TCF1-LEF1 co-expression identifies a multipotent progenitor cell (TH2-MPP) across human allergic diseases.

Repetitive exposure to antigen in chronic infection and cancer drives T cell exhaustion, limiting adaptive immunity. In contrast, aberrant, sustained T cell responses can persist over decades in human allergic disease. To understand these divergent outcomes, we employed bioinformatic, immunophenotyping and functional approaches with human diseased tissues, identifying an abundant population of type 2 helper T (TH2) cells with co-expression of TCF7 and LEF1, and features of chronic activation. These cells, which we termed TH2-multipotent progenitors (TH2-MPP) could self-renew and differentiate into cytokine-producing effector cells, regulatory T (Treg) cells and follicular helper T (TFH) cells. Single-cell T-cell-receptor lineage tracing confirmed lineage relationships between TH2-MPP, TH2 effectors, Treg cells and TFH cells. TH2-MPP persisted despite in vivo IL-4 receptor blockade, while thymic stromal lymphopoietin (TSLP) drove selective expansion of progenitor cells and rendered them insensitive to glucocorticoid-induced apoptosis in vitro. Together, our data identify TH2-MPP as an aberrant T cell population with the potential to sustain type 2 inflammation and support the paradigm that chronic T cell responses can be coordinated over time by progenitor cells.

Targeting LEF1-mediated epithelial-mesenchymal transition reverses lenvatinib resistance in hepatocellular carcinoma.

Acquired resistance is a significant hindrance to clinical application of lenvatinib in unresectable hepatocellular carcinoma (HCC). Further in-depth investigation of resistance mechanisms can help to develop additional therapeutic strategies to overcome or delay resistance. In our study, two lenvatinib-resistant (LR) HCC cell lines were established by treatment with gradient increasing concentration of lenvatinib, named Hep3B-LR and HepG2-LR. Interestingly, continuous lenvatinib treatment reinforced epithelial-mesenchymal transition (EMT), cell migration, and cell invasion. Gene set enrichment analysis (GSEA) enrichment analysis of RNA-sequencing from Hep3B-LR and corresponding parental cells revealed that activation of Wnt signaling pathway was involved in this adaptive process. Active ß-catenin and its downstream target lymphoid enhancer binding factor 1 (LEF1) were significantly elevated in LR HCC cells, which promoted lenvatinib resistance through mediating EMT-related genes. Data analysis based on Gene Expression Omnibus (GEO) and the Cancer Genome Atlas Program (TCGA) databases suggests that LEF1, as a key regulator of EMT, was a novel molecular target linked to lenvatinib resistance and poor prognosis in HCC. Using a small-molecule specific inhibitor ICG001 and knocking down LEF1 showed that targeting LEF1 restored the sensitivity of LR HCC cells to lenvatinib. Our results uncover upregulation of LEF1 confers lenvatinib resistance by facilitating EMT, cell migration, and invasion of LR HCC cells, indicating that LEF1 is a novel therapeutic target for overcoming acquired lenvatinib resistance.

Downregulation of TCF7 and LEF1 is a key determinant of tumor-infiltrating regulatory T-cell function.

Forkhead box P3 (Foxp3)-expressing regulatory T (Treg) cells play essential roles in immune homeostasis but also contribute to establish a favorable environment for tumor growth by suppressing anti-tumor immune responses. It is thus necessary to specifically target tumor-infiltrating Treg cells to minimize effects on immune homeostasis in cancer immunotherapy. However, molecular features that distinguish tumor-infiltrating Treg cells from those in secondary lymphoid organs remain unknown. Here we characterize distinct features of tumor-infiltrating Treg cells by global analyses of the transcriptome and chromatin landscape. They exhibited activated phenotypes with enhanced Foxp3-dependent transcriptional regulation, yet being distinct from activated Treg cells in secondary lymphoid organs. Such differences may be attributed to the extensive clonal expansion of tumor-infiltrating Treg cells. Moreover, we found that TCF7 and LEF1 were specifically downregulated in tumor-infiltrating Treg cells both in mice and humans. These factors and Foxp3 co-occupied Treg suppressive function-related gene loci in secondary lymphoid organ Treg cells, whereas the absence of TCF7 and LEF1 accompanied altered gene expression and chromatin status at these gene loci in tumor-infiltrating Treg cells. Functionally, overexpression of TCF7 and LEF1 in Treg cells inhibited the enhancement of Treg suppressive function upon activation. Our results thus show the downregulation of TCF7 and LEF1 as markers of highly suppressive Treg cells in tumors and suggest that their absence controls the augmentation of Treg suppressive function in tumors. These molecules may be potential targets for novel cancer immunotherapy with minimum effects on immune homeostasis.

