Forkhead Box P4 Research Articles

Incomplete Thermal Ablation-Induced FOXP4-Mediated Promotion of Malignant Progression in Liver Cancer via NDST2.

The explosive progression of residual hepatocellular carcinoma (HCC) following incomplete thermal ablation is challenging, and the underlying mechanisms require further exploration. We investigated the mechanism by which Forkhead box P4 (FOXP4) promotes the malignant transformation of residual HCC cells through N-deacetylase and N-sulfotransferase 2 (NDST2) after incomplete thermal ablation. The clinical significance of FOXP4 and NDST2 in HCC was evaluated using big data analysis. FOXP4 expression was detected in clinical samples of HCC. The gene expression levels in an in vitro heat-stressed HCC cell model were determined using quantitative real-time PCR (RT-qPCR) and Western blotting. The effects of the genes on heat-stressed HCC cells were investigated using Cell Counting Kit-8 (CCK-8), scratch, Transwell migration, and invasion assays. Additionally, the regulatory relationship between FOXP4 and NDST2 was validated using the Cleavage Under Targets and Tagmentation (CUT&Tag) experiments and phenotypic assays. High FOXP4 expression was correlated with liver cancer occurrence and development. In the heat-stressed HCC cell model, downregulating FOXP4 inhibited cancer cell progression. Besides, there was a positive association between FOXP4 and NDST2 in liver cancer. Suppressing FOXP4 reduced NDST2 expression in the heat-stressed HCC cells. Furthermore, reducing NDST2 expression weakened the biological behavior of heat-stressed HCC cells. FOXP4 and NDST2 are crucial in the incomplete thermal ablation of residual cancer. FOXP4 might regulate the biological progression of residual HCC after incomplete thermal ablation through NDST2.

Nuclear KRT19 is a transcriptional corepressor promoting histone deacetylation and liver tumorigenesis.

Epigenetic reprogramming and escape from terminal differentiation are poorly understood enabling characteristics of liver cancer. Keratin 19 (KRT19), classically known to form the intermediate filament cytoskeleton, is a marker of stemness and worse prognosis in liver cancer. This study aimed to address the functional roles of KRT19 in liver tumorigenesis and to elucidate the underlying mechanisms. Using multiplexed genome editing of hepatocytes in vivo, we demonstrated that KRT19 promoted liver tumorigenesis in mice. Cell fractionation revealed a previously unrecognized nuclear fraction of KRT19. Tandem affinity purification identified histone deacetylase 1 and REST corepressor 1, components of the corepressor of RE-1 silencing transcription factor (CoREST) complex as KRT19-interacting proteins. KRT19 knockout markedly enhanced histone acetylation levels. Mechanistically, KRT19 promotes CoREST complex formation by enhancing histone deacetylase 1 and REST corepressor 1 interaction, thus increasing the deacetylase activity. ChIP-seq revealed hepatocyte-specific genes, such as hepatocyte nuclear factor 4 alpha ( HNF4A ), as direct targets of KRT19-CoREST. In addition, we identified forkhead box P4 as a direct activator of aberrant KRT19 expression in liver cancer. Furthermore, treatment of primary liver tumors and patient-derived xenografts in mice suggest that KRT19 expression has the potential to predict response to histone deacetylase 1 inhibitors especially in combination with lenvatinib. Our data show that nuclear KRT19 acts as a transcriptional corepressor through promoting the deacetylase activity of the CoREST complex, resulting in dedifferentiation of liver cancer. These findings reveal a previously unrecognized function of KRT19 in directly shaping the epigenetic landscape in cancer.

Participation of Long Noncoding RNA FOXP4-AS1 in the Development and Progression of Endometrioid Carcinoma with Epigenetically Silencing DUSP5.

