancer-binding protein homologous protein (CHOP)/TRIB3/Akt/mTOR, TRAIL/TRAIL-receptor, c-Met and Ephrin-Eph, the mitochondria and extrinsic signalling pathways. Furthermore, hypericin-medicated photodynamic therapy in conjunction with chemotherapeutic agents or targeted therapies is more effective in inhibiting the growth of tumour cells. Summary During the past few decades, the anticancer properties of photoactivated hypericin have been extensively investigated. Hypericin-medicated photodynamic therapy can modulate a variety of proteins and genes and exhibit a great potential to be used as a therapeutic agent for various types of cancer.

Identifying disease-causing pathways and drugs that target them in Parkinson's disease (PD) has remained challenging. We uncovered a PD-relevant pathway in which the stress-regulated heterodimeric transcription complex CHOP/ATF4 induces the neuron prodeath protein Trib3 that in turn depletes the neuronal survival protein Parkin. Here we sought to determine whether the drug adaptaquin, which inhibits ATF4-dependent transcription, could suppress Trib3 induction and neuronal death in cellular and animal models of PD. Neuronal PC12 cells and ventral midbrain dopaminergic neurons were assessed in vitro for survival, transcription factor levels and Trib3 or Parkin expression after exposure to 6-hydroxydopamine or 1-methyl-4-phenylpyridinium with or without adaptaquin co-treatment. 6-hydroxydopamine injection into the medial forebrain bundle was used to examine the effects of systemic adaptaquin on signaling, substantia nigra dopaminergic neuron survival and striatal projections as well as motor behavior. In both culture and animal models, adaptaquin suppressed elevation of ATF4 and/or CHOP and induction of Trib3 in response to 1-methyl-4-phenylpyridinium and/or 6-hydroxydopamine. In culture, adaptaquin preserved Parkin levels, provided neuroprotection and preserved morphology. In the mouse model, adaptaquin treatment enhanced survival of dopaminergic neurons and substantially protected their striatal projections. It also significantly enhanced retention of nigrostriatal function. These findings define a novel pharmacological approach involving the drug adaptaquin, a selective modulator of hypoxic adaptation, for suppressing Parkin loss and neurodegeneration in toxin models of PD. As adaptaquin possesses an oxyquinoline backbone with known safety in humans, these findings provide a firm rationale for advancing it towards clinical evaluation in PD.

Metformin has beneficial effects of preventing and treating cancers on type 2 diabetic patients. However, the role of metformin in non-diabetic cancer patients and the precise molecular mechanisms against cancer have not yet been sufficiently elucidated. We recently reported that the pseudokinase protein TRIB3 acts as a stress sensor linking metabolic stressors to cancer promotion by inhibiting autophagy and ubiquitin-proteasomal degradation systems; genetically abrogating of TRIB3 expression reduces tumourigenesis and cancer progression. Thus, TRIB3 is a potential therapeutic target for diverse cancers. In this study, we found that metformin attenuates melanoma growth and metastasis by reducing TRIB3 expression in non-diabetic C57BL/6 mice and diabetic KK-Ay mice; overexpression of TRIB3 protects metformin from the activation of autophagic flux, the clearance of accumulated tumour-promoting factors and the attenuation of tumour progression. We further elucidated that TRIB3 acts as an adaptor to recruit lysine acetyltransferase 5 (KAT5) to SMAD3 and induce a phosphorylation-dependent K333 acetylation of SMAD3, which sustains transcriptional activity of SMAD3 and subsequently enhances TRIB3 transcription. Metformin suppresses SMAD3 phosphorylation and decreases the KAT5/SMAD3 interaction, to attenuate the KAT5-mediated K333 acetylation of SMAD3, reduce the SMAD3 transcriptional activity and subsequent TRIB3 expression, thereby antagonizes melanoma progression. Together, our study not only defines a molecular mechanism by which metformin protects against melanoma progression by disturbing the KAT5/TRIB3/SMAD3 positive feedback loop in diabetes and non-diabetes mice, but also suggests a candidate diverse utility of metformin in tumour prevention and therapy because of suppressing stress protein TRIB3 expression.

While acute promyelocytic leukemia has a good prognosis with all-trans retinoic acid (ATRA) treatment, ATRA resistance is a major obstacle. It is now demonstrated that TRIBBLES 3 (TRIB3) stabilizes TWIST1, leading to ATRA resistance. Peptides that disrupt this interaction lead to the degradation of TWIST1 and overcome ATRA resistance. See related article by Lin et al., p. 6228

