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The T allele at rs7903146 in TCF7L2 increases the rate of conversion from prediabetes to type 2 diabetes. This has been associated with impaired β-cell function and with defective suppression of α-cell secretion by glucose. However, the temporal relationship of these abnormalities is uncertain. To study the longitudinal changes in islet function, we recruited 128 subjects, with 67 homozygous for the diabetes-associated allele (TT) at rs7903146 and 61 homozygous for the protective allele. Subjects were studied on two occasions, 3 years apart, using an oral 75-g glucose challenge. The oral minimal model was used to quantitate β-cell function; the glucagon secretion rate was estimated from deconvolution of glucagon concentrations. Glucose tolerance worsened in subjects with the TT genotype. This was accompanied by impaired postchallenge glucagon suppression but appropriate β-cell responsivity to rising glucose concentrations. These data suggest that α-cell abnormalities associated with the TT genotype (rs7903146) occur early and may precede β-cell dysfunction in people as they develop glucose intolerance and type 2 diabetes., (© 2024 by the American Diabetes Association.)

Background: Hepatocellular carcinoma (HCC) is one of the most common malignant tumors of the digestive system, and the exact mechanism of HCC is still unclear. Transcription factor 7 like 2 (TCF7L2) plays a pivotal role in cell proliferation and stemness maintenance. However, the exact mechanism of TCF7L2 in HCC remains unclear., Methods: Clinical samples and public databases were used to analyze the expression and prognosis of TCF7L2 in HCC. The function of TCF7L2 in HCC was studied in vitro and in vivo. ChIP and luciferase assays were used to explore the molecular mechanism of TCF7L2. The relationship between TCF7L2 and NEDD9 was verified in HCC clinical samples by tissue microarrays., Results: The expression of TCF7L2 was upregulated in HCC, and high expression of TCF7L2 was associated with poor prognosis of HCC patients. Overexpression of TCF7L2 promoted the metastasis of HCC in vitro and in vivo, while Knockdown of TCF7L2 showed the opposite effect. Mechanically, TCF7L2 activated neural precursor cell expressed developmentally downregulated protein 9 (NEDD9) transcription by binding to the -1522/-1509 site of the NEDD9 promoter region, thereby increasing the phosphorylation levels of AKT and mTOR. The combination of TCF7L2 and NEDD9 could distinguish the survival of HCC patients., Conclusions: This study demonstrated that TCF7L2 promotes HCC metastasis by activating AKT/mTOR pathway in a NEDD9-dependent manner, suggesting that potential of TCF7L2 and NEDD9 as prognostic markers and therapeutic targets for HCC., (© 2024. The Author(s).)

Background: The three-dimensional (3D) genome architecture plays a critical role inregulating gene expression. However, the specific alterations in thisarchitecture within somatotroph tumors and their implications for gene expression remain largely unexplored., Methods: We employed Hi-C and RNA-seq analyses to compare the 3D genomic structures of somatotroph tumors with normal pituitary tissue. This comprehensive approachenabled the characterization of A/B compartments, topologically associateddomains (TADs), and chromatin loops, integrating these with gene expression patterns., Results: We observed a decrease in both the frequency of chromosomal interactions andthe size of TADs in tumor tissue compared to normal tissue. Conversely, the number of TADs and chromatin loops was found to be increased in tumors. Integrated analysis of Hi-C and RNA-seq data demonstrated that changes inhigher-order chromat in structure were associated with alterations in gene expression. Specifically, genes in A compartments showed higher density and increased expression relative to those in B compartments. Moreover, the weakand enhanced insulation boundaries were identified, and the associated genes were enriched in the Wnt/β-Catenin signaling pathway. We identified the gainedand lost loops in tumor and integrated these differences with transcriptional changes to examine the functional relevance of the identified loops. Notably, we observed an enhanced insulation boundary and a greater number of loops in the TCF7L2 gene region within tumors, which was accompanied by an upregulation of TCF7L2 expression. Subsequently, TCF7L2 expression was confirmed through qRT-PCR, and upregulated TCF7L2 prompted cell proliferation and growth hormone (GH) secretion in vitro., Conclusion: Our results provide comprehensive 3D chromatin architecture maps of somatotroph tumors and offer a valuable resource for furthering the understanding of the underlying biology and mechanisms of gene expression regulation., (© 2024 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Deregulation of the Wnt/β-catenin pathway is associated with the development of human cancer including colorectal and liver cancer. Although we previously showed that histidine ammonia lyase (HAL) was transcriptionally reduced by the β-catenin/TCF complex in liver cancer cells, the mechanism(s) of its down-regulation by the complex remain to be clarified. In this study, we search for the transcription factor(s) regulating HAL, and identify CEBPA and FOXA1, two factors whose expression is suppressed by the knockdown of β-catenin or TCF7L2. In addition, RNA-seq analysis coupled with genome-wide mapping of CEBPA- and FOXA1-binding regions reveals that these two factors also increase the expression of arginase 1 (ARG1) that catalyzes the hydrolysis of arginine. Metabolome analysis discloses that activated Wnt signaling augments intracellular concentrations of histidine and arginine, and that the signal also increases the level of lactic acid suggesting the induction of the Warburg effect in liver cancer cells. Further analysis reveals that the levels of metabolites of the urea cycle and genes coding its related enzymes are also modulated by the Wnt signaling. These findings shed light on the altered cellular metabolism in the liver by the Wnt/β-catenin pathway through the suppression of liver-enriched transcription factors including CEBPA and FOXA1., (© 2024. The Author(s).)

Background: Paired box 1 (PAX1) is a transcription factor and essential for the development of pharyngeal pouches-derived tissues, including thymus. PAX1 mutations are identified in Severe Combined Immunodeficiency (SCID) patients with Otofaciocervical Syndrome Type 2 (OTFCS2). However, despite the critical roles of PAX1 in embryonic development and diseases, detailed insights into its molecular mode of action are critically missing., Methods: The repressing roles of PAX1 and SCID associated mutants on Wnt signaling pathway were investigated by luciferase reporter assays, qRT-PCR and in situ hybridization in HEK293FT, HCT116 cells and zebrafish embryos, respectively. Co-immunoprecipitation (co-IP) and western blotting assays were carried out to identify the molecular mechanisms underlying PAX1's role on Wnt signaling pathway. hESC based endoderm differentiation, flow cytometry, high-throughput sequencing data analysis, and qRT-PCR assays were utilized to determine the roles of PAX1 during endoderm differentiation., Results: Here, we show that PAX1 represses canonical Wnt signaling pathway in vertebrate cells. Mechanically, PAX1 competes with SUMO E3 ligase PIASy to bind to TCF7L2, thus perturbing TCF7L2 SUMOylation level, further reducing its transcriptional activity and protein stability. Moreover, we reveal that PAX1 plays dual roles in hESC-derived definitive and foregut/pharyngeal endoderm cells, which give rise to the thymus epithelium, by inhibiting Wnt signaling. Importantly, our data show PAX1 mutations found in SCID patients significantly compromise the suppressing ability of PAX1 on Wnt signaling., Conclusions: Our study presents a novel molecular mode of action of PAX1 in regulation of canonical Wnt signaling and endoderm differentiation, thus providing insights for the molecular basis of PAX1 associated SCID, offering better understanding of the behavior of PAX1 in embryogenesis., (© 2024. The Author(s).)

The Wnt/β-catenin pathway contains multiple high-confidence risk genes that are linked to neurodevelopmental disorders, including autism spectrum disorder. However, its ubiquitous roles across brain cell types and developmental stages have made it challenging to define its impact on neural circuit development and behavior. Here, we show that TCF7L2, which is a key transcriptional effector of the Wnt/β-catenin pathway, plays a cell-autonomous role in postnatal astrocyte maturation and impacts adult social behavior. TCF7L2 was the dominant Wnt effector that was expressed in both mouse and human astrocytes, with a peak during astrocyte maturation. The conditional knockout of Tcf7l2 in postnatal astrocytes led to an enlargement of astrocytes with defective tiling and gap junction coupling. These mice also exhibited an increase in the number of cortical excitatory and inhibitory synapses and a marked increase in social interaction by adulthood. These data reveal an astrocytic role for developmental Wnt/β-catenin signaling in restricting excitatory synapse numbers and regulating adult social behavior., (© 2023. The Author(s).)