Background: Long noncoding RNAs (lncRNAs), as emerging regulators of a wide variety of biological processes via diverse mechanisms, have been demonstrated to be of increasing importance in biology. Genome-wide association studies of tumor samples have identified several lncRNAs as either oncogenes or tumor suppressors in various types of cancers. In recent years, the importance of lncRNAs, especially in endometrioid cancer (EEC), has become increasingly well understood. The lncRNA Forkhead box P4 antisense RNA 1 (FOXP4-AS1) has been reported to fulfill roles in several types of cancers; however, the main biological function and associated underlying molecular mechanism of FOXP4-AS1 in EEC have yet to be fully elucidated. The present study therefore aimed to investigate how RNA FOXP4-AS1 may participate in the development and progression of endometrioid carcinoma tissues. Materials and Methods: In the present study, the expression level of FOXP4-AS1 was investigated in endometrioid carcinoma tissues and matching nearby normal endometrial tissues collected from patients receiving surgery at the hospital. A series of molecular biological assays were performed to investigate the effect of FOXP4-AS1 on cell proliferation, cell migration, and cell invasion. Results: An increased concentration of FOXP4-AS1 was identified in endometrioid carcinoma samples and cell lines compared with the corresponding controls, and this lncRNA was found to be positively correlated with advanced FIGO stages in patients with endometrial cancer. Furthermore, knocking down endogenous FOXP4-AS1 led to a significant reduction in the colony formation number and a significant inhibition of cell proliferation, cell migration, and cell invasion in endometrioid carcinoma cells. Moreover, dual-specificity phosphatase 5 (DUSP5), which is lowly expressed in endometrioid carcinoma tissues cells and negatively modulated by FOXP4-AS1, was identified as the downstream target molecule of FOXP4-AS1. Subsequently, the mechanistic experiments confirmed that, through binding to enhancer of zeste homolog 2 (EZH2; one of the catalytic subunits of polycomb repressive complex 2 [PRC2]), FOXP4-AS1 could epigenetically suppress the expression of DUSP5. Finally, the oncogenic function of the FOXP4-AS1/EZH2/DUSP5 axis in endometrioid carcinoma was confirmed via rescue assays. Conclusions: The findings of the present study have highlighted how FOXP4-AS1 fulfills an oncogenic role in endometrioid carcinoma, and targeting FOXP4-AS1 and its pathway may provide new biomarkers for patients with endometrioid carcinoma.

Forkhead Box P4 promotes the proliferation of cells in colorectal adenocarcinoma

Abstract Objectives Forkhead Box P4 (FOXP4) is a transcription factor that promotes tumor formation and progression. However, studies on its roles in colorectal adenocarcinoma (CRAC) and cell proliferation regulation are few to date. This work investigates the expression of FOXP4 in CRAC, explores the characteristic of FOXP4 in different clinicopathological features, and analyzes its regulation of cell proliferation. Methods The GEPIA database was used to predict the trend of FOXP4 expression in colon cancer and normal mucosa. Tumor tissue and normal paracancerous mucosal tissue were sampled from 64 cases diagnosed with CRAC and who were receiving radical surgery at Tianjin Hospital from January 2017 and December 2022. FOXP4 and proliferating cell nuclear antigen (PCNA) were detected by the immunohistochemistry EnVision method. The colon cancer cell lines SW480, HCT15, and SW620 and the normal colon cell line NCM460 were selected, and expression of FOXP4 was detected by the Western blot method. The siRNA-FOXP4 plasmid was synthesized and transfected with SW480 and HCT15 cell lines, respectively, to establish si-FOXP4 groups, and empty vector transfection group (NC-FOXP4) and blank control group (NC) was set up. The expression levels of FOXP4 and PCNA were detected by the Western blot method, while the cell proliferation activity was assessed using CCK-8. Normally distributed quantitative data were compared between two and more groups with ANOVA (SNK-based pairwise comparison), while intergroup enumeration data comparisons were performed through χ 2 test and assessed through linear correlation analysis. Results GEPIA-based prediction shows a potential rise in FOXP4 expression in colon cancer. The rate of positive FOXP4 expression is significantly higher in CRAC tissue than in normal mucosa (p<0.05). The difference in FOXP4 is statistically significant in the comparison of maximum tumor diameter and depth of invasion in CRAC (p<0.05) but not in the comparison of gender, age, degree of differentiation, tumor focus, tumor embolism, and lymph node metastasis (p>0.05). The expression levels of FOXP4 and PCNA in CRAC are positively correlated (p<0.05). FOXP4 expression is significantly higher in cell lines SW480, HCT15, and SW620 than in cell line NCM460. The cell proliferation activity and PCNA expression are significantly lower in si-FOXP4 group than in NC-FOXP4 and NC groups for cell lines SW480 and HCT15. Conclusions FOXP4 is highly expressed and has a proliferative effect on tumor cells in CRAC.

Hsa_circ_0017956 Acts as miR-758-3p Sponge to Facilitate the Progression of Non-small-Cell Lung Cancer by Regulating FOXP4 Expression.