A distinct isoform of lymphoid enhancer binding factor 1 (LEF1) epigenetically restricts EBV reactivation to maintain viral latency.

As a human tumor virus, EBV is present as a latent infection in its associated malignancies where genetic and epigenetic changes have been shown to impede cellular differentiation and viral reactivation. We reported previously that levels of the Wnt signaling effector, lymphoid enhancer binding factor 1 (LEF1) increased following EBV epithelial infection and an epigenetic reprogramming event was maintained even after loss of the viral genome. Elevated LEF1 levels are also observed in nasopharyngeal carcinoma and Burkitt lymphoma. To determine the role played by LEF1 in the EBV life cycle, we used in silico analysis of EBV type 1 and 2 genomes to identify over 20 Wnt-response elements, which suggests that LEF1 may bind directly to the EBV genome and regulate the viral life cycle. Using CUT&RUN-seq, LEF1 was shown to bind the latent EBV genome at various sites encoding viral lytic products that included the immediate early transactivator BZLF1 and viral primase BSLF1 genes. The LEF1 gene encodes various long and short protein isoforms. siRNA depletion of specific LEF1 isoforms revealed that the alternative-promoter derived isoform with an N-terminal truncation (ΔN LEF1) transcriptionally repressed lytic genes associated with LEF1 binding. In addition, forced expression of the ΔN LEF1 isoform antagonized EBV reactivation. As LEF1 repression requires histone deacetylase activity through either recruitment of or direct intrinsic histone deacetylase activity, siRNA depletion of LEF1 resulted in increased histone 3 lysine 9 and lysine 27 acetylation at LEF1 binding sites and across the EBV genome. Taken together, these results indicate a novel role for LEF1 in maintaining EBV latency and restriction viral reactivation via repressive chromatin remodeling of critical lytic cycle factors.

LEF1 isoforms regulate cellular senescence and aging.

The study of aging and its mechanisms, such as cellular senescence, has provided valuable insights into age-related pathologies, thus contributing to their prevention and treatment. The current abundance of high-throughput data combined with the surge of robust analysis algorithms has facilitated novel ways of identifying underlying pathways that may drive these pathologies. For the purpose of identifying key regulators of lung aging, we performed comparative analyses of transcriptional profiles of aged versus young human subjects and mice, focusing on the common age-related changes in the transcriptional regulation in lung macrophages, T cells, and B immune cells. Importantly, we validated our findings in cell culture assays and human lung samples. Our analysis identified lymphoid enhancer binding factor 1 (LEF1) as an important age-associated regulator of gene expression in all three cell types across different tissues and species. Follow-up experiments showed that the differential expression of long and short LEF1 isoforms is a key regulatory mechanism of cellular senescence. Further examination of lung tissue from patients with idiopathic pulmonary fibrosis, an age-related disease with strong ties to cellular senescence, revealed a stark dysregulation of LEF1. Collectively, our results suggest that LEF1 is a key factor of aging, and its differential regulation is associated with human and murine cellular senescence.

Mechanism of LEF1-AS1 regulating HUVEC cells by targeting miR-489-3p/S100A11 axis.