Colorectal carcinoma (CRC), one of the most life-threatening cancer types, is associated with aberrant expression of epigenetic modifiers and activation of the Wnt pathway. However, the role of epigenetic regulators in driving cancer cell proliferation and their potential as therapeutic targets affecting the Wnt pathway remain unclear. In this study, BRD4 was found to promote the progression of CRC both in vitro and in vivo. The expression of BRD4 correlated with shortened CRC patient survival. In addition, BRD4 function was strongly correlated with the Wnt pathway, but rather through regulation of TCF7L2 at transcriptional levels. BRD4 and H3K27ac have overlapping occupancies in the cis-regulatory elements of TCF7L2, suggesting enhancer-based epigenetic regulation. Numerous YY1 binding sites were found in the abovementioned region. YY1 recruited BRD4 to bind to cis-regulatory elements of TCF7L2, thereby regulating the expression of TCF7L2. Altogether, this study validates that BRD4 performs a canonical epigenetic regulatory function in CRC and can be used in the treatment of Wnt pathway-dependent CRC or other malignancies with clinically available bromodomain inhibitors., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

At the top of the midbrain is the inferior colliculus (IC), which functions as the major hub for processing auditory information. Despite the functional significance of neurons in the IC, our understanding of their formation is limited. In this study, we identify the embryonic patterning gene Dbx1 as a key molecular player that governs genetic programs for IC survival. We find that Dbx1 plays a critical role in preventing apoptotic cell death in postnatal IC by transcriptionally repressing c-Jun and pro-apoptotic BH3 only factors. Furthermore, by employing combined approaches, we uncover that Tcf7l2 functions downstream of Dbx1. Loss of Tcf7l2 function causes IC phenotypes with striking similarity to those of Dbx1 mutant mice, which include defective embryonic maturation and postnatal deletion of the IC. Finally, we demonstrate that the Dbx1-Tcf7l2 cascade functions upstream of Ap-2δ, which is essential for IC development and survival. Together, these results unravel a novel molecular mechanism for IC maintenance, which is indispensable for normal brain development., (© 2023. The Author(s), under exclusive licence to ADMC Associazione Differenziamento e Morte Cellulare.)

Aims/hypothesis: Non-alcoholic fatty liver disease (NAFLD) associated with type 2 diabetes may more easily progress towards severe forms of non-alcoholic steatohepatitis (NASH) and cirrhosis. Although the Wnt effector transcription factor 7-like 2 (TCF7L2) is closely associated with type 2 diabetes risk, the role of TCF7L2 in NAFLD development remains unclear. Here, we investigated how changes in TCF7L2 expression in the liver affects hepatic lipid metabolism based on the major risk factors of NAFLD development., Methods: Tcf7l2 was selectively ablated in the liver of C57BL/6N mice by inducing the albumin (Alb) promoter to recombine Tcf7l2 alleles floxed at exon 5 (liver-specific Tcf7l2-knockout [KO] mice: Alb-Cre;Tcf7l2 f/f ). Alb-Cre;Tcf7l2 f/f and their wild-type (Tcf7l2 f/f ) littermates were fed a high-fat diet (HFD) or a high-carbohydrate diet (HCD) for 22 weeks to reproduce NAFLD/NASH. Mice were refed a standard chow diet or an HCD to stimulate de novo lipogenesis (DNL) or fed an HFD to provide exogenous fatty acids. We analysed glucose and insulin sensitivity, metabolic respiration, mRNA expression profiles, hepatic triglyceride (TG), hepatic DNL, selected hepatic metabolites, selected plasma metabolites and liver histology., Results: Alb-Cre;Tcf7l2 f/f essentially exhibited increased lipogenic genes, but there were no changes in hepatic lipid content in mice fed a normal chow diet. However, following 22 weeks of diet-induced NAFLD/NASH conditions, liver steatosis was exacerbated owing to preferential metabolism of carbohydrate over fat. Indeed, hepatic Tcf7l2 deficiency enhanced liver lipid content in a manner that was dependent on the duration and amount of exposure to carbohydrates, owing to cell-autonomous increases in hepatic DNL. Mechanistically, TCF7L2 regulated the transcriptional activity of Mlxipl (also known as ChREBP) by modulating O-GlcNAcylation and protein content of carbohydrate response element binding protein (ChREBP), and targeted Srebf1 (also called SREBP1) via miRNA (miR)-33-5p in hepatocytes. Eventually, restoring TCF7L2 expression at the physiological level in the liver of Alb-Cre;Tcf7l2 f/f mice alleviated liver steatosis without altering body composition under both acute and chronic HCD conditions., Conclusions/interpretation: In mice, loss of hepatic Tcf7l2 contributes to liver steatosis by inducing preferential metabolism of carbohydrates via DNL activation. Therefore, TCF7L2 could be a promising regulator of the NAFLD associated with high-carbohydrate diets and diabetes since TCF7L2 deficiency may lead to development of NAFLD by promoting utilisation of excess glucose pools through activating DNL., Data Availability: RNA-sequencing data have been deposited into the NCBI GEO under the accession number GSE162449 ( www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE162449 )., (© 2023. The Author(s).)

Vocalization is an essential medium for social signaling in birds and mammals. Periaqueductal gray (PAG) a conserved midbrain structure is believed to be responsible for innate vocalizations, but its molecular regulation remains largely unknown. Here, through a mouse forward genetic screening we identified one of the key Wnt/β-catenin effectors TCF7L2/TCF4 controls ultrasonic vocalization (USV) production and syllable complexity during maternal deprivation and sexual encounter. Early developmental expression of TCF7L2 in PAG excitatory neurons is necessary for the complex trait, while TCF7L2 loss reduces neuronal gene expressions and synaptic transmission in PAG. TCF7L2-mediated vocal control is independent of its β-catenin-binding domain but dependent of its DNA binding ability. Patient mutations associated with developmental disorders, including autism spectrum disorders, disrupt the transcriptional repression effect of TCF7L2, while mice carrying those mutations display severe USV impairments. Therefore, we conclude that TCF7L2 orchestrates gene expression in midbrain to control vocal production through its DNA binding but not transcription activation domain., (© 2023. The Author(s).)

Clinical and basic neuroscience research is greatly benefited from the identification and characterization of lineage specific and developmental stage-specific markers. In the glial research community, histological markers that specifically label newly differentiated premyelinating oligodendrocytes are still scarce. Premyelinating oligodendrocyte markers, especially those of nuclear localization, enable researchers to easily quantify the rate of oligodendrocyte generation regardless of developmental ages. We propose that the transcription factor 7-like 2 (TCF7l2, mouse gene symbol Tcf7l2) is a useful nuclear marker that specifically labels newly generated premyelinating oligodendrocytes and promotes oligodendroglial lineage progression. Here, we highlight the controversial research history of TCF7l2 expression and function in oligodendroglial field and discuss previous experimental data justifying TCF7l2 as a specific nuclear marker for premyelinating oligodendrocytes during developmental myelination and remyelination. We conclude that TCF7l2 can be used alone or combined with pan-oligodendroglial lineage markers to identify newly differentiated or newly regenerated oligodendrocytes and quantify the rate of oligodendrocyte generation., (© 2022 Wiley Periodicals LLC.)