Accumulating studies have demonstrated the important role of circular RNAs (circRNAs) in the progression of different human tumors, including non-small-cell lung cancer (NSCLC). The purpose of this study was to deeply study the function and mechanism of circ_0017956 in NSCLC. Real-time quantitative polymerase chain reaction (RT-qPCR) was applied to detect the expression of circ_0017956, microRNA-758-3p (miR-758-3p), and Forkhead Box P4 (FOXP4). Western blot was performed to determine the protein levels. Cell proliferation was examined by cell counting kit-8 (CCK-8) assay and 5-ethynyl-2'-deoxyuridine (EdU) assay. Flow cytometry was used to evaluate the apoptosis of NSCLC cells. Transwell assay was applied to detect cell migratory and invasive capacities. The angiogenesis ability was evaluated by tube formation experiment. The target relationship between miR-758-3p and circ_0017956 or FOXP4 was confirmed by dual-luciferase reporter assay. Animal experiment was conducted to assess the effect of circ_0017956 in vivo. Circ_0017956 and FOXP4 were upregulated, while miR-758-3p was downregulated in NSCLC tissues and cells. Silencing of circ_0017956 significantly suppressed cell proliferation, migration, invasion, and angiogenesis, but promoted cell apoptosis in NSCLC cells. Mechanically, circ_0017956 functioned as a sponge for miR-758-3p and miR-758-3p could directly interact with FOXP4. Moreover, silencing of miR-758-3p or overexpression of FOXP4 could overturn the anticancer influence of circ_0017956 interference on NSCLC cells. Besides that, circ_0017956 knockdown hindered tumor growth in vivo. Altogether, circ_0017956 promoted the progression of NSCLC by regulating FOXP4 through sponging miR-758-3p.

The essential role of forkhead box P4 (FOXP4) in thyroid cancer: a study related to The Cancer Genome Atlas and experimental data.

Thyroid cancer (THCA) is the most common endocrine cancer in the world. Although most patients with THCA have a good prognosis, the prognosis of those with THCA who have an extra-glandular invasion, vascular invasion, and distant metastasis is poor. Therefore, it is very important to find potential biomarkers that can effectively predict the prognosis and progression of highly aggressive THCAs. It has been identified that forkhead box P4 (FOXP4) may be a new biomarker for the proliferation and prognosis for tumor diagnosis. However, the expression and function of FOXP4 in THCA remain to be determined. In the present study, the function of FOXP4 in cells was investigated through the comprehensive analysis of data in The Cancer Genome Atlas and combined with experiments including immunohistochemistry (IHC), colony formation, Cell Counting Kit-8 assay, wound scratch healing, and transwell invasion assay. In the present study, relevant bioinformatic data showed that FOXP4 was highly expressed in THCA, which was consistent with the results of the IHC and cell experiments. Meanwhile, 10 FOXP4-related hub genes were identified as potential diagnostic genes for THCA. It was found in further experiments that FOXP4 was located in the nucleus of THCA cells, and the expression of FOXP4 in the nucleus was higher than that in the cytoplasm. FOXP4 knockdown inhibited in vitro proliferation of the THCA cells, whereas overexpression promoted the proliferation and migration of THCA cells. Furthermore, deficiency of FOXP4 induced cell-cycle arrest. FOXP4 might be a potential target for diagnosing and treating THCA.

The clinical prognostic value of lncRNA FOXP4-AS1 in cancer patients: A meta-analysis and bioinformatics analysis based on TCGA datasets.

