A hsa_circ_001726 axis regulated by E2F6 contributes to metastasis of hepatocellular carcinoma.

BACKGROUND: CircRNAs participate in the development of hepatocellular carcinoma (HCC). This work aims to explore the key tumor promoting circRNA as a gene therapy target. METHODS: The differentially expressed gene circRNAs in HCC tumor tissues was identified by mining GSE121714 dataset. EdU staining, wound healing, transwell invasion assay, TUNEL staining and western blotting examined proliferation, migration, invasion, apoptosis and epithelial mesenchymal transition (EMT). Xenograft mouse model and orthotopic transplantation tumor mouse model were constructed to verify the role of hsa_circ_001726 in growth and metastasis of HCC. The relationship among CCT2, E2F6, hsa_circ_001726, miR-671-5p and PRMT9 was identified by RNA-fluorescence in situ hybridization, luciferase reporter assay and RNA Immunoprecipitation. RESULTS: Eleven differentially expressed circRNAs were found in HCC tumors. Among them, hsa_circ_001726 was highly expressed in HCC tumors and cells, which was transcribed from CCT2. As a transcription factor of CCT2, E2F6 knockdown inactivated CCT2 promoter and reduced hsa_circ_001726 expression. Moreover, hsa_circ_001726 elevated PRMT9 expression by sponging miR-671-5p, and then activated Notch signaling pathway. Additionally, hsa_circ_001726 deficiency repressed malignant phenotypes of HCC cells, including proliferation, migration, invasion, EMT and apoptosis. In vivo, hsa_circ_001726 deficiency reduced tumor growth and lung metastasis of HCC in xenograft mouse models and orthotopic transplantation tumor mouse models. CONCLUSION: Hsa_circ_001726 functioned as an oncogene in HCC, which was derived from CCT2 and regulated by E2F6. Hsa_circ_001726 elevated PRMT9 expression by sponging miR-671-5p, and then activated Notch signaling pathway, thereby accelerating malignant phenotypes of HCC. Therefore, targeting hsa_circ_001726 may be a new avenue for HCC treatment.

Predictive role of E2F6 in cancer prognosis and responses of immunotherapy.

BACKGROUND: E2F6 is a member of the E2F transcription factor family. Numerous studies have demonstrated that E2F6 is critical to cancer development and progression, but its role in cancer immunotherapy remains unclear. METHODS: Genotype-Tissue Expression (GTEx) and The Cancer Genome Atlas (TCGA) databases were used to obtain RNA-seq data for cancer and normal tissues, and we utilized the cBioPortal to analyze E2F6 genomic alterations in pan-cancer. The protein localization of E2F6 was obtained using the Human Protein Atlas (HPA), and the upregulation of E2F6 expression in clinical glioblastoma multiforme (GBM) tissues was detected by Western blot analysis. The ComPPI website was used to analyze the protein interaction information of E2F6. To evaluate the role of E2F6 in pan-cancer prognosis, we used univariate Cox regression and Kaplan-Meier methods, and gene set enrichment analysis (GSEA) was utilized to identify markers associated with E2F6 expression in tumors. TIMER 2.0 was used to study E2F6-related immune cell infiltration in tumor tissues, and the correlation of E2F6 with immunotherapy biomarkers was investigated using Spearman correlation analysis. The role of E2F6 in the cell cycle was analyzed by flow cytometry, and the Cell Counting Kit-8 (CCK-8) and colony formation assays were utilized to determine the proliferative ability of cells. RESULTS: In most tumor types, E2F6 was highly expressed and was a good predictor of prognosis. E2F6 was significantly related to markers of immune activation, tumor immune cell infiltration, and immune regulators. Furthermore, E2F6 knockdown significantly attenuated the proliferative ability of glioma cells. Finally, E2F6 effectively predicted anti-programmed cell death 1 (PD1) treatment response. CONCLUSION: E2F6 is an effective biomarker that predicts the prognosis of cancer patients treated with anti-immune checkpoint therapy.

lncRNA ENST00000585827 Contributes to the Progression of Endometrial Carcinoma via Regulating miR-424/E2F6/E2F7 Axis.