It is a great challenge to develop a safe and effective treatment strategy for age-related osteoporosis and fracture healing. As one of the four FOXO transcription factors, FOXO1 is essential for cell proliferation, survival, senescence, energy metabolism, and oxidative stress in various cells. Our previous study demonstrated that specific Foxo1 gene deletion in osteoblasts in young mice results in bone loss while that in aged mice shows the opposite effect. However, the mechanism underlying the differential regulation of bone metabolism by FOXO1 remains to be elucidated. In this study, we generated osteoblast-specific Foxo1 knockout mice by using Foxo1 fl/fl and Bglap-Cre mice. In young mice, Foxo1 gene deletion inhibits osteoblast differentiation, leading to a decreased osteoblast number and decreased bone formation rate because of the weakened ability to resist oxidative stress, eventually resulting in bone loss and delayed healing of bone defects. In aged mice, high levels of reactive oxygen species (ROS) promote the diversion of CTNNB1 (β-catenin) from T cell factor 4 (TCF4)- to FOXO1-mediated transcription, thereby inhibiting Wnt/β-catenin signaling and leading to decreased osteogenic activity. Conversely, FOXO1 deficiency indirectly promotes the binding of β-catenin and TCF4 and activates Wnt/β-catenin signaling, thereby alleviating age-related bone loss and improving bone defect healing. Our study proves that FOXO1 has differential effects on bone metabolism in young and aged mice and elucidates its underlying mechanism. Further, this study provides a new perspective on the treatment of age-related osteoporosis., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Overexpression of ubiquitin-specific protease 28 (USP28) is found in hepatic carcinoma. It is unclear whether the deubiquitinase plays a role in hepatocarcinogenesis. Deregulation of the Wnt signaling pathway is frequently associated with liver cancer. Transcription factor 7-like 2 (TCF7L2) is an important downstream transcription factor of the Wnt/β-catenin signaling pathway, but the mechanisms by which TCF7L2 itself is regulated have not yet been revealed. Here, we report that USP28 promotes the activity of the Wnt signaling pathway through maintaining the stability of TCF7L2. We further show that FBXW7 is the E3 ubiquitin ligase for TCF7L2. By regulating the levels of TCF7L2, USP28 modulates the Wnt/β-catenin signaling in liver cancer and USP28 depletion or inhibition by a small molecule inhibitor leads to a halt of growth in liver cancer cells. These results suggest that USP28 could be a potential therapeutic target for liver cancer., (© 2022 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Background: NF-κB signaling is widely linked to the pathogenesis and treatment resistance in cancers. Increasing attention has been paid to its anti-oncogenic roles, due to its key functions in cellular senescence and the senescence-associated secretory phenotype (SASP). Therefore, thoroughly understanding the function and regulation of NF-κB in cancers is necessary prior to the application of NF-κB inhibitors., Methods: We established glioblastoma (GBM) cell lines expressing ectopic TCF4N, an isoform of the β-catenin interacting transcription factor TCF7L2, and evaluated its functions in GBM tumorigenesis and chemotherapy in vitro and in vivo. In p65 knock-out or phosphorylation mimic (S536D) cell lines, the dual role and correlation of TCF4N and NF-κB signaling in promoting tumorigenesis and chemosensitivity was investigated by in vitro and in vivo functional experiments. RNA-seq and computational analysis, immunoprecipitation and ubiquitination assay, minigene splicing assay and luciferase reporter assay were performed to identify the underlying mechanism of positive feedback regulation loop between TCF4N and the p65 subunit of NF-κB. A eukaryotic cell-penetrating peptide targeting TCF4N, 4N, was used to confirm the therapeutic significance., Results: Our results indicated that p65 subunit phosphorylation at Ser 536 (S536) and nuclear accumulation was a promising prognostic marker for GBM, and endowed the dual functions of NF-κB in promoting tumorigenesis and chemosensitivity. p65 S536 phosphorylation and nuclear stability in GBM was regulated by TCF4N. TCF4N bound p65, induced p65 phosphorylation and nuclear translocation, inhibited its ubiquitination/degradation, and subsequently promoted NF-κB activity. p65 S536 phosphorylation was essential for TCF4N-led senescence-independent SASP, GBM tumorigenesis, tumor stem-like cell differentiation and chemosensitivity. Activation of p65 was closely connected to alterative splicing of TCF4N, a likely positive feedback regulation loop between TCF4N and p65 in GBM. 4N increased chemosensitivity, highlighting a novel anti-cancer strategy., Conclusion: Our study defined key roles of TCF4N as a novel regulator of NF-κB through mutual regulation with p65 and provided a new avenue for GBM inhibition., (© 2022 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Huntington's disease (HD) is characterized by defective oligodendroglial differentiation and white matter disease. Here, we investigate the role of oligodendrocyte progenitor cell (OPC) dysfunction in adult myelin maintenance in HD. We first note a progressive, age-related loss of myelin in both R6/2 and zQ175 HD mice compared with wild-type controls. Adult R6/2 mice then manifest a significant delay in remyelination following cuprizone demyelination. RNA-sequencing and proteomic analysis of callosal white matter and OPCs isolated from both R6/2 and zQ175 mice reveals a systematic downregulation of genes associated with oligodendrocyte differentiation and myelinogenesis. Gene co-expression and network analysis predicts repressed Tcf7l2 signaling as a major driver of this expression pattern. In vivo Tcf7l2 overexpression restores both myelin gene expression and remyelination in demyelinated R6/2 mice. These data causally link impaired TCF7L2-dependent transcription to the poor development and homeostatic retention of myelin in HD and provide a mechanism for its therapeutic restoration., Competing Interests: Declaration of interests S.A.G. is a part-time employee and stockholder of Sana Biotechnology, a cell therapy company, and his lab receives sponsored research support from Sana. S.A.G. is also a co-founder and adviser to CNS2 Therapeutics, another cell therapy company. None of the work described in this paper overlaps with his work with those companies. D.C.-M. is also a consultant to Sana., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

To explore the role and possible mechanism of miRNA-212 in heart failure (HF). The rat model of abdominal aortic constriction was constructed, the changes of myocardial morphology were observed by hematoxylin-eosin (HE) staining, and the hypertrophy-related marker molecules were detected by quantitative real-time polymerase chain reaction (qRT-PCR). At the cellular level, phenylephrine and angiotensin II were added to induce cardiomyocyte hypertrophy. The overexpression of miR-212 adenovirus was constructed, and the expression of miR-212 was overexpressed, and its effect on cardiac hypertrophy (CH) was detected by immunofluorescence and qRT-PCR. Then, the mechanism of miR-212 regulating CH was verified by website prediction, luciferase reporter gene assay, qRT-PCR, and western blotting assay. In the successfully constructed rat model of abdominal aortic constriction and cardiomyocyte hypertrophy, ANP and myh7 were dramatically increased, myh6 expression was decreased, and miRNA-212 expression was increased. Overexpression of miRNA-212 in cardiomyocytes can promote cardiomyocyte hypertrophy, while knocking down miR-212 in cardiomyocytes can partially reverse cell hypertrophy. In addition, miR-212 targets TCF7L2 and inhibits the expression of this gene. miRNA-212 targets TCF7L2 and inhibits the expression of this gene, possibly through this pathway to promote cardiomyocyte hypertrophy., Competing Interests: The authors declare that they have no competing interests., (Copyright © 2022 Jinxia Yuan and Guoliang Yuan.)

Mutations in the human kinesin family member 5A (KIF5A) gene were recently identified as a genetic cause of amyotrophic lateral sclerosis (ALS). Several KIF5A ALS variants cause exon 27 skipping and are predicted to produce motor proteins with an altered C-terminal tail (referred to as ΔExon27). However, the underlying pathogenic mechanism is still unknown. Here, we confirm the expression of KIF5A mutant proteins in patient iPSC-derived motor neurons. We perform a comprehensive analysis of ΔExon27 at the single-molecule, cellular, and organism levels. Our results show that ΔExon27 is prone to form cytoplasmic aggregates and is neurotoxic. The mutation relieves motor autoinhibition and increases motor self-association, leading to drastically enhanced processivity on microtubules. Finally, ectopic expression of ΔExon27 in Drosophila melanogaster causes wing defects, motor impairment, paralysis, and premature death. Our results suggest gain-of-function as an underlying disease mechanism in KIF5A-associated ALS., (© 2022 The Authors.)