Background:The mortality and recurrence of patients with cancer is of high prevalence. Long non-coding RNA (lncRNA) forkhead box P4 antisense RNA 1 (FOXP4-AS1) is a promising lncRNA. There is increasing evidence that lncRNA FOXP4-AS1 is abnormally expressed in various tumors and is associated with cancer prognosis. This study was designed to identify the prognostic value of lncRNA FOXP4-AS1 in human malignancies.Methods:We searched electronic databases up to April 29, 2022, including PubMed, Cochrane Library, Embase, MEDLINE, and Web of Science. Eligible studies that evaluated the clinicopathological and prognostic role of lncRNA FOXP4-AS1 in patients with malignant tumors were included. The pooled odds ratios (ORs) and the hazard ratios (HRs) were calculated to assess the role of lncRNA FOXP4-AS1 using Stata/SE 16.1 software.Results:A total of 6 studies on cancer patients were included in the present meta-analysis. The combined results revealed that high expression of lncRNA FOXP4-AS1 was significantly associated with unfavorable overall survival (OS) (HR = 1.99, 95% confidence interval [CI]: 1.65–2.39, P < .00001), and poor disease-free survival (DFS) (HR = 1.81, 95% CI: 1.54–2.13, P < .00001) in a variety of cancers. In additional, the increase in lncRNA FOXP4-AS1 expression was also correlated with tumor size ((larger vs smaller) (OR = 3.16, 95% CI: 2.12–4.71, P < .00001), alpha-fetoprotein (≥400 vs <400) (OR = 3.81, 95%CI: 2.38–6.11, P = .83), lymph node metastasis (positive vs negative) (OR = 2.93, 95%CI: 1.51–5.68, P = .001), and age (younger vs older) (OR = 2.06, 95% CI: 1.41–3.00, P = .00002) in patients with cancer. Furthermore, analysis results using The Cancer Genome Atlas (TCGA) dataset showed that the expression level of lncRNA FOXP4-AS1 was higher in most tumor tissues than in the corresponding normal tissues, which predicted a worse prognosis.Conclusions:In this meta-analysis, we demonstrate that high lncRNA FOXP4-AS1 expression may become a potential marker to predict cancer prognosis.

FOXP4 promotes proliferation of human spermatogonial stem cells.

Continuous self-renewal and differentiation of spermatogonial stem cells (SSCs) is vital for maintenance of adult spermatogenesis. Although several spermatogonial stem cell regulators have been extensively investigated in rodents, regulatory mechanisms of human SSC self-renewal and differentiation have not been fully established. We analyzed single-cell sequencing data from the human testis and found that forkhead box P4 (FOXP4) expression gradually increased with development of SSCs. Further analysis of its expression patterns in human testicular tissues revealed that FOXP4 specifically marks a subset of spermatogonia with stem cell potential. Conditional inactivation of FOXP4 in human SSC lines suppressed SSC proliferation and significantly activated apoptosis. FOXP4 expressions were markedly suppressed in tissues with dysregulated spermatogenesis. These findings imply that FOXP4 is involved in human SSC proliferation, which will help elucidate on the mechanisms controlling the fate decisions in human SSCs.

Foxp1 and Foxp4 Deletion Causes the Loss of Follicle Stem Cell Niche and Cyclic Hair Shedding by Inducing Inner Bulge Cell Apoptosis.

Quiescent hair follicle stem cells (HFSCs) reside in specialized bulge niche where they undergo activation and differentiation upon sensing niche-dependent signals during hair follicle (HF) homeostasis and wound repair. The underlying mechanism of HFSCs and bulge niche maintenance is poorly understood. Our previous study has reported that a transcription factor, forkhead box P1 (Foxp1), functions to maintain the quiescence of HFSCs. Here, we further discovered that forkhead box P4 (Foxp4), a close family member of Foxp1, had similar expression profiles in various components of HFs and formed a complex with Foxp1 in vitro and in vivo. The HF-specific deficiency of Foxp4 resulted in the precocious activation of HFSCs during hair cycles. In contrast to single Foxp1 or Foxp4 conditional knockout (cKO) mice, Foxp1/4 double cKO exerted an additive effect in the spectrum and severity of phenotypes in HFSC activation, hair cycling acceleration and hair loss, coupled with remarkable downregulation of fibroblast growth factor 18 (Fgf18) and bone morphogenetic protein 6 (Bmp6) expression in bulge cells. In addition, the double KO of Foxp1/4 induced the apoptosis of K6-positive (K6+) inner bulge cells, a well-established stem cell (SC) niche, thus resulting in the destruction of the bulge SC niche and recurrent hair loss. Our investigation reveals the synergistic role of Foxp1/4 in sustaining K6+ niche cells for the quiescence of HFSCs.

FOXP4-AS1 Inhibits Papillary Thyroid Carcinoma Proliferation and Migration Through the AKT Signaling Pathway.