Endoplasmic reticulum (ER) stress has been demonstrated to prompt various cardiovascular risks although the underlying mechanism remains elusive. Protein tyrosine phosphatase-1B (PTP1B) serves as an essential negative regulator for insulin signaling. This study examined the role of PTP1B in ER stress-induced myocardial anomalies and underlying mechanism involved with a focus on autophagy. WT and PTP1B knockout mice were subjected to the ER stress inducer tunicamycin (1mg/kg). Cardiac function was evaluated with echocardiography and an Ion-Optix MyoCam system. Western blot analysis was used to monitor the levels of ER stress, autophagy and insulin signaling including insulin receptor substrate (IRS), tribbles homolog 3 (TRIB3), Atg5/7, p62 and LC3-II. Our results showed that ER stress resulted in compromised echocardiographic and cardiomyocyte contractile function, intracellular Ca2+ mishandling, ER stress, O2- production, apoptosis, the effects of which (with the exception of ER stress) were significantly attenuated or negated by PTP1B ablation. Levels of serine phosphorylation of IRS-1, TRIB3, Atg5/7, LC3B and the autophagy adaptor p62 were significantly upregulated while IRS-1 tyrosine phosphorylation was reduced by tunicamycin, the effect of which were obliterated by PTP1B ablation. In vitro study revealed that the autophagy inducer rapamycin and TRIB3 overexpression cancelled PTP1B ablation-offered beneficial effects on cardiomyocyte function or O2- production in murine cardiomyocytes or H9C2 myoblasts. Antioxidant or gene silencing of TRIB3 mimicked PTP1B ablation-induced protective effects. These findings collectively suggested that PTP1B ablation protects against ER stress-induced cardiac anomalies through regulation of autophagy.

Although bone morphogenetic protein-2 (BMP2) has demonstrated extraordinary potential in bone formation, its clinical applications require supraphysiological milligram-level doses that increase postoperative inflammation and inappropriate adipogenesis, resulting in well-documented life-threatening cervical swelling and cyst-like bone formation. Recent promising alternative biomolecular strategies are toward promoting pro-osteogenic activity of BMP2 while simultaneously suppressing its adverse effects. Here, we demonstrated that small molecular phenamil synergized osteogenesis and bone formation with BMP2 in a rat critical size mandibular defect model. Moreover, we successfully elicited the BMP2 adverse outcomes (i.e. adipogenesis and inflammation) in the mandibular defect by applying high dose BMP2. Phenamil treatment significantly improves the quality of newly formed bone by inhibiting BMP2 induced fatty cyst-like structure and inflammatory soft-tissue swelling. The observed positive phenamil effects were associated with upregulation of tribbles homolog 3 (Trib3) that suppressed adipogenic differentiation and inflammatory responses by negatively regulating PPARγ and NF-κB transcriptional activities. Thus, use of BMP2 along with phenamil stimulation or Trib3 augmentation may be a promising strategy to improve clinical efficacy and safety of current BMP therapeutics.

Numerous animal studies have suggested that sevoflurane can induce neuronal cell death and damage social and emotional behaviors and endoplasmic reticulum stress is a crucial underlying mechanism. The goal of this study was to investigate the role and the mechanisms of long noncoding RNA X-inactive specific transcript (XIST) in sevoflurane-induced social and emotional impairment and neuron apoptosis in neonatal mice. The performance in social and emotional tests and the expression levels of lncRNA-XIST after sevoflurane exposure in neonatal mice and hippocampal neuronal cells were measured. Also, the effects of the suppression of lncRNA-XIST on neuron apoptosis and endoplasmic reticulum stress were determined. Subsequently, the association between lncRNA-XIST and miR-98-5p as well as EDEM1 in hippocampal neuronal cells was explored. Our results showed that social and emotional behaviors were impaired in neonatal mice and lncRNA-XIST and EDEM1were increasingly expressed in hippocampal neuronal cells after sevoflurane exposure and suppression of lncRNA-XIST decreased sevoflurane-induced neuron apoptosis and endoplasmic reticulum stress in vitro. Further in vitro studies showed that overexpression of miR-98-5p decreased apoptosis and endoplasmic reticulum stress while lncRNA Xist negative regulated miR-98-5p expression and the effects of lncRNA Xist on apoptosis and endoplasmic reticulum stress through sponging miR-98-5p. Moreover, suppression of lncRNA-XIST improved sevoflurane-induced social and emotional impairment and alleviated sevoflurane-induced apoptosis and endoplasmic reticulum stress accompanied in neonatal mice. Our findings reveal that knockdown of lncRNA-XIST may contribute to ameliorating sevoflurane-induced social and emotional impairment via inhibiting neuron apoptosis and endoplasmic reticulum stress, which may be achieved by regulating miR-98-5p/EDEM1 axis. Funding Statement: This work was supported by the National Natural Science Foundation of China (81400929, 81471240, 81641042 and 81603545), the Bureau of Chinese Medicine, Zhejiang, China (2018ZB065) and the Innovative Talents Project of Zhejiang province (2016). Declaration of Interests: There is no conflict of interest in the authors. Ethics Approval Statement: The study protocol was approved by the Zhejiang University Institutional review board of Animal studies (ZJU2019-16778). The Guidance Recommendations for Experimental Animal Care and Use of the Ministry of Science and Technology of the People’s Republic of China supervised all experimental procedures.