Diabetic retinopathy (DR) is one of the most common blindness in working-age adults. Transcription factor 7 like 2 (TCF7L2) is a susceptibility gene of DR, however, its roles in the pathogenesis of DR are still largely unknown. In this study, we found that TCF7L2 was mainly located in the cell nucleus of retinal ganglion cell layer (GCL) and inner nuclear layer (INL), while it was not expressed in the cell nucleus of retinal outer nuclear layer (ONL). Expression of TCF7L2 was significantly elevated in the retinas of db/db diabetic mice and oxygen-induced retinopathy (OIR) mice. Also, in Ad-hTCF7L2 treated hiPSCs-derived retinal progenitor cells (RPCs), activating transcription factor 6 (ATF6)-related endoplasmic reticulum (ER) stress signaling was remarkably activated. Moreover, knockdown of TCF7L2 significantly inhibited ATF6-related ER stress signaling. Furthermore, the data of endothelial permeability assay showed that RPCs pretreated with Ad-hTCF7L2 lead to enhanced monolayer permeability of human umbilical vein endothelial cells (HUVECs), and knockdown of TCF7L2 or ATF6 in RPCs could alleviate the monolayer permeability of HUVECs. Thus, our results showed that TCF7L2 could trigger ATF6-related ER stress signaling and promote vein endothelial cell permeability, which will provide important insight into the role of TCF7L2 in the pathogenesis of DR and contribute to designing potential therapies., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Introduction: Non-coding genetic variation at TCF7L2 is the strongest genetic determinant of type 2 diabetes (T2D) risk in humans. TCF7L2 encodes a transcription factor mediating the nuclear effects of WNT signaling in adipose tissue (AT). In vivo studies in transgenic mice have highlighted important roles for TCF7L2 in adipose tissue biology and systemic metabolism., Objective: To map the expression of TCF7L2 in human AT, examine its role in human adipose cell biology in vitro, and investigate the effects of the fine-mapped T2D-risk allele at rs7903146 on AT morphology and TCF7L2 expression., Methods: Ex vivo gene expression studies of TCF7L2 in whole and fractionated human AT. In vitro TCF7L2 gain- and/or loss-of-function studies in primary and immortalized human adipose progenitor cells (APCs) and mature adipocytes (mADs). AT phenotyping of rs7903146 T2D-risk variant carriers and matched controls., Results: Adipose progenitors (APs) exhibited the highest TCF7L2 mRNA abundance compared to mature adipocytes and adipose-derived endothelial cells. Obesity was associated with reduced TCF7L2 transcript levels in whole subcutaneous abdominal AT but paradoxically increased expression in APs. In functional studies, TCF7L2 knockdown (KD) in abdominal APs led to dose-dependent activation of WNT/β-catenin signaling, impaired proliferation and dose-dependent effects on adipogenesis. Whilst partial KD enhanced adipocyte differentiation, near-total KD impaired lipid accumulation and adipogenic gene expression. Over-expression of TCF7L2 accelerated adipogenesis. In contrast, TCF7L2-KD in gluteal APs dose-dependently enhanced lipid accumulation. Transcriptome-wide profiling revealed that TCF7L2 might modulate multiple aspects of AP biology including extracellular matrix secretion, immune signaling and apoptosis. The T2D-risk allele at rs7903146 was associated with reduced AP TCF7L2 expression and enhanced AT insulin sensitivity., Conclusions: TCF7L2 plays a complex role in AP biology and has both dose- and depot-dependent effects on adipogenesis. In addition to regulating pancreatic insulin secretion, genetic variation at TCF7L2 might also influence T2D risk by modulating AP function., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Gastric cancer (GC) is the fifth most frequently diagnosed malignant tumour and the third leading cause of cancer-related death. EBV infection is one of the major causes of GC, accounting for approximately 10% of GC cases. Studies revealed that EBV infection could induce malignancy through multilevel mechanisms, including epigenetics state alterations, miRNA regulations, etc. However, it is poorly understood if EBV positive GC has any characteristic oncogenic singling activation. By comparing the transcriptomic of EB positive and negative cell lines, TCGA GC samples, and GC single cells, we identified that canonical WNT signalling is commonly activated in EBV positive GC. Next, we proved that the WNT activation is crucial for the invasiveness of the GC via regulating epithelial-mesenchymal transition (EMT) gene expressions. Then we confirmed that the WNT signalling activation in EBV + GC is through the WNT/CTNNB1 (β-catenin)/TCF7L2 axis with the expression of TCF7L2 indicating worse outcome and more metastasis in EBV+ GCs. Finally, we found that KLF5 is potentially involved as a cooperative transcription factor in the early phase of the EMT process through single-cell pseudotime analysis., Competing Interests: Declaration of competing interest None., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Oligodendrocyte (OL) death and remyelination failure lead to progressive neurological deficits in multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). Matrine (MAT), a quinolizidine alkaloid component derived from the root of Sophora flavescens, has the capacity to effectively inhibit central nervous system (CNS) inflammation and to promote neuroregeneration. In the present study we explored its regulatory mechanism on the Wnt/β-catenin/TCF7L2 pathway, a negative modulator for myelination, in MOG35--55 peptide-induced EAE. Our results clearly indicate that MAT treatment reduced the activation of Wnt3a and β-catenin in the CNS of EAE mice, accompanied by the activation of GSK3β and decreased expression of cyclin D1 and Axin2, two target genes of the Wnt3a/β-catenin pathway. In addition, MAT increased OL maturation and myelination, as evidenced by the decreased number of NG2 + Olig2 + cells and the increased numbers of MBP + and CC1 + Olig2 + cells. Taken together, these findings indicate that MAT treatment promoted the maturation of OLs and myelin repair, which is closely related to the modulation of the Wnt/β-catenin/TCF7L2 signaling pathway., (Copyright © 2022 Elsevier B.V. All rights reserved.)

T-cell-like factor (TCF)7l2, a key effector of canonical Wnt signaling, is highly expressed in bone but nothing is known about its role in regulating osteoblast function. To test this, we generated mice with conditional disruption of Tcf7l2 gene in osteoblast lineages using Tcf7l2 floxed and Col1α2 -Cre mice. Skeletal parameters were evaluated using heterozygous conditional knockdown (HCKD) mice since homozygous conditional knockout died during pregnancy or immediately after birth. At 5 wk of age, trabecular bone mass of long bones was reduced by 35% as measured by microcomputed tomography (μCT). Histology data showed a 42% reduction in femur trabecular bone mass caused by reduced bone formation. Knockdown of Tcf7l2 expression in osteoblasts decreased proliferation and differentiation by 20%-40%. Expression levels of genes ( Hif1α , Vegf , and β-catenin) targeted by TCF7L2 were decreased by 50% in Tcf7l2 -deficient osteoblasts and bones of HCKD mice. We found that the Hif1α gene promoter contained multiple putative TCF7L2 motifs and stabilization of HIF1α protein levels rescued expression of TCF7L2 target genes and alkaline phosphatase (ALP) activity in Tcf7l2 -deficient osteoblasts. Furthermore, Tcf7l2 overexpression increased proliferation in the presence of canonical Wnt3a that was not affected by β-catenin inhibitor providing evidence for a noncanonical signaling in mediating TCF7L2 effects. Tcf7l2 expression was increased in response to mechanical strain (MS) in vitro and in vivo, and disruption of Tcf7l2 expression in osteoblasts reduced MS-induced ALP activity by 35%. We conclude that Tcf7l2 , a mechanoresponsive gene, is an important regulator of osteoblast function acting, in part, via hypoxia signaling. NEW & NOTEWORTHY TCF7L2 is expressed by bone but it was not known whether TCF7L2 expression influenced bone development. By using a mouse model with conditional disruption of Tcf7l2 in osteoblast lineage cells, we have demonstrated for the first time, that TCF7L2 plays an important role in regulating osteoblasts via a noncanonical pathway.

Nucleus pulposus (NP) cells play a critical role in maintaining intervertebral disc integrity through producing the components of extracellular matrix (ECM). NP cell dysfunction, including senescence and hyper-apoptosis, has been regarded as critical events during intervertebral disc degeneration development. In the present study, we found that Transcription Factor 7-Like 2 (TCF7L2) was overexpressed within degenerative intervertebral disc tissue samples, and TCF7L2 silencing improved lipopolysaccharide (LPS)-induced repression on NP cell proliferation, ECM synthesis, and LPS-induced NP cell senescence. miR-1260b directly targeted TCF7L2 and inhibited TCF7L2 expression. miR-1260b overexpression improved LPS-induced degenerative changes in NP cells; more importantly, TCF7L2 overexpression significantly reversed the effects of miR-1260b overexpression on LPS-stimulated degenerative changes within NP cells. For the first time, we demonstrated the function of the miR-1260b/TCF7L2 axis on the phenotypic maintenance of chondrocyte-like NP cells and ECM synthesis by NP cells under LPS stimulation., (© 2022. The Author(s) under exclusive licence to Japan Human Cell Society.)

Various studies have manifested that microRNAs (miRNAs) are involved in the modulation of the occurrence and development of osteosarcoma (OS). However, whether miR-22-3p is associated with OS growth remains unclear. In the study, the potential molecular mechanisms of miR-22-3p in OS was explored. It was affirmed that miR-22-3p was associated with distant metastasis and tumor size in OS patients, and reduced in OS tissues and cells while transcription factor 7-like 2 (TCF7L2) was elevated. Elevated miR-22-3p repressed OS cell progression, and the Wnt/β-catenin pathway, while elevated TCF7L2 was opposite. MiR-22-3p targeted TCF7L2 in OS. In functional rescue experiments, knockdown of miR-22-3p on OS progression and promotion of Wnt/β-catenin were reversed by simultaneous knockdown of TCF7L2. Transplantation experiments in nude mice showed that elevated miR-22-3p repressed OS tumor growth and decreased TCF7L2, Wnt and β-catenin. Shortly, this study suggest that miR-22-3p refrains the Wnt/β-catenin pathway by targeting TCF7L2 and thereby preventing OS deterioration. MiR-22-3p/TCF7L2 axis is supposed to be a candidate molecular target for future OS treatment.