Papillary thyroid carcinoma, also known as PTC, is one of the commonest malignancies in the endocrine system. Long non-coding RNAs (lncRNAs) in PTC could maintain proliferative signaling, induce therapeutic resistance, activate invasion and migration, and sustain stem cell-like characteristics. In this paper, results showed that lncRNA forkhead box P4 antisense RNA 1 (FOXP4-AS1) is downregulated in PTC tissues and cell lines. Patients in TCGA cohort with a higher FOXP4-AS1 expression showed a higher disease-free interval (DFI) rate, and the expression of FOXP4-AS1 is shown to be linked to the clinical stage, T stage, N stage, and extraglandular invasion condition of the TC patients. FOXP4-AS1 is localized in the cell cytoplasmic domain of PTC cells. Functionally, upregulated FOXP4-AS1 inhibited PTC cell proliferation, apoptosis, and migration, whereas it downregulated FOXP4-AS1-promoted progression of PTC. In vivo assay also confirmed the tumor inhibitory effect of FOXP4-AS1 in PTC growth. Mechanism analysis indicated that FOXP4-AS1 can play its functions by regulating the AKT signaling pathway, and AKT inhibitor treatment could attenuate the impact of FOXP4-AS1 on PTC progression. Furthermore, FOXP4-AS1 also negatively regulates the expression of its host gene FOXP4. Collectively, we showed that FOXP4-AS1 inhibited PTC progression although AKT signaling and FOXP4-AS1 plays a tumor-suppressor role in PTC tumorigenesis.

Downregulation of microRNA‑423‑5p suppresses TGF‑β1‑induced EMT by targeting FOXP4 in airway fibrosis.

Airway fibrosis (AF) is a common disease that can severely affect patient prognosis. Epithelial-mesenchymal transition (EMT) participates in the pathophysiological development of AF and several studies have demonstrated that some microRNAs (miRNAs) contribute to the development of EMT. The aim of this study was to investigate the function of miR-423-5p in the EMT process and its possible underlying mechanism in BEAS-2B cells. The present study utilized the BEAS-2B cell line to model EMT in AF. Online tools, fluorescence in situ hybridization analysis and an RNA pull-down assay were used to identify potential target genes of miR-423-5p. In addition, immunohistochemistry, wound healing assays, Transwell migration assays, flow cytometry, enzyme-linked immunosorbent assay, reverse transcription-quantitative PCR, western blot analysis and immunofluorescence staining were used to determine the function of miR-423-5p and its target gene in the EMT process in AF. The results indicated that the miR-423-5p expression in AF tissues and BEAS-2B cells stimulated with 10 ng/ml TGF-β1 for 24 h was significantly increased compared with that in the control group. Overexpression of miR-423-5p facilitated TGF-β1-induced EMT in BEAS-2B cells; by contrast, downregulation of miR-423-5p suppressed TGF-β1-induced EMT in BEAS-2B cells. Furthermore, forkhead box p4 (FOXP4) was identified as a potential target gene of miR-423-5p and changes in the miR-423-5p and FOXP4 expression were shown to significantly affect the expression of PI3K/AKT/mTOR pathway members. In summary, overexpression of miR-423-5P promoted the EMT process in AF by downregulating FOXP4 expression and the underlying mechanism may partly involve activation of the PI3K/AKT/mTOR pathway.

FOXP4 differentially controls cold-induced beige adipocyte differentiation and thermogenesis.

Beige adipocytes have a discrete developmental origin and possess notable plasticity in their thermogenic capacity in response to various environmental cues, but the transcriptional machinery controlling beige adipocyte development and thermogenesis remains largely unknown. By analyzing beige adipocyte-specific knockout mice, we identified a transcription factor, forkhead box P4 (FOXP4), that differentially governs beige adipocyte differentiation and activation. Depletion of Foxp4 in progenitor cells impaired beige cell early differentiation. However, we observed that ablation of Foxp4 in differentiated adipocytes profoundly potentiated their thermogenesis capacity upon cold exposure. Of note, the outcome of Foxp4 deficiency on UCP1-mediated thermogenesis was confined to beige adipocytes, rather than to brown adipocytes. Taken together, we suggest that FOXP4 primes beige adipocyte early differentiation, but attenuates their activation by potent transcriptional repression of the thermogenic program.