Background: The venous malformation is the most common congenital vascular malformation and exhibits the characteristics of local invasion and lifelong progressive development. Long noncoding RNA (lncRNA) regulates endothelial cells, vascular smooth muscle cells, macrophages, vascular inflammation, and metabolism and also affects the development of venous malformations. This study aimed to elucidate the role of the lncRNA LEF1-AS1 in the development of venous malformations and examine the interaction among LEF1-AS1, miR-489-3p, and S100A11 in HUVEC cells. Methods: Venous malformation tissues, corresponding normal venous tissues, and HUVEC cells were used. Agilent human lncRNA microarray gene chip was used to screen differential genes, RNA expression was detected using quantitative reverse transcription PCR, and protein expression was detected using Western blotting. The proliferation, migration, and angiogenesis of HUVEC cells were assessed using CCK8, transwell, and in vitro angiogenesis tests. Results: A total of 1,651 lncRNAs were screened using gene chip analysis, of which 1015 were upregulated and 636 were downregulated. The lncRNA LEF1-AS1 was upregulated with an obvious difference multiple, and the fold-change value was 11.03273. The results of the analysis performed using the StarBase bioinformatics prediction website showed that LEF1-AS1 and miR-489-3p possessed complementary binding sites and that miR-489-3p and S100A11 also had complementary binding sites. The findings of tissue experiments revealed that the expressions of LEF1-AS1 and S100A11 were higher in tissues with venous malformations than in normal tissues, whereas the expression of miR-489-3p was lower in venous malformations than in normal tissues. Cell culture experiments indicated that LEF1-AS1 promoted the proliferation, migration, and angiogenesis of HUVEC cells. In these cells, LEF1-AS1 targeted miR-489-3p, which in turn targeted S100A11. LEF1-AS1 acted as a competitive endogenous RNA and promoted the expression of S100A11 by competitively binding to miR-489-3p and enhancing the proliferation, migration, and angiogenesis of HUVEC cells. Thus, LEF1-AS1 participated in the occurrence and development of venous malformation. Conclusions: The expression of LEF1-AS1 was upregulated in venous malformations, and the expression of S100A11 was increased by the adsorption of miR-489-3p to venous endothelial cells, thus enhancing the proliferation, migration, and angiogenesis of HUVEC cells. In conclusion, LEF1-AS1 is involved in the occurrence and development of venous malformations by regulating the miR-489-3p/S100A11 axis, which provides valuable insights into the pathogenesis of this disease and opens new avenues for its treatment.

Constant small-cell changes and variable LEF1 expression in DUSP22-rearranged primary cutaneous anaplastic large-cell lymphoma: Analysis of the repeated biopsies of three patients.

DUSP22-rearranged primary cutaneous anaplastic large-cell lymphoma (pcALCL) has a biphasic histological pattern defined by large dermal atypical lymphocytes and epidermotropic small lymphocytes resembling pagetoid reticulosis, but the positivity rate of the biphasic pattern in DUSP22-rearranged pcALCL is unknown. Immunohistochemically, LEF1 expression in >75% of tumor cells is associated with DUSP22-rearrangement (DUSP22-R) in systemic ALCL. However, whether this association applies to pcALCL remains unclear. To analyze these pathological clues for screening DUSP22-R, we reviewed 11 skin biopsies from three patients with DUSP22-rearranged pcALCL. All specimens showed a biphasic pattern, of which three showed nonpagetoid infiltration of the epidermis. In all lesions, small-cell changes of tumor cells were observed not only within the epidermis but also under the epidermis. LEF1 positivity rates varied by lesion (range: 30%-90%, mean: 59.6%) with only three patients expressing LEF1 in more than 75% of tumor cells. In conclusion, the biphasic pattern was a constant finding in DUSP22-rearranged pcALCL, but it was not always pagetoid reticulosis-like. The recognition of small-cell change outside the epidermis may be helpful in diagnosing DUSP22-rearranged pcALCL. However, LEF1 expression was variable and its diagnostic usefulness may be limited.