Background: Epithelial-to-mesenchymal transition (EMT) is a process linked to metastasis and drug resistance with non-coding RNAs (ncRNAs) playing pivotal roles. We previously showed that miR-100 and miR-125b, embedded within the third intron of the ncRNA host gene MIR100HG, confer resistance to cetuximab, an anti-epidermal growth factor receptor (EGFR) monoclonal antibody, in colorectal cancer (CRC). However, whether the MIR100HG transcript itself has a role in cetuximab resistance or EMT is unknown., Methods: The correlation between MIR100HG and EMT was analyzed by curating public CRC data repositories. The biological roles of MIR100HG in EMT, metastasis and cetuximab resistance in CRC were determined both in vitro and in vivo. The expression patterns of MIR100HG, hnRNPA2B1 and TCF7L2 in CRC specimens from patients who progressed on cetuximab and patients with metastatic disease were analyzed by RNAscope and immunohistochemical staining., Results: The expression of MIR100HG was strongly correlated with EMT markers and acted as a positive regulator of EMT. MIR100HG sustained cetuximab resistance and facilitated invasion and metastasis in CRC cells both in vitro and in vivo. hnRNPA2B1 was identified as a binding partner of MIR100HG. Mechanistically, MIR100HG maintained mRNA stability of TCF7L2, a major transcriptional coactivator of the Wnt/β-catenin signaling, by interacting with hnRNPA2B1. hnRNPA2B1 recognized the N6-methyladenosine (m 6 A) site of TCF7L2 mRNA in the presence of MIR100HG. TCF7L2, in turn, activated MIR100HG transcription, forming a feed forward regulatory loop. The MIR100HG/hnRNPA2B1/TCF7L2 axis was augmented in specimens from CRC patients who either developed local or distant metastasis or had disease progression that was associated with cetuximab resistance., Conclusions: MIR100HG and hnRNPA2B1 interact to control the transcriptional activity of Wnt signaling in CRC via regulation of TCF7L2 mRNA stability. Our findings identified MIR100HG as a potent EMT inducer in CRC that may contribute to cetuximab resistance and metastasis by activation of a MIR100HG/hnRNPA2B1/TCF7L2 feedback loop., (© 2022. The Author(s).)

FGF19/FGF15 is an endocrine regulator of hepatic bile salt and lipid metabolism, which has shown promising effects in the treatment of NASH in clinical trials. FGF19/15 is transcribed and released from enterocytes of the small intestine into enterohepatic circulation in response to bile-induced FXR activation. Previously, the TSS of FGF19 was identified to bind Wnt-regulated TCF7L2/encoded transcription factor TCF4 in colorectal cancer cells. Impaired Wnt signaling and specifical loss of function of its coreceptor LRP6 have been associated with NASH. We, therefore, examined if TCF7L2/TCF4 upregulates Fgf19 in the small intestine and restrains NASH through gut-liver crosstalk. We examined the mice globally overexpressing, haploinsufficient, and conditional knockout models of TCF7L2 in the intestinal epithelium. The TCF7L2 +/- mice exhibited increased plasma bile salts and lipids and developed diet-induced fatty liver disease while mice globally overexpressing TCF7L2 were protected against these traits. Comprehensive in vivo analysis revealed that TCF7L2 transcriptionally upregulates FGF15 in the gut, leading to reduced bile synthesis and diminished intestinal lipid uptake. Accordingly, Vilin Creert2 ; Tcf7L2 fl/fl mice showed reduced Fgf19 in the ileum, and increased plasma bile. The global overexpression of TCF7L2 in mice with metabolic syndrome-linked LRP6 R611C substitution rescued the fatty liver and fibrosis in the latter. Strikingly, the hepatic levels of TCF4 were reduced and CYP7a1 was increased in human NASH, indicating the relevance of TCF4-dependent regulation of bile synthesis to human disease. These studies identify the critical role of TCF4 as an upstream regulator of the FGF15-mediated gut-liver crosstalk that maintains bile and liver triglyceride homeostasis., (© 2022 Federation of American Societies for Experimental Biology.)

Epithelial-mesenchymal transition (EMT) is thought to be involved in the tissue remodeling and long-term inflammatory process of chronic sinusitis (CRS), but the driving mechanism is still unclear. Using high-resolution mass spectrometry, we performed a proteomic screen of CRS nasal mucosal tissue to identify differentially expressed proteins. Data are available via ProteomeXchange with identifier PXD030884. Specifically, we identified S100 calcium binding protein A4 (S100A4), an effective factor in inflammation-related diseases, and its downstream protein closely related to tissue fibrosis collagen type I alpha 1 chain (COL1A1), which suggested its involvement in nasal mucosal tissue remodeling. In addition, stimulation of human nasal epithelial cells (HNEpCs) with lipopolysaccharide (LPS) mimicked the inflammatory environment of CRS and showed that S100A4 is involved in regulating EMT and thus accelerating tissue remodeling in the nasal mucosa, both in terms of increased cell motility and overexpression of mesenchymal-type proteins. Additionally, we further investigated the regulation mechanism of S100A4 involved in EMT in CRS. Our research results show that in the inflammatory environment of CRS nasal mucosal epithelial cells, TCF-4 will target to bind to S100A4 and regulate its transcription. The transcription of S100A4 in turn affects the execution of the important signaling pathway in EMT, the Wnt/GSK-3β/β-catenin pathway, through the TCF-4/β-catenin complex. In conclusion, this study confirmed that the expression of S100A4 was significantly increased during the progressive EMT process of CRS mucosal epithelial cells, and revealed that the transcriptional regulation of S100A4 plays an important role in the occurrence and development of EMT. This finding will help us to better understand the pathogenesis behind the remodeling in CRS patients, and identify target molecules for the treatment of CRS., Competing Interests: Author HD was employed by the company Qilu Pharmaceutical Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gong, Shi, Bing, Li, Hu, Zhang, Yang, Guo, Du, Xia and Liu.)

Bipolar disorder (BD) and obesity are highly comorbid. We previously performed a genome-wide association study (GWAS) for BD risk accounting for the effect of body mass index (BMI), which identified a genome-wide significant single-nucleotide polymorphism (SNP) in the gene encoding the transcription factor 7 like 2 (TCF7L2). However, the molecular function of TCF7L2 in the central nervous system (CNS) and its possible role in the BD and BMI interaction remained unclear. In the present study, we demonstrated by studying human induced pluripotent stem cell (hiPSC)-derived astrocytes, cells that highly express TCF7L2 in the CNS, that the BD-BMI GWAS risk SNP is associated with glucocorticoid-dependent repression of the expression of a previously uncharacterized TCF7L2 transcript variant. That transcript is a long non-coding RNA (lncRNA-TCF7L2) that is highly expressed in the CNS but not in peripheral tissues such as the liver and pancreas that are involved in metabolism. In astrocytes, knockdown of the lncRNA-TCF7L2 resulted in decreased expression of the parent gene, TCF7L2, as well as alterations in the expression of a series of genes involved in insulin signaling and diabetes. We also studied the function of TCF7L2 in hiPSC-derived astrocytes by integrating RNA sequencing data after TCF7L2 knockdown with TCF7L2 chromatin-immunoprecipitation sequencing (ChIP-seq) data. Those studies showed that TCF7L2 directly regulated a series of BD risk genes. In summary, these results support the existence of a CNS-based mechanism underlying BD-BMI genetic risk, a mechanism based on a glucocorticoid-dependent expression quantitative trait locus that regulates the expression of a novel TCF7L2 non-coding transcript., (© 2021. The Author(s).)

The leucine-rich repeat-containing G-protein-coupled receptor 6 (LGR6) is considered to be a stem cell marker in many normal tissues and promotes tissue development, regeneration, and repair. LGR6 is also related to the initiation and progression of some malignant tumors. However, the role of LGR6 in cervical cancer has not been reported. Here, immunohistochemistry and western blotting showed that LGR6 was significantly upregulated in cervical cancer, compared with the normal cervix. By analyzing The Cancer Genome Atlas database, LGR6 was found to be correlated with a poor prognosis of cervical cancer. Then, a small population of LGR6 high cells isolated by using the fluorescence-activated cell sorting exhibited enhanced properties of cancer stem cells including self-renewal, differentiation, and tumorigenicity. Moreover, RNA sequencing revealed that LGR6 was correlated with the Wnt signaling pathway and TOP/FOP, reverse transcription-PCR, and western blotting further proved that LGR6 could activate the Wnt/β-catenin signaling pathway. Interestingly, LGR6 upregulated the expression of TCF7L2 by activating the Wnt/β-catenin pathway. Then, TCF7L2 combining with β-catenin in the nucleus enhanced LGR6 transcription by binding the promoter of LGR6, which further activated the Wnt signaling to form a positive feedback loop. Thus, our study demonstrated that LGR6 activated a novel β-catenin/TCF7L2/LGR6-positive feedback loop in LGR6 high cervical cancer stem cells (CSCs), which provided a new therapeutic strategy for targeting cervical CSCs to improve the prognosis of cervical cancer patients., (© 2021. The Author(s).)