Long non-coding RNA FOXP4-AS1 facilitates the biological functions of hepatocellular carcinoma cells via downregulating ZC3H12D by mediating H3K27me3 through recruitment of EZH2

BackgroundSome studies have reported the effect of long non-coding RNA forkhead box P4 antisense RNA 1 (lncRNA FOXP4-AS1) on hepatocellular carcinoma (HCC). Here, we aimed to discuss the effects of FOXP4-AS1/enhancer of zeste homolog 2 (EZH2)/trimethylation of lysine 27 on histone H3 (H3K27me3)/zinc finger CCCH-type containing 12D (ZC3H12D) axis on HCC.MethodsThe expression of FOXP4-AS1, EZH2, and ZC3H12D, and abundance of H3K27me3 in HCC tissues and cells were tested. The relationship between FOXP4-AS1 expression and prognosis of HCC patients was analyzed. The biological functions of HCC cells were detected via loss- and gain-of-function assays. The tumor weight and volume in vivo were tested. The interaction between FOXP4-AS1 and EZH2 as well as that between EZH2 and H3K27me3 was verified.ResultsFOXP4-AS1 and EZH2 expression and H3K27me3 abundance were enhanced while ZC3H12D expression was depressed in HCC tissues and cells. Knockdown of FOXP4-AS1 suppressed biological functions of HCC cells as well as the weight and volume of HCC transplanted tumor. Depleting ZC3H12D reversed the effect of downregulated FOXP4-AS1 on HCC cells. FOXP4-AS1 suppressed ZC3H12D expression via mediating H3K27me3 by recruitment of EZH2.ConclusionThe key findings of the present study demonstrate that FOXP4-AS1 suppresses ZC3H12D expression via mediating H3K27me3 by recruitment of EZH2, thus promoting the progression of HCC.Graphical abstract

LncRNA RP11-116G8.5 promotes the progression of lung squamous cell carcinoma through sponging miR-3150b-3p/miR-6870-5p to upregulate PHF12/FOXP4

BackgroundLung squamous cell carcinoma (LUSC) is one of the commonest malignancies worldwide. Long noncoding RNAs (lncRNAs) have been revealed to engage in cancer development. LncRNA RP11-116G8.5 is a new founded lncRNA that has not been clearly elucidated in LUSC. Materials and methodsExpression levels of RNAs in LUSC cells were measured through qRT-PCR. To identify the functions of RP11-116G8.5, CCK-8 assay, colony formation assay and EdU assay were conducted in indicated LUSC cells. Mechanism experiments, including RNA pull down assay, Ago2-RIP assay and luciferase reporter assay were performed to demonstrate the interaction between RP11-116G8.5 and miR-3150b-3p/miR-6870-5p. Meanwhile, the interaction between miR-3150b-3p/miR-6870-5p and their downstream targets PHD finger protein 12 (PHF12), and forkhead box P4 (FOXP4) were also proven in the same methods. ResultsRP11–116G8.5 was expressed at high level in LUSC cell lines. Down-regulated RP11–116G8.5 repressed cell proliferation, migration and invasion, but accelerated apoptosis. Furthermore, it was proven that RP11-116G8.5 could act as sponges for miR-3150b-3p and miR-6870-5p these miRNAs were found to act as cancer suppressors in LUSC cells. PHF12 and FOXP4 were verified as the target gene of miR-3150b-3p and miR-6870-5p separately. Overexpression of PHF12 and FOXP4 could reverse the repressive effect of RP11-116G8.5 knockdown on LUSC progression. Additionally, Paired Box 5 (PAX-5) was proven to be the transcription factor for RP11–116G8.5 in LUSC cells. ConclusionsLncRNA RP11-116G8.5 promotes malignant behaviors of LUSC through sponging miR-3150b-3p/miR-6870-5p to upregulate PHF12/FOXP4 expression. Availability of dataThe research data is confidential.

FOXP4-AS1 is a favorable prognostic-related enhancer RNA in ovarian cancer.

Ovarian cancer (OV) is the main cause of deaths worldwide in female reproductive system malignancies. Enhancer RNAs (eRNAs) are derived from the transcription of enhancers and has attracted increasing attention in cancers recently. However, the biological functions and clinical significance of eRNAs in OV have not been well described presently. We used an integrated data analysis to identify prognostic-related eRNAs in OV. Tissue-specific enhancer-derived RNAs and their regulating genes were considered as putative eRNA–target pairs using the computational pipeline PreSTIGE. Gene expression profiles and clinical data of OV and 32 other cancer types were obtained from the UCSC Xena platform. Altogether, 71 eRNAs candidates showed significant correlation with overall survival (OS) of OV samples (Kaplan–Meier log-rank test, P<0.05). Among which, 23 were determined to be correlated with their potential target genes (Spearman’s r > 0.3, P<0.001). It was found that among the 23 prognostic-related eRNAs, the expression of forkhead box P4 antisense RNA 1 (FOXP4-AS1) had the highest positive correlation with its predicted target gene FOXP4 (Spearman’s r = 0.61). Moreover, the results were further validated by RT-qPCR analysis in an independent OV cohort. Our results suggested the eRNA FOXP4-AS1 expression index may be a favorable independent prognostic biomarker candidate in OV.