Carcinogenesis promotion in oral squamous cell carcinoma: KDM4A complex-mediated gene transcriptional suppression by LEF1.

Oral squamous cell carcinoma (OSCC) is the most prevalent cancer of the mouth, characterised by rapid progression and poor prognosis. Hence, an urgent need exists for the development of predictive targets for early diagnosis, prognosis determination, and clinical therapy. Dysregulation of lymphoid enhancer-binding factor 1 (LEF1), an important transcription factor involved in the Wnt-ß-catenin pathway, contributes to the poor prognosis of OSCC. Herein, we aimed to explore the correlation between LEF1 and histone lysine demethylase 4 A (KDM4A). Results show that the KDM4A complex is recruited by LEF1 and specifically binds the LATS2 promoter region, thereby inhibiting its expression, and consequently promoting cell proliferation and impeding apoptosis in OSCC. We also established NOD/SCID mouse xenograft models using CAL-27 cells to conduct an in vivo analysis of the roles of LEF1 and KDM4A in tumour growth, and our findings show that cells stably suppressing LEF1 or KDM4A have markedly decreased tumour-initiating capacity. Overall, the results of this study demonstrate that LEF1 plays a pivotal role in OSCC development and has potential to serve as a target for early diagnosis and treatment of OSCC.

PD1 is transcriptionally regulated by LEF1 in mature T cells.

The role of programmed cell death 1 (PD1) in cancer immune evasion is of considerable importance, prompting the development of monoclonal antibodies that specifically target PD-1 to enhance the immune system for cancer therapy. Nevertheless, the efficacy of PD1/programmed cell death-Ligand 1 (PD-L1) blocking antibodies is limited to certain patients or tumor types. Although researchers have demonstrated the influence of PD-1 on the positive selection of T cells, its effect on the T-cell repertoire remains uncertain. Lymphoid enhancer binding factor 1 (LEF1) has been known to play a critical role as a transcription factor in the development and maturation of T cells. Despite the greater focus on the study of its homologous protein, T cell factor 1 (TCF1), we discovered that LEF1 had a positive regulatory effect on the transcription of PD1 in mature T cells, including CD4+ T cells, CD8+ T cells, and Treg cells. This finding was observed in LEF1 knockout and LEF1-stimulated mice models. Additionally, we confirmed the direct regulation of PD1 by LEF1 in tumor-infiltrating lymphocytes through tumor-implantation experiments. The direct regulation of PD1 by LEF1 was further validated in the LEF1 knockout cell line. The results of our study provide novel perspectives on the regulation of PD1 in immune responses and investigate potential approaches for clinical anti-PD1 therapy.

OTUD7B Activates Wnt Signaling Pathway through the Interaction with LEF1.

The Wnt signaling pathway plays a critical role in regulating normal cellular processes, including proliferation, differentiation, and apoptosis. Dysregulation of Wnt signaling has been implicated in various human diseases, including cancer. ß-catenin and LEF1 are key mediators of Wnt signaling, and their dysregulation is a hallmark of many cancer types. In this study, we aimed to identify the deubiquitinases (DUBs) that regulate the Wnt signaling pathway through the essential component LEF1. Screening candidate DUBs from the human DUB library, we discovered that OTUD7B interacts with LEF1 and activates Wnt signaling. OTUD7B and LEF1 interact with each other through the UBA and HMG domains, respectively. Furthermore, OTUD7B promotes the nuclear localization of LEF1, leading to an increased interaction with ß-catenin in the nucleus while not noticeably affecting ubiquitination on LEF1. Using qPCR array analysis, we found that OTUD7B overexpression leads to an upregulation of 75% of the tested Wnt target genes compared to the control. These findings suggest that OTUD7B may serve as a potential therapeutic target in human diseases, including cancers where Wnt signaling is frequently dysregulated.