Colorectal cancer (CRC) liver metastasis is a significant clinical problem for which better therapies are urgently needed. Tumor-associated macrophage, a major cell population in the tumor microenvironment, is a known contributor to primary cancer progression and cancer metastasis. Here, we found TAM recruitment and M2 polarization were increased in the hepatic metastatic lesion compared with the primary site of human CRC tissues. Moreover, Pearson correlation analysis showed that TAM recruitment and polarization were closely correlated with the elevated TCF4 expression in the metastatic site. To investigate the role of TCF4 in CRC liver metastasis, we generated a syngeneic mouse model using MC38 cells splenic injection. Results from in vivo experiments and mouse models revealed that TCF4 deficiency in MC38 cells does not affect their proliferation and invasion; however, it reduces TAM infiltration and M2 polarization in the metastasis site. Further studies indicated that these effects are mediated by the TCF4 regulated CCL2 and CCR2 expression. TCF4 or CCL2 silencing in the tumor cells prevent CRC liver metastasis in the mouse model. Altogether, these findings suggest that the TCF4-CCL2-CCR2 axis plays an essential role in CRC liver metastasis by enhancing TAMs recruitment and M2 polarization., (© 2021. The Author(s).)

Mouse embryonic stem cells (mESCs) can be maintained in vitro in defined N2B27 medium supplemented with two chemical inhibitors for GSK3 and MEK (2i) and the cytokine leukemia inhibitory factor (LIF), which act synergistically to promote self-renewal and pluripotency. Here, we find that genetic deletion of the four genes encoding the TCF/LEF transcription factors confers mESCs with the ability to self-renew in N2B27 medium alone. TCF/LEF quadruple knockout (qKO) mESCs display dysregulation of several genes, including Aire, Dnmt3l, and IcosL, located adjacent to each other within a topologically associated domain (TAD). Aire, Dnmt3l, and IcosL appear to be regulated by TCF/LEF in a β-catenin independent manner. Moreover, downregulation of Aire and Dnmt3l in wild-type mESCs mimics the loss of TCF/LEF and increases mESC survival in the absence of 2iL. Hence, this study identifies TCF/LEF effectors that mediate exit from the pluripotent state., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Transcriptional regulation by Wnt signalling is primarily thought to be accomplished by a complex of β-catenin and TCF family transcription factors (TFs). Although numerous studies have suggested that additional TFs play roles in regulating Wnt target genes, their mechanisms of action have not been investigated in detail. We characterised a Wnt-responsive element (WRE) downstream of the Wnt target gene Axin2 and found that TCFs and Caudal type homeobox (CDX) proteins were required for its activation. Using a new separation-of-function TCF mutant, we found that WRE activity requires the formation of a TCF/CDX complex. Our systematic mutagenesis of this enhancer identified other sequences essential for activation by Wnt signalling, including several copies of a novel CAG DNA motif. Computational and experimental evidence indicates that the TCF/CDX/CAG mode of regulation is prevalent in multiple WREs. Put together, our results demonstrate the complex nature of cis- and trans- interactions required for signal-dependent enhancer activity., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Thoracic aortic aneurysm (TAA) is characterized by dilation of the aortic root or ascending/descending aorta. TAA is a heritable disease that can be potentially life threatening. While 10%-20% of TAA cases are caused by rare, pathogenic variants in single genes, the origin of the majority of TAA cases remains unknown. A previous study implicated common variants in FBN1 with TAA disease risk. Here, we report a genome-wide scan of 1,351 TAA-affected individuals and 18,295 control individuals from the Cardiovascular Health Improvement Project and Michigan Genomics Initiative at the University of Michigan. We identified a genome-wide significant association with TAA for variants within the third intron of TCF7L2 following replication with meta-analysis of four additional independent cohorts. Common variants in this locus are the strongest known genetic risk factor for type 2 diabetes. Although evidence indicates the presence of different causal variants for TAA and type 2 diabetes at this locus, we observed an opposite direction of effect. The genetic association for TAA colocalizes with an aortic eQTL of TCF7L2, suggesting a functional relationship. These analyses predict an association of higher expression of TCF7L2 with TAA disease risk. In vitro, we show that upregulation of TCF7L2 is associated with BCL2 repression promoting vascular smooth muscle cell apoptosis, a key driver of TAA disease., Competing Interests: Declaration of interests The spouse of C.J.W. is an employee of Regeneron. All other authors declare no competing interests., (Copyright © 2021 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Aim: MicroRNAs play an important role in the maintenance of cellular functions by fine-tuning gene expression levels. The aim of the current study was to identify genetically caused changes in microRNA expression which associate with islet dysfunction in diabetic mice., Methods: To identify novel microRNAs involved in islet dysfunction, transcriptome and miRNome analyses were performed in islets of obese, diabetes-susceptible NZO and diabetes-resistant B6-ob/ob mice and results combined with quantitative trait loci (QTL) and functional in vitro analysis., Results: In islets of NZO and B6-ob/ob mice, 94 differentially expressed microRNAs were detected, of which 11 are located in diabetes QTL. Focusing on conserved microRNAs exhibiting the strongest expression difference and which have not been linked to islet function, miR-205-5p was selected for further analysis. According to transcriptome data and target prediction analyses, miR-205-5p affects genes involved in Wnt and calcium signalling as well as insulin secretion. Over-expression of miR-205-5p in the insulinoma cell line INS-1 increased insulin expression, left-shifted the glucose-dependence of insulin secretion and supressed the expression of the diabetes gene TCF7L2. The interaction between miR-205-5p and TCF7L2 was confirmed by luciferase reporter assay., Conclusion: MiR-205-5p was identified as relevant microRNA involved in islet dysfunction by interacting with TCF7L2., (© 2021 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.)

Morphine is an opioid agonist and a nonselective mu, kappa and delta receptor agonist. It is a commonly used analgesic drug for the treatment of acute and chronic pain as well as cancer pain. Morphine is particularly important to address the problem of morphine tolerance. Tcf7l2, known as a risk gene for schizophrenia and autism, encodes a member of the LEF1/TCF transcription factor family. TCF7L2 is an important transcription factor that is upregulated in neuropathic pain models. However, the relationship between TCF7L2 and morphine tolerance has not been reported. In this study, we found that morphine tolerance led to the upregulation of TCF7L2 in the spinal cord, and also led to the upregulation of TCF7L2 expression in glial cells, which promoted inflammation related signal, and activated TLR4 / NF-κB/NLRP3 pathway. In addition, TCF7L2 regulated microglial cell activation induced by chronic morphine treatment. Mechanically, we found that TCF7L2 transcriptionally regulated TLR4 expression, and the depletion of TCF7L2 alleviated morphine tolerance induced by chronic morphine treatment, and further alleviated pain hypersensitivity induced by chronic morphine treatment. We therefore suggested that TCF7L2 regulates the activation of TLR4/ NF-κB/NLRP3 pathway in microglia, and is involved in the formation of morphine tolerance. Our results provide a new idea for the regulation mechanism of morphine tolerance., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Super-enhancers or stretch enhancers (SEs) consist of large clusters of active transcription enhancers which promote the expression of critical genes that define cell identity during development and disease. However, the role of many super-enhancers in tumor cells remains unclear. This study aims to explore the function and mechanism of a new super-enhancer in various tumor cells. A new super-enhancer that exists in a variety of tumors named EphA2-Super-enhancer (EphA2-SE) was found using multiple databases and further identified. CRISPR/Cas9-mediated deletion of EphA2-SE results in the significant downregulation of its target gene EphA2. Mechanistically, we revealed that the core active region of EphA2-SE comprises E1 component enhancer, which recruits TCF7L2 and FOSL2 transcription factors to drive the expression of EphA2, induce cell proliferation and metastasis. Bioinformatics analysis of RNA-seq data and functional experiments in vitro illustrated that EphA2-SE deletion inhibited cell growth and metastasis by blocking PI3K/AKT and Wnt/β-catenin pathway in HeLa, HCT-116 and MCF-7 cells. Overexpression of EphA2 in EphA2-SE -/- clones rescued the effect of EphA2-SE deletion on proliferation and metastasis. Subsequent xenograft animal model revealed that EphA2-SE deletion suppressed tumor proliferation and survival in vivo. Taken together, these findings demonstrate that EphA2-SE plays an oncogenic role and promotes tumor progression in various tumors by recruiting FOSL2 and TCF7L2 to drive the expression of oncogene EphA2.