Endometrial cancer (EC) ranks fourth among the most common gynecologic malignancies. Despite advances in medical technology, the pathogenesis is still unclear. Numerous reports have identified the involvement of lncRNA in the malignant progression of endometrial cancer. The aim of the study was to investigate the expression level of lncRNA ENST00000585827 (lncRNA E27) in endometrial cancer and the molecular mechanism that regulates the development of endometrial cancer. Combined with the results of the previous study, PCR analysis confirmed that lncRNA E27 was significantly upregulated in endometrial cancer cell lines. The results of CCK-8, wound healing assay, and transwell experiments showed that lncRNA E27 could significantly inhibit cell proliferation, migration, and invasion. Flow cytometry results confirmed that lncRNA E27 could promote apoptosis. Furthermore, based on bioinformatics predictions, dual-luciferase assay and RT-qPCR analysis confirmed that miR-424, as its downstream molecule, competitively regulates the expression of E2F6/E2F7. Rescue experiments further supported that lncRNA E27 inhibited proliferation, migration, invasion, and promoted apoptosis of endometrial cancer through miR-424/E2F6/E2F7 signaling axis. Conclusively, our findings revealed the role of lncRNA E27 in regulating the miR-424/E2F6/E2F7 signaling axis during EC progression, opening up new strategies for the treatment of endometrial cancer.

Circ_0004676 exacerbates triple-negative breast cancer progression through regulation of the miR-377-3p/E2F6/PNO1 axis.

BACKGROUND: The significant roles of circular RNAs (circRNAs) in different cancers and diseases have been reported. We now focused on the possible role of a newly recognized circRNA, circ_0004674 in triple-negative breast cancer (TNBC), and the related downstream mechanism. METHODS: The expression of circ_0004674 in TNBC tissues and cells was determined followed by analysis of the correlation between circ_0004674 and TNBC patients' prognosis. The interaction between circ_0004674, miR-377-3p, E2F6, and PNO1 was then identified using bioinformatics analysis combined with FISH, RIP, RNA pull-down, RT-qPCR, and Western blot analysis. Using gain-of-function and loss-of-function methods, we analyzed the effect of circ_0004674, miR-377-3p, E2F6, and PNO1 on TNBC in vivo and in vitro. RESULTS: Increased circ_0004674 and E2F6 but decreased miR-377-3p were observed in TNBC tissues and MDA-MB-231 TNBC cells, all of which findings were associated with poor prognosis in patients with TNBC. Silencing of circ_0004676 remarkably suppressed the proliferation, cell cycle progression, and migration of TNBC cells in vitro, as well as inhibiting tumorigenesis and metastasis in vivo. Additionally, circ_0004676 served as a sponge of miR-377-3p which bound to the transcription factor E2F6. In the presence of overexpression of circ_0004676, E2F6 expression and its target PNO1 expression were elevated, while miR-377-3p expression was decreased. Interestingly, overexpression of E2F6 could reverse the inhibitory effect on tumor growth caused by downregulation of circ_0004676. CONCLUSION: Our study highlighted the carcinogenic effect of circ_0004676 on TNBC through regulation of the miR-377-3p/E2F6/PNO1 axis. 1. Circ_0004674 is highly expressed in TNBC tissues and cells. 2. Circ_0004674 upregulates the expression of E2F6 by sponging miR-377-3p. 3. E2F6 upregulates PNO1 by binding to the PNO1 promoter. 4. Circ_0004674 favors TNBC progression by regulating the miR-377-3p/E2F6/PNO1 axis. 5. This study provides a new target for the treatment of TNBC.

A positive feedback loop of CENPU/E2F6/E2F1 facilitates proliferation and metastasis via ubiquitination of E2F6 in hepatocellular carcinoma.

Centromere protein U (CENPU), a centromere-binding protein required for cellular mitosis, has been reported to be closely associated with carcinogenesis in multiple malignancies; however, the role of CENPU in hepatocellular carcinoma (HCC) is still unclear. Herein, we investigated its biological role and molecular mechanism in the development of HCC. High CENPU expression in HCC tissue was observed and correlated positively with a poor prognosis in HCC patients. CENPU knockdown inhibited the proliferation, metastasis, and G1/S transition of HCC cells in vivo and in vitro, while ectopic expression of CENPU exerted the opposite effects. Mechanistically, CENPU physically interacted with E2F6 and promoted its ubiquitin-mediated degradation, thus affecting the transcription level of E2F1 and further accelerating the G1/S transition to promote HCC cell proliferation. E2F1 directly binds to the CENPU promoter and increases the transcription of CENPU, thereby forming a positive regulatory loop. Collectively, our findings indicate a crucial role for CENPU in E2F1-mediated signalling for cell cycle progression and reveal a role for CENPU as a predictive biomarker and therapeutic target for HCC patients.

Metabolic Reprogramming of Alloreactive T Cells Through TCR/MYC/mTORC1/E2F6 Signaling in aGvHD Patients.