The ß-catenin-LINC00183-miR-371b-5p-Smad2/LEF1 axis promotes adult T-cell lymphoblastic lymphoma progression and chemoresistance.

BACKGROUND: High-intensity chemotherapy regimens are often used in adult T-cell lymphoblastic lymphoma (T-LBL) patients. Nevertheless, the response rate remains unsatisfactory due to emergence of chemoresistance. Growing evidence has shown that long non-coding RNAs (lncRNAs) are involved in tumor progression and chemoresistance. Herein, we investigated the potential role of lncRNAs in T-LBLs. METHODS: RNAseq was used to screen and identify candidate lncRNAs associated with T-LBL progression and chemoresistance. Luciferase reporter assay was used to examine the binding of miR-371b-5p to the 3'UTR of Smad2 and LEF1, and the binding of TCF-4/LEF1 to the promoter of LINC00183. Chromatin immunoprecipitation assay was undertaken to analyze the connection between LEF1 and the LINC00183 promoter region. RNA immunoprecipitation assays were used to explore the mechanism whereby LINC00183 regulated miR-371b-5p. MTT and flow cytometry assays were used to measure apoptosis of T-LBL cells. RESULTS: LINC00183 was upregulated in T-LBL progression and chemoresistant tissues in both the Sun Yat-sen University Cancer Center dataset and the First Affiliated Hospital of Anhui Medical University dataset. High expression of LINC00183 was correlated with poorer overall survival and progression-free survival of T-LBL patients compared to those with low expression of LINC00183. Furthermore, miR-371b-5p was negatively regulated by LINC00183. In vivo and in vitro assays showed that LINC00183-mediated T-LBL chemoresistance depended on miR-371b-5p expression. The direct binding of miR-371b-5p to Smad2 and LEF1 was verified by luciferase assays. It was shown that TCF4/LEF1 could bind to the LINC00183 promoter site and increase its transcript level. Downregulation of miR-371b-5p led to increased expression of Smad2/LEF1, and in turn increased LINC00183 expression. Additionally, phospho-Smad2 promotes nuclear translocation of ß-catenin, LINC00183 downregulation decreased chemoresistance induced by ß-catenin and TGF-ß1 in T-LBL cells. CONCLUSION: We unraveled a ß-catenin-LINC00183-miR-371b-5p-Smad2/LEF1 feedback loop that promotes T-LBL progression and chemoresistance, indicating that LINC00183 may serve as a potential therapeutic target in T-LBLs.

Altered levels of lymphocyte enhancer-binding factor-1 modulates the pigmentation in acral and non-acral lesions of non-segmental vitiligo patients: a follow-up-based study in North India.

BACKGROUND: Lymphocyte enhancer-binding factor-1 (LEF1) is responsible for melanocyte proliferation, migration and differentiation and its downregulation may result in depigmentation in vitiligo. Narrowband UVB (NB-UVB) phototherapy is known to enhance melanocyte migration from hair follicles to lesional epidermis; hence, it may have a role in the upregulation of LEF1. OBJECTIVES: We intended to assess the expression of LEF1 both before and after NB-UVB therapy and correlate it with the extent of re-pigmentation. MATERIALS AND METHODS: In this prospective cohort study, 30 patients of unstable non-segmental vitiligo were administered NB-UVB phototherapy for 24 weeks. Skin biopsies were obtained from acral and non-acral sites in all patients, both prior to initiation and after completion of phototherapy and LEF1 expression was measured. RESULTS: Amongst the 16 patients who completed the study, at 24 weeks, all patients achieved > 50% re-pigmentation. However, > 75% re-pigmentation was achieved in only 11.1% of acral patches, whereas it was achieved in a significantly higher number of non-acral patches (66.6%) (p = 0.05). A significant increase was observed in the mean fluorescent intensity of the LEF1 gene in both acral as well as non-acral areas at 24 weeks as compared to baseline (p = 0.0078), However, no difference was observed between acral and non-acral lesions in the LEF1 expression at 24 weeks or the change in LEF1 expression from baseline. CONCLUSION: LEF1 expression modulates the re-pigmentation of vitiligo lesions after treatment with NBUVB phototherapy.