MiR-491-5p inhibits the proliferation and migration of A549 cells by FOXP4.

Aberrant expression of microRNAs (miRNAs/miRs) plays a key role in the development of non-small cell lung cancer (NSCLC). In the present study, lower miRNA (miR)-491-5p levels and a higher forkhead box P4 (FOXP4) mRNA level were observed in NSCLC tissues and cell lines, compared to adjacent tissues and the normal human lung epithelial cell line BEAS-2B, respectively. A549 cell proliferation and migration were inhibited upon transfection of miR-491-5p mimics compared to miR-negative control (NC) mimics. In addition, compared to miR-NC mimics, overexpression of miR-491-5p decreased FOXP4 expression, while downregulation of miR-491-5p increased FOXP4 expression in A549 cells. The dual luciferase assay confirmed that the 3'untranslated region of FOXP4 was a target for miR-491-5p in A549 cells. Moreover, compared with the control short hairpin (sh)RNA, there was lower expression levels of TGF-β and its downstream targets (MMP-2 and MMP-9) in the FOXP4 shRNA group. Similarly, compared to miR-NC mimics, overexpression of miR-491-5p decreased MMP-2 and MMP-9 expression levels. In FOXP4-knockdown A549 cells, overexpression of miR-491-5p showed little effect on cell proliferation/migration. In A549 cells, overexpression of FOXP4 partially reversed the miR-491-5p mimics-induced inhibition on the cell proliferation and migration. These results may provide new insights into the role of miR-491-5p in NSCLC.

Circ‑0000212 promotes cell proliferation of colorectal cancer by sponging miR‑491 and modulating FOXP4 expression

Colorectal cancer (CRC) is a lethal and common malignancy worldwide. Non-coding (nc)RNAs have been shown to modulate tumor progression in several types of cancer. The present study aimed to investigate the role of hsa_circ_0000212 in CRC, as a sponge of microRNA (miR)-491. The expression levels of miR-491 and forkhead box P4 (FOXP4) were analyzed using data from The Cancer Genome Atlas. The association between miR-491 and FOXP4 and the clinicopathological characteristics were also analyzed. A novel circular (circ)RNA, hsa_circ_0000212, was found to sponge miR-491 based on bioinformatics analysis. The potential binding site between miR-491 and FOXP4 or circ-0000212 was validated using luciferase and RNA immunoprecipitation assays. The expression levels and distribution of circ-0000212 was also determined. Cell Counting Kit-8 and colony formation assays were performed to determine the role of miR-491 or circ-0000212 on the proliferation of the CRC cells. Decreased miR-491 or increased FOXP4 expression levels were associated with the pathological stage in patients with CRC. In addition, miR-491 inhibited cell proliferation by targeting FOXP4. circ-0000212 was increased in CRC tissues and was predominantly localized in the cytoplasm. Furthermore, circ-0000212 augmented viability of the CRC cells by sponging miR-491 and modulating FOXP4. In conclusion, circ-0000212 may serve as a novel tumor-promoter and drug target in CRC.

The forkhead box transcription factor FoxP4 regulates thermogenic programs in adipocytes

Forkhead box transcription factors have been shown to be involved in various developmental and differentiation processes. In particular, members of the FoxP family have been previously characterized in depth for their participation in the regulation of lung and neuronal cell differentiation and T-cell development and function; however, their role in adipocyte functionality has not yet been investigated. Here, we report for the first time that Forkhead box P4 (FoxP4) is expressed at high levels in subcutaneous fat depots and mature thermogenic adipocytes. Through molecular and gene expression analyses, we revealed that FoxP4 is induced in response to thermogenic stimuli, both in vivo and in isolated cells, and is regulated directly by the heat shock factor protein 1 through a heat shock response element identified in the proximal promoter region of FoxP4. Further detailed analysis involving chromatin immunoprecipitation and luciferase assays demonstrated that FoxP4 directly controls the levels of uncoupling protein 1, a key regulator of thermogenesis that uncouples fatty acid oxidation from ATP production. In addition, through our gain-of-function and loss-of-function studies, we showed that FoxP4 regulates the expression of a number of classic brown and beige fat genes and affects oxygen consumption in isolated adipocytes. Overall, our data demonstrate for the first time the novel role of FoxP4 in the regulation of thermogenic adipocyte functionality.