Promoting oligodendrocyte (OL) differentiation represents a promising option for remyelination therapy for treating the demyelinating disease multiple sclerosis (MS). The Wnt effector transcription factor 7-like 2 (TCF7l2) was upregulated in MS lesions and had been proposed to inhibit OL differentiation. Recent data suggest the opposite yet underlying mechanisms remain elusive. Here, we unravel a previously unappreciated function of TCF7l2 in controlling autocrine bone morphogenetic protein (BMP)4-mediated signaling. Disrupting TCF7l2 in mice of both sexes results in oligodendroglial-specific BMP4 upregulation and canonical BMP4 signaling activation in vivo Mechanistically, TCF7l2 binds to Bmp4 gene regulatory element and directly represses its transcriptional activity. Functionally, enforced TCF7l2 expression promotes OL differentiation by reducing autocrine BMP4 secretion and dampening BMP4 signaling. Importantly, compound genetic disruption demonstrates that oligodendroglial-specific BMP4 deletion rescues arrested OL differentiation elicited by TCF7l2 disruption in vivo Collectively, our study reveals a novel connection between TCF7l2 and BMP4 in oligodendroglial lineage and provides new insights into augmenting TCF7l2 for promoting remyelination in demyelinating disorders such as MS. SIGNIFICANCE STATEMENT Incomplete or failed myelin repairs, primarily resulting from the arrested differentiation of myelin-forming oligodendrocytes (OLs) from oligodendroglial progenitor cells, is one of the major reasons for neurologic progression in people affected by multiple sclerosis (MS). Using in vitro culture systems and in vivo animal models, this study unraveled a previously unrecognized autocrine regulation of bone morphogenetic protein (BMP)4-mediated signaling by the Wnt effector transcription factor 7-like 2 (TCF7l2). We showed for the first time that TCF7l2 promotes oligodendroglial differentiation by repressing BMP4-mediated activity, which is dysregulated in MS lesions. Our study suggests that elevating TCF7l2 expression may be possible in overcoming arrested oligodendroglial differentiation as observed in MS patients., (Copyright © 2021 the authors.)

Background: Breast cancer is the most prevalent malignancy occurring in females. In recent years, emerging evidence has suggested that circular RNAs are involved in the development of multiple cancers. Circ-PRMT5 has recently attracted attention as a tumor-promoting circular RNA. In the present study, we focused on exploring the biological effects of circ-PRMT5 in breast cancer., Methods: A quantitative real-time polymerase chain reaction was used to determine the expression of circ-PRMT5 in breast cancer. In vitro experiments, including cell-counting kit-8, 5-ethynyl-2'-deoxyuridine, flow cytometry and tube formation assays, were performed to test the effects of circ-PRMT5 on the cellular progression of breast cancer. Bioinformatic analysis, luciferase reporter, radioimmunoprecipitation and RNA-pull down assays were performed to predict the potential microRNAs interacting with circ-PRMT5 and mRNAs that can be targeted by miR-509-3p., Results: Circ-PRMT5 is up-regulated in breast cancer tissues and cells. Importantly, an elevation of circ-PRMT5 indicates a poor prognosis in patients with breast cancer. Functionally, knockdown of circ-PRMT5 suppresses cell proliferation and angiogenesis and increases cell apoptosis in breast cancer. Mechanistically, we identified that circ-PRMT5 up-regulates TCF7L2 expression by acting as a miR-509-3p sponge. The negative expression correlation between miR-509-3p and circ-PRMT5 or TCF7L2 in clinical tissues was further demonstrated. Rescue assays showed that TCF7L2 overexpression reverses the antitumoral effects of circ-PRMT5 knockdown on breast cancer cell processes. Additionally, we demonstrated that circ-PRMT5 activates the phosphoinositide 3-kinase (PI3K)/AKT pathway by up-regulation of TCF7L2., Conclusions: Overall, our data indicate that the circ-PRMT5/miR-509-3p/TCF7L2 axis can aggravate the malignant character of breast cancer cells by the regulation of the PI3K/AKT pathway., (© 2020 John Wiley & Sons, Ltd.)

In colon cancer, downregulation of the transmembrane heparan sulfate proteoglycan syndecan-1 (Sdc-1) is associated with increased invasiveness, metastasis, and dedifferentiation. As Sdc-1 modulates signaling pathways relevant to stem cell function, we tested the hypothesis that it may regulate a tumor-initiating cell phenotype. Sdc-1 small-interfering RNA knockdown in the human colon cancer cell lines Caco2 and HT-29 resulted in an increased side population (SP), enhanced aldehyde dehydrogenase 1 activity, and higher expression of CD133, LGR5, EPCAM, NANOG, SRY (sex-determining region Y)-box 2, KLF2, and TCF4/TCF7L2. Sdc-1 knockdown enhanced sphere formation, cell viability, Matrigel invasiveness, and epithelial-to-mesenchymal transition-related gene expression. Sdc-1-depleted HT-29 xenograft growth was increased compared to controls. Decreased Sdc-1 expression was associated with an increased activation of β1-integrins, focal adhesion kinase (FAK), and wingless-type (Wnt) signaling. Pharmacological FAK and Wnt inhibition blocked the enhanced stem cell phenotype and invasive growth. Sequential flow cytometric SP enrichment substantially enhanced the stem cell phenotype of Sdc-1-depleted cells, which showed increased resistance to doxorubicin chemotherapy and irradiation. In conclusion, Sdc-1 depletion cooperatively enhances activation of integrins and FAK, which then generates signals for increased invasiveness and cancer stem cell properties. Our findings may provide a novel concept to target a stemness-associated signaling axis as a therapeutic strategy to reduce metastatic spread and cancer recurrence. DATABASES: The GEO accession number of the Affymetrix transcriptomic screening is GSE58751., (© 2020 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Wnt/β-catenin signaling is important for development and progression of colorectal cancer (CRC). The degradation complex for β-catenin is functionally impaired in CRC cells, thereby resulting in the accumulation of β-catenin and its translocation into the nucleus. Nuclear β-catenin interacts with and co-activates T cell factor4 (TCF4), resulting in β-catenin/TCF4-dependent transcription. Therefore, nuclear β-catenin has been categorized as the main driving force in the tumorigenesis of CRC. Recent studies reveal that Jun activation domain-binding protein 1 (JAB1) enhances the degradation of seven in absentia homolog-1 (SIAH-1), a putative E3 ubiquitin ligase of β-catenin, and positively regulates the expression of total β-catenin in human CRC cells. An another recent study also shows that nuclear β-catenin is ubiquitinated and degraded by an E3 ubiquitin ligase, tripartite motif-containing protein 33 (TRIM33). However, the regulatory mechanism for the expression of nuclear β-catenin remains to be fully understood. In this study, we have demonstrated that JAB1 positively regulates the expression of nuclear β-catenin, c-MYC as a β-catenin/TCF4 target, and cell cycle regulators, such as Ki-67 and topoisomerase IIα, in human CRC cells. Taken together, these results suggest that JAB1 is considered as a promising target for novel CRC therapy., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2020. Published by Elsevier Inc.)

The Wnt/β-catenin pathway is one of the major pathways that regulates embryonic development, adult homeostasis, and stem cell self-renewal. In this pathway, transcription factors T-cell factor and lymphoid enhancer factor (TCF/LEF) serve as a key switch to repress or activate Wnt target gene transcription by recruiting repressor molecules or interacting with the β-catenin effector, respectively. It has become evident that the protein stability of the TCF/LEF family members may play a critical role in controlling the activity of the Wnt/β-catenin signaling pathway. However, factors that regulate the stability of TCF/LEFs remain largely unknown. Here, we report that pVHL binding protein 1 (VBP1) regulates the Wnt/β-catenin signaling pathway by controlling the stability of TCF/LEFs. Surprisingly, we found that either overexpression or knockdown of VBP1 decreased Wnt/β-catenin signaling activity in both cultured cells and zebrafish embryos. Mechanistically, VBP1 directly binds to all four TCF/LEF family members and von Hippel-Lindau tumor-suppressor protein (pVHL). Either overexpression or knockdown of VBP1 increases the association between TCF/LEFs and pVHL and then decreases the protein levels of TCF/LEFs via proteasomal degradation. Together, our results provide mechanistic insights into the roles of VBP1 in controlling TCF/LEFs protein stability and regulating Wnt/β-catenin signaling pathway activity., Competing Interests: Conflict of interest—The authors declared that they have no conflicts of interest with the contents of this article., (© 2020 Zhang et al.)