Cancer-testis non-coding RNA LEF1-AS1 regulates the nuclear translocation of PDCD5 and suppresses its interaction with p53 signaling: a novel target for immunotherapy in esophageal squamous cell carcinoma.

Despite the improvement of current classical treatment, the prognosis of esophageal squamous cell carcinoma (ESCC) remains poor. Immunotherapy, as a new treatment method, has revolutionized the therapy of various cancer types and created more attractive for ESCC. Cancer-testis genes (CTGs), because of its characteristic expression and immunomodulation property, are considered as the ideal targets for tumor immunotherapy. However, the ESCC-specific CTGs, especially long non-coding RNA (lncRNA), has not been elucidated. In the present study, a systematic strategy was adopted to screen ESCC-specific cancer-testis lncRNA (CT-lncRNA). Collectively, 447 genes were recognized as ESCC-specific CT-lncRNAs, in particularly LEF1-AS1 showed the most aberrantly expression and clinically associated with poor outcome. Functional assays revealed that H3K27 acetylation in LEF1-AS1 promoter might give rise to the activation of LEF1-AS1 during ESCC tumorigenesis. The activated LEF1-AS1 was predominantly localized in the cytoplasm implicated in regulation of apoptosis and proliferation capacities of ESCC cells in vitro and in vivo. Further mechanistic studies unveiled that LEF1-AS1 participated in ESCC by interacting with RNA binding protein PDCD5 through weakened its nuclear translocation binding to TP53, leading to p53 degradation and disruption the transcription of downstream genes. Taken together, our findings suggest that LEF1-AS1 acts as a CT-lncRNA and might be an ideal immunotherapeutic target for clinical intervention for ESCC.

Lymphoid enhancer binding factor 1 is associated with nose color in Yunong black pigs.

Yunong black pig is an indigenous black pig breed being cultivated that has a pure black whole body. However, some individuals appear with a white spot on the nose. We performed case-control association studies and FST approaches in 76 animals with nose color records (26 white-nosed pigs vs. 50 black-nosed pigs) by Illumina Porcine SNP50 BeadChip data. In total, 76 SNPs, which included 2 genome-wide significant SNPs and 18 chromosome-wide suggestive SNPs, were identified by association study. The top-ranked 0.1% windows of FST results as signals under selection and 24 windows were selected. The lymphoid enhancer binding factor 1 was identified as candidate gene with strong signal in analyses of genome-wide association study and FST in black- and white-nosed pigs. Overall, our findings provide evidence that nose color is a heritable trait influenced by many loci. The results contribute to expand our understanding of pigmentation in pigs and provide SNP markers for skin color and related traits selection in Yunong black pigs. Additional research on the genetic link between nose pigmentation is needed.

Transcriptional activity mediated by ß-CATENIN and TCF/LEF family members is completely dispensable for survival and propagation of multiple human colorectal cancer cell lines.

Unrestrained transcriptional activity of ß-CATENIN and its binding partner TCF7L2 frequently underlies colorectal tumor initiation and is considered an obligatory oncogenic driver throughout intestinal carcinogenesis. Yet, the TCF7L2 gene carries inactivating mutations in about 10% of colorectal tumors and is non-essential in colorectal cancer (CRC) cell lines. To determine whether CRC cells acquire TCF7L2-independence through cancer-specific compensation by other T-cell factor (TCF)/lymphoid enhancer-binding factor (LEF) family members, or rather lose addiction to ß-CATENIN/TCF7L2-driven gene expression altogether, we generated multiple CRC cell lines entirely negative for TCF/LEF or ß-CATENIN expression. Survival of these cells and the ability to propagate them demonstrate their complete ß-CATENIN- and TCF/LEF-independence. Nonetheless, one ß-CATENIN-deficient cell line eventually became senescent, and absence of TCF/LEF proteins and ß-CATENIN consistently impaired CRC cell proliferation, reminiscent of mitogenic effects of WNT/ß-CATENIN signaling in the healthy intestine. Despite this common phenotype, ß-CATENIN-deficient cells exhibited highly cell-line-specific gene expression changes with little overlap between ß-CATENIN- and TCF7L2-dependent transcriptomes. Apparently, ß-CATENIN and TCF7L2 independently control sizeable fractions of their target genes. The observed divergence of ß-CATENIN and TCF7L2 transcriptional programs, and the finding that neither ß-CATENIN nor TCF/LEF activity is strictly required for CRC cell survival has important implications when evaluating these factors as potential drug targets.