Exosomal miR‐3180‐3p inhibits proliferation and metastasis of non‐small cell lung cancer by downregulating FOXP4

BackgroundNon‐small cell lung cancer (NSCLC) is one of the most malignant cancers worldwide and its pathogenesis is not completely clear. In this study, we explored the functions and mechanisms of exosomes transferring miR‐3180‐3p in NSCLC progression.MethodsThe expression levels of miR‐3180‐3p in NSCLC tissues and paracarcinoma tissues was obtained from the GEO database (GEO: GSE53882). Exosomes derived from A549 cells were identified. Proliferation, migration and invasion were measured after treatment with exosomal miR‐3180‐3p or transfection using miR‐3180‐3p mimics. The relationship between miR‐3180‐3p and forkhead box P4 (FOXP4) was predicted using a bioinformatic tool and measured using a dual‐luciferase reporter gene assay and western blotting. Finally, a mouse xenograft model of NSCLC cells was established to verify the function of exosomal miR‐3180‐3p in vivo.ResultsWe found that miR‐3180‐3p decreased in both NSCLC cell lines and patient tissues. Overexpression of miR‐3180‐3p or treatment with exosomal miR‐3180‐3p significantly suppressed cell proliferation and metastasis in NSCLC cell lines. Subsequently, we found miR‐3180‐3p downregulated FOXP4 protein expression levels. Furthermore, the volumes and weights of nude mouse tumors expressing exosomal miR‐3180‐3p were significantly reduced.ConclusionsExosomal miR‐3180‐3p suppresses NSCLC progression by downregulating FOXP4 expression.Key pointsSignificant findings of the studyWe found that exosomal miR‐3180‐3p suppressed NSCLC progression and also identified a miR‐3180‐3p target gene. These findings provide a foundation to determine innovative therapeutic strategies.What this study addsThis study contributes to research investigating exosomal containing miRNAs.

LncRNA FOXP4‑AS1 predicts poor prognosis and accelerates the progression of mantle cell lymphoma through the miR‑423‑5p/NACC1 pathway

Long non-coding RNA (lncRNA) forkhead box P4 antisense RNA 1 (FOXP4-AS1) has been determined to function as an oncogene in various types of cancer. However, the biological function and the underlying mechanisms of FOXP4-AS1 in mantle cell lymphoma (MCL) remain to be uncovered. The expression and the associated clinicopathological characteristics and prognostic significance of FOXP4-AS1 were explored in MCL clinical samples. The effects of FOXP4-AS1 on MCL cellular behaviors, including proliferation, migration and invasion were analyzed using CCK-8, crystal violet and Transwell assays. The downstream molecules of FOXP4-AS1 were explored using bioinformatics analysis and dual luciferase assay. Our results showed that FOXP4-AS1 expression was upregulated in MCL patients, and that the high expression of FOXP4-AS1 was correlated with the unfavorable prognosis of patients. Functionally, while FOXP4-AS1 downregulation inhibited proliferation, migration and invasion of MCL cells, FOXP4-AS1 overexpression had promotive effects on these cellular processes. Mechanistically, FOXP4-AS1 was found to act as a competing endogenous (ce)RNA for miR-423-5p to regulate the expression of nucleus accumbens-associated 1 (NACC1). The negative regulation of FOXP4-AS1 on miR-423-5p compared to that of miR-423-5p on NACC1 was determined at the mRNA or protein levels in MCL cells. Moreover, an inverse expression correlation between FOXP4-AS1 and miR-423-5p, and that between miR-423-5p and NACC1 was confirmed in MCL clinical samples. In addition, rescue assay showed that miR-423-5p upregulation or NACC1 knockdown abolished the promoting effects of FOXP4-AS1 on MCL cell proliferation, migration and invasion. In conclusion, FOXP4-AS1 promotes MCL progression through the upregulation of NACC1 expression by inhibiting miR-423-5p. FOXP4-AS1 may serve as a novel therapeutic target for patients with MCL.