Acute graft-versus-host disease (aGvHD) is the most common complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and significantly linked with morbidity and mortality. Although much work has been engaged to investigate aGvHD pathogenesis, the understanding of alloreactive T-cell activation remains incomplete. To address this, we studied transcriptional activation of carbohydrate, nucleotide, tricarboxylic acid (TCA) cycle, and amino acid metabolism of T cells before aGvHD onset by mining the Gene Expression Omnibus (GEO) datasets. Glycolysis had the most extensive correlation with other activated metabolic sub-pathways. Through Pearson correlation analyses, we found that glycolytic activation was positively correlated with activated CD4 memory T-cell subset and T-cell proliferation and migration. T-cell receptor (TCR), mechanistic target of rapamycin complex 1 (mTORC1), myelocytomatosis oncogene (MYC) signaling pathways and E2F6 might be "master regulators" of glycolytic activity. aGvHD predictive model constructed by glycolytic genes (PFKP, ENO3, and GAPDH) through logistic regression showed high predictive and discriminative value. Furthermore, higher expressions of PFKP, ENO3, and GAPDH in alloreactive T cells were confirmed in our pre-aGvHD patient cohort. And the predictive value of the aGvHD risk model was also validated. In summary, our study demonstrated that glycolytic activation might play a pivotal function in alloreactive T-cell activation before aGvHD onset and would be the potential target for aGvHD therapy.

rtcisE2F promotes the self-renewal and metastasis of liver tumor-initiating cells via N6-methyladenosine-dependent E2F3/E2F6 mRNA stability.

Liver cancer is highly heterogeneous, and the tumor tissue harbors a variety of cell types. Liver tumor initiating cells (TICs) well contribute to tumor heterogeneity and account for tumor initiation and metastasis, but the molecular mechanisms of liver TIC self-renewal are elusive. Here, we identified a functional read-through rt-circRNA, termed rtcisE2F, that is highly expressed in liver cancer and liver TICs. rtcisE2F plays essential roles in the self-renewal and activities of liver TICs. rtcisE2F targets E2F6 and E2F3 mRNAs, attenuates mRNA turnover, and increases E2F6/E2F3 expression. Mechanistically, rtcisE2F functions as a scaffold of N-methyladenosine (m6A) reader IGF2BP2 and E2F6/E2F3 mRNA. rtcisE2F promotes the association of E2F6/E2F3 mRNAs with IGF2BP2, and inhibits their association with another m6A reader, YTHDF2. IGF2BP2 inhibits E2F6/E2F3 mRNA decay, whereas YTHDF2 promotes E2F6/E2F3 mRNA decay. By switching m6A readers, rtcisE2F enhances E2F6/E2F3 mRNA stability. E2F6 and E2F3 are both required for liver TIC self-renewal and Wnt/ß-catenin activation, and inhibition of these pathways is a potential strategy for preventing liver tumorigenesis and metastasis. In conclusion, the rtcisE2F-IGF2BP2/YTHDF2-E2F6/E2F3-Wnt/ß-catenin axis drives liver TIC self-renewal and initiates liver tumorigenesis and metastasis, and may provide a strategy to eliminate liver TICs.

circ_0089153 exacerbates breast cancer cells proliferation and metastasis via sponging miR-2467-3p/E2F6.

The role of circ_0089153 in breast cancer (BCa) malignancy development was explored. circ_0089153 expression in BCa was analyzed by Gene Expression Omnibus database. Clinical tissues were obtained from 90 BCa patients. Cell counting kit-8 assay, 5-ethnyl-2 deoxyuridine assay and colony formation experiment were applied for proliferation detection. Wound healing assay and Transwell experiment were used for migration and invasion detection. Dual luciferase reporter gene assay, RNA immunoprecipitation assay and RNA pull-down assay were conducted. In vivo growth and metastasis of BCa cells were performed. Quantitative real-time polymerase chain reaction, Western blot and immunohistochemistry were applied for RNAs and proteins expression. The up-modulated circ_0089153 indicated an unfavorable survival of BCa patients. circ_0089153 knockdown attenuated BCa cells proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) (P < .01). circ_0089153 was miR-2467-3p sponge. Low miR-2467-3p expression indicated a worse survival of BCa patients. miR-2467-3p overexpression reduced BCa cells proliferation, migration, invasion and EMT (P < .05). circ_0089153 enhanced BCa cells proliferation, migration, invasion and EMT by sponging miR-2467-3p (P < .05). E2F6 was directly suppressed by miR-2467-3p. E2F6 high expression in BCa patients associated with worse survival. circ_0089153 knockdown suppressed in vivo BCa cells growth and lung metastasis (P < .01). circ_0089153 was an oncogene in breast cancer, which enhanced proliferation and metastasis through sponging miR-2467-3p/E2F6. circ_0089153 was suggested to be a potential target for BCa target treatment.

Long Noncoding RNA LAMTOR5-AS1 Interference Affects MicroRNA-506-3p/E2F6-Mediated Behavior of Non-Small Cell Lung Cancer Cells.