MiR-34a-5p promotes autophagy and apoptosis of ovarian granulosa cells via the Hippo-YAP signaling pathway by targeting LEF1 in chicken.

Follicular atresia is a natural physiological phenomenon in poultry reproduction. It is well known that follicular atresia is caused by both autophagy and apoptosis of granulosa cells. In current experiment, we evaluated the function of miR-34a-5p on autophagy and apoptosis in chicken follicular atresia. First, the follicular atresia model of chicken was successfully constructed by subcutaneous injection of tamoxifen (TMX), and found the expression of miR-34a-5p in the atresia follicles obviously increased. Then, we confirmed that miR-34a-5p accelerates autophagy and apoptosis of chicken granulose cells in vitro, and miR-34a-5p could induce apoptosis by mediating autophagy. Mechanistically, lymphoid enhancer binding factor 1 (LEF1) was deemed as a target gene for miR-34a-5p. On the contrary, LEF1 overexpression attenuated the autophagy and apoptosis of chicken granular cells. In addition, it was confirmed that the miR-34a-5p/LEF1 axis plays a regulatory role in chicken granulosa cells by mediating the Hippo-YAP signaling pathway. Taken together, this study demonstrated that miR-34a-5p contributes to autophagy and apoptosis of chicken follicular granulosa cells by targeting LEF1 to mediate the Hippo-YAP signaling pathway.

Expression dynamics of lymphoid enhancer-binding factor 1 in terminal Schwann cells, dermal papilla, and interfollicular epidermis.

BACKGROUND: Transcription factor lymphoid enhancer-binding factor 1 (LEF1) is a downstream mediator of the Wnt/ß-catenin signaling pathway. It is expressed in dermal papilla and surrounding cells in the hair follicle, promoting cell proliferation, and differentiation. RESULTS: Here, we report that LEF1 is also expressed all through the hair cycle in the terminal Schwann cells (TSCs), a component of the lanceolate complex located at the isthmus. The timing of LEF1 appearance at the isthmus coincides with that of hair follicle innervation. LEF1 is not found at the isthmus in the aberrant hair follicles in nude mice. Instead, LEF1 in TSCs is found in the de novo hair follicles reconstituted on nude mice by stem cells chamber graft assay. Cutaneous denervation experiment demonstrates that the LEF1 expression in TSCs is independent of nerve endings. At last, LEF1 expression in the interfollicular epidermis during the early stage of skin development is significantly suppressed in transgenic mice with T-cell factor 3 (TCF3) overexpression. CONCLUSION: We reveal the expression dynamics of LEF1 in skin during development and hair cycle. LEF1 expression in TSCs indicates that the LEF1/Wnt signal might help to establish a niche at the isthmus region for the lanceolate complex, the bulge stem cells and other neighboring cells.

LEF1 Drives a Central Memory Program and Supports Antitumor Activity of Natural Killer T Cells.