The canonical Wnt signaling pathway is mediated by interaction of β-catenin with the T-cell factor/lymphoid enhancer-binding factor (TCF/LEF) transcription factors and subsequent transcription activation of Wnt-target genes. In the hematopoietic system, the function of the pathway has been mainly investigated by rather unspecific genetic manipulations of β-catenin that yielded contradictory results. Here, we used a mouse expressing a truncated dominant negative form of the human TCF4 transcription factor (dnTCF4) that specifically abrogates β-catenin-TCF/LEF interaction. Disruption of the β-catenin-TCF/LEF interaction resulted in the accumulation of immature cells and reduced granulocytic differentiation. Mechanistically, dnTCF4 progenitors exhibited downregulation of the Csf3r gene, reduced granulocyte colony-stimulating factor (G-CSF) receptor levels, attenuation of downstream Stat3 phosphorylation after G-CSF treatment, and impaired G-CSF-mediated differentiation. Chromatin immunoprecipitation assays confirmed direct binding of TCF/LEF factors to the promoter and putative enhancer regions of CSF3R. Inhibition of β-catenin signaling compromised activation of the emergency granulopoiesis program, which requires maintenance and expansion of myeloid progenitors. Consequently, dnTCF4 mice were more susceptible to Candida albicans infection and more sensitive to 5-fluorouracil-induced granulocytic regeneration. Importantly, genetic and chemical inhibition of β-catenin-TCF/LEF signaling in human CD34+ cells reduced granulocytic differentiation, whereas its activation enhanced myelopoiesis. Altogether, our data indicate that the β-catenin-TCF/LEF complex directly regulates G-CSF receptor levels, and consequently controls proper differentiation of myeloid progenitors into granulocytes in steady-state and emergency granulopoiesis. Our results uncover a role for the β-catenin signaling pathway in fine tuning the granulocytic production, opening venues for clinical intervention that require enhanced or reduced production of neutrophils., (© 2020 by The American Society of Hematology.)

Lipoprotein lipase (LPL) plays a central role in hydrolyzing triglyceride and its deficiency leads to atherosclerosis. Artesunate (ART), a derivative of artemisinin, has been demonstrated that ART reduces the formation of atherosclerotic plaques. However, it remains unclear whether ART-alleviated atherosclerotic lesion is involved in regulating lipid metabolism. ApoE -/- mice were fed a high-fat diet to form atherosclerotic plaques and then injected with artesunate or not. Oil Red O, HE and Masson staining were performed to assess atherosclerotic plaques. Both Western blot and qRT-PCR were applied to detect protein expression. The Luciferase reporter gene and Chromatin immunoprecipitation assays were used to assess the interaction between proteins. Immunofluorescence assay was performed to show the localization of target proteins. In vitro, our data shown that ART increased LPL expression and inhibition of NRF2 blocked the binding of TCF7L2 to LPL promoter region in VSMCs. Downregulated Klf2 could decrease the nuclear enrichment of NRF2, TCF7L2 and LPL expression. In vivo, ART decreased atherosclerotic plaque formation and increased VSMC counts and LPL expression within atherosclerotic plaques. We observed the reduced tendency of serum lipids, and increased in serum LPL activity in mice. In support of vitro data, the markedly increased KLF2, TCF7L2 and LPL expression have been detected in aorta. Our study suggests that ART may be a novel therapeutic drug for inhibition of atherosclerotic plaque formation. The molecular mechanism may involve in upregulation of LPL expression via the KLF2/NRF2/TCF7L2 pathway in VSMCs., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Most signals detected by genome-wide association studies map to non-coding sequence and their tissue-specific effects influence transcriptional regulation. However, key tissues and cell-types required for functional inference are absent from large-scale resources. Here we explore the relationship between genetic variants influencing predisposition to type 2 diabetes (T2D) and related glycemic traits, and human pancreatic islet transcription using data from 420 donors. We find: (a) 7741 cis-eQTLs in islets with a replication rate across 44 GTEx tissues between 40% and 73%; (b) marked overlap between islet cis-eQTL signals and active regulatory sequences in islets, with reduced eQTL effect size observed in the stretch enhancers most strongly implicated in GWAS signal location; (c) enrichment of islet cis-eQTL signals with T2D risk variants identified in genome-wide association studies; and (d) colocalization between 47 islet cis-eQTLs and variants influencing T2D or glycemic traits, including DGKB and TCF7L2. Our findings illustrate the advantages of performing functional and regulatory studies in disease relevant tissues.

Predicting where transcription factors bind in the genome from their in vitro DNA-binding affinity is confounded by the large number of possible interactions with nearby transcription factors. To characterize the in vivo binding logic for the Wnt effector Tcf7l2, we developed a high-throughput screening platform in which thousands of synthesized DNA phrases are inserted into a specific genomic locus, followed by measurement of Tcf7l2 binding by DamID. Using this platform at two genomic loci in mouse embryonic stem cells, we show that while the binding of Tcf7l2 closely follows the in vitro motif-binding strength and is influenced by local chromatin accessibility, it is also strongly affected by the surrounding 99 bp of sequence. Through controlled sequence perturbation, we show that Oct4 and Klf4 motifs promote Tcf7l2 binding, particularly in the adjacent ∼50 bp and oscillating with a 10.8-bp phasing relative to these cofactor motifs, which matches the turn of a DNA helix., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Aberrant activation of Wnt signaling plays a critical role in the initiation and progression of colorectal cancer (CRC). Chlorquinaldol (CQD) is a topical antimicrobial agent used to treat skin infections. Little is known about the anticancer activity of CQD and its underlying mechanisms. In this study, CQD was demonstrated to inhibit Wnt/β-catenin signaling through targeting the downstream part of this pathway. The results showed that CQD could inhibit the acetylation of β-catenin and disrupt the interaction of β-catenin with T-cell factor 4 (TCF4), leading to reduced binding of β-catenin to the promoters of Wnt target genes and downregulation of the expression of these target genes. Moreover, treatment with CQD suppressed the proliferation, migration, invasion and stemness of CRC cells. In APC min/+ mice and CRC cell xenografts, administration of CQD suppressed tumor growth and the expression of Wnt target genes c-Myc and Leucine-rich G protein-coupled receptor-5 (LGR5). These results strongly suggest that CQD may be a promising therapeutic agent in the treatment of CRC., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Alternative splicing contributes to diversification of gene function, yet consequences of splicing on functions of specific gene products is poorly understood. The major transcription factor TCF7L2 undergoes alternative splicing but the biological significance of TCF7L2 isoforms has remained largely to be elucidated. Here, we find that the TCF7L2 E-isoforms maintain, whereas the M and S isoforms disrupt morphogenesis of 3D-epithelial cell-derived organoids via regulation of epithelial-mesenchymal transition (EMT). Remarkably, TCF7L2E2 antagonizes, whereas TCF7L2M2/S2 promotes EMT-like effects in epithelial cells induced by transforming growth factor beta (TGFβ) signaling. In addition, we find TGFβ signaling reduces the proportion of TCF7L2E to TCF7L2M/S protein in cells undergoing EMT. We also find that TCF7L2 operates via TGFβ-Smad3 signaling to regulate EMT. Collectively, our findings unveil novel isoform-specific functions for the major transcription factor TCF7L2 and provide novel links between TCF7L2 and TGFβ signaling in the control of EMT-like responses and epithelial tissue morphogenesis.

In this study, we investigated the mechanisms underlying the altered hepatic glucose metabolism and enhanced diabetes risk in individuals with the TCF7L2 rs290487 C allele. Analysis of 195 cirrhotic patients revealed a higher insulin resistance index and incidence of hepatogenous diabetes in patients with the rs290487 C/C genotype compared to those with the C/T or T/T genotype. The in vitro experiments using targeted mutant PLC-PRF-5 cell line showed that cells with the rs290487 C/C genotype (C/C cells) had higher glucose production, lower glucose uptake, and lower TCF7L2 mRNA and protein levels than those with the C/T genotype (C/T cells). Integrated multi-omics analysis of ChIP-seq, ATAC-seq, RNA-seq, and metabolomics data showed genome-wide alterations in the DNA binding affinity of TCF7L2 in the C/C cells, including gain (e.g., PFKP and PPARGC1A ) and loss (e.g., PGK1 and PGM1 ) of binding sites in several glucose metabolism-related genes. These allele-specific changes in transcriptional regulation lead to increased expression of gluconeogenesis-related genes ( PCK1 , G6PC and PPARGC1A ) and their downstream metabolites (oxaloacetate and β-D-fructose 2,6-bisphosphate). These findings demonstrate that the TCF7L2 rs290487 C allele enhances gluconeogenesis through allele-specific changes in transcription and chromatin binding.