Vα24-invariant natural killer T cells (NKT) possess innate antitumor properties that can be exploited for cancer immunotherapy. We have shown previously that the CD62L+ central memory-like subset of these cells drives the in vivo antitumor activity of NKTs, but molecular mediators of NKT central memory differentiation remain unknown. Here, we demonstrate that relative to CD62L- cells, CD62L+ NKTs express a higher level of the gene encoding the Wnt/ß-catenin transcription factor lymphoid enhancer binding factor 1 (LEF1) and maintain active Wnt/ß-catenin signaling. CRISPR/Cas9-mediated LEF1 knockout reduced CD62L+ frequency after antigenic stimulation, whereas Wnt/ß-catenin activator Wnt3a ligand increased CD62L+ frequency. LEF1 overexpression promoted NKT expansion and limited exhaustion following serial tumor challenge and was sufficient to induce a central memory-like transcriptional program in NKTs. In mice, NKTs expressing a GD2-specific chimeric-antigen receptor (CAR) with LEF1 demonstrated superior control of neuroblastoma xenograft tumors compared with control CAR-NKTs. These results identify LEF1 as a transcriptional activator of the NKT central memory program and advance development of NKT cell-based immunotherapy. See related Spotlight by Van Kaer, p. 144.

Lateral confined growth of cells activates Lef1 dependent pathways to regulate cell-state transitions.

Long-term sustained mechano-chemical signals in tissue microenvironment regulate cell-state transitions. In recent work, we showed that laterally confined growth of fibroblasts induce dedifferentiation programs. However, the molecular mechanisms underlying such mechanically induced cell-state transitions are poorly understood. In this paper, we identify Lef1 as a critical somatic transcription factor for the mechanical regulation of de-differentiation pathways. Network optimization methods applied to time-lapse RNA-seq data identify Lef1 dependent signaling as potential regulators of such cell-state transitions. We show that Lef1 knockdown results in the down-regulation of fibroblast de-differentiation and that Lef1 directly interacts with the promoter regions of downstream reprogramming factors. We also evaluate the potential upstream activation pathways of Lef1, including the Smad4, Atf2, NFkB and Beta-catenin pathways, thereby identifying that Smad4 and Atf2 may be critical for Lef1 activation. Collectively, we describe an important mechanotransduction pathway, including Lef1, which upon activation, through progressive lateral cell confinement, results in fibroblast de-differentiation.

MicroRNA-621 functions as a metastasis suppressor in colorectal cancer by directly targeting LEF1 and suppressing Wnt/ß-catenin signaling.

AIMS: Colorectal liver metastasis (CRLM) is the leading death-causing among colorectal cancer (CRC) patients. Recently, a novel tumor-related microRNA, miR-621, has been identified as a tumor suppressor in diverse tumor types, but its role in CRLM remains unclear and requires further investigation. MAIN METHODS: To elucidate novel regulators of CRLM progression, we used a well-established CRLM animal model. After serially transplanting human colon carcinoma cell lines Caco-2 into the liver, we obtained liver metastatic variants that exhibited a strong ability for invasion and metastasis. High-throughput sequencing was conducted on these newly established cell lines. After comparison and prediction between the two cell lines: parental Caco-2 (hereafter referred to as F0) and F3, miR-621 was identified as a candidate regulator for lymphoid enhancer-binding factor 1 (LEF1) expression. Further validation was achieved with dual-luciferase reporter assay. KEY FINDINGS: The gain- and loss-of-function validation showed that miR-621 inhibits cell viability, cell cycle progression, colony formation, and proliferation in vitro. Meanwhile, miR-621 could reverse EMT malignant phenotype. LEF1, an important downstream mediator of activated Wnt/ß-catenin signaling pathway, was validated as the direct functional target of miR-621. miR-621 interacts directly with the LEF1 3'-UTR and post-transcriptionally suppresses LEF1 expression. Moreover, LEF1 overexpression reversed the effect of miR-621. LEF1 silencing counteracted miR-621 down-regulation-induced effects. Further in vivo experiments revealed that miR-621 over-expression suppressed CRLM, but LEF1 abrogated the inhibitory effect of miR-621. SIGNIFICANCE: MiR-621 is a vital tumor suppressor in CRC and could be a promising anti-cancer therapeutic target.