Long noncoding RNA LAMTOR5 antisense RNA 1 (LAMTOR5-AS1) has been certified as a risk predictor and diagnostic biomarker of prostate cancer. However, the expression and exact roles of LAMTOR5-AS1 in non-small cell lung cancer (NSCLC) remain unclear. Thus, we measured LAMTOR5-AS1 expression in NSCLC and gauged its clinical value. The detailed roles and downstream working mechanism of LAMTOR5-AS1 in NSCLC were comprehensively unraveled. qRT-PCR was applied to measure gene expression. Functionally, utilizing small interfering RNA, LAMTOR5-AS1 was ablated, and the functional alterations were addressed by means of different experiments. The targeting activities between LAMTOR5-AS1 and microRNA-506-3p (miR-506-3p) and between miR-506-3p and E2F transcription factor 6 (E2F6) were confirmed by RNA immunoprecipitation and luciferase reporter assays. LAMTOR5-AS1 overexpression in NSCLC was verified in TCGA datasets and our own cohort and manifested an evident relationship with poor prognosis. Interference with LAMTOR5-AS1 led to repression of the proliferation, cloning, and metastasis abilities of NSCLC cells in vitro. We further confirmed an obvious increase in LAMTOR5-AS1-silenced NSCLC cell apoptosis. Furthermore, the absence of LAMTOR5-AS1 restricted tumor growth in vivo. Mechanistically, LAMTOR5-AS1 sponged miR-506-3p in NSCLC cells. Furthermore, E2F6, a downstream target of miR-506-3p, was under the control of LAMTOR5-AS1, which was realized by decoying miR-506-3p. Rescue experiments showed that miR-506-3p suppression or E2F6 reintroduction was capable of remitting LAMTOR5-AS1 deficiency-triggered anticarcinogenic actions in NSCLC. Our study confirmed the exact roles of LAMTOR5-AS1 for the first time and revealed that LAMTOR5-AS1 knockdown disrupts the malignancy of NSCLC by targeting the miR-506-3p/E2F6 axis. Targeting the LAMTOR5-AS1/miR-506-3p/E2F6 pathway may be instrumental for managing patients with NSCLC.

Deubiquitination of the repressor E2F6 by USP22 facilitates AKT activation and tumor growth in hepatocellular carcinoma.

Dysregulated ubiquitination of tumor-related proteins plays a critical role in tumor development and progression. The deubiquitinase USP22 is aberrantly expressed in certain types of cancer and contributes to aggressive tumor progression. However, the precise mechanism underlying the pro-tumorigenic function of USP22 in hepatocellular carcinoma (HCC) remains unclear. Here, we report that E2F6, a pocket protein-independent transcription repressor, is essential for HCC cell growth, and that its activities are controlled by USP22-mediated deubiquitination. USP22 interacts with and stabilizes E2F6, resulting in the transcriptional repression of phosphatase DUSP1. Moreover, the process involving DUSP1 repression by E2F6 strengthens AKT activation in HCC cells. Therefore, these findings provide mechanistic insights into the USP22-mediated control of oncogenic AKT signaling, emphasizing the importance of USP22-E2F6 regulation in HCC development.

Long noncoding RNA SLC9A3­AS1 increases E2F6 expression by sponging microRNA­486­5p and thus facilitates the oncogenesis of nasopharyngeal carcinoma.

Long noncoding RNA SLC9A3 antisense RNA 1 (SLC9A3­AS1) plays a central role in lung cancer; yet, its functions in nasopharyngeal carcinoma (NPC) have not been elucidated. The present study revealed the roles of SLC9A3­AS1 in NPC and dissected the mechanisms downstream of SLC9A3­AS1. SLC9A3­AS1 levels in NPC were assessed by applying RT­qPCR. The modulatory role of SLC9A3­AS1 interference on NPC cells was examined using numerous functional experiments. High expression of SLC9A3­AS1 was observed in NPC samples. Patients with NPC with a high level of SLC9A3­AS1 experienced a shorter overall survival than those with a low SLC9A3­AS1 level. Loss of SLC9A3­AS1 reduced NPC cell proliferation, colony formation, migration, and invasion but induced cell apoptosis in vitro. Animal experiments further revealed that the depletion of SLC9A3­AS1 hindered NPC tumour growth in vivo. As a competitive endogenous RNA, SLC9A3­AS1 sponged microRNA­486­5p (miR­486­5p), consequently upregulating E2F transcription factor 6 (E2F6). Finally, the effects of SLC9A3­AS1 silencing on NPC cells were reversed by inhibiting miR­486­5p or overexpressing E2F6. In summary, SLC9A3­AS1 exerted carcinogenic effects on NPC cells by adjusting the miR­486­5p/E2F6 axis. Accordingly, the newly identified SLC9A3­AS1/miR­486­5p/E2F6 pathway may offer attractive therapeutic targets for future development.

Expression patterns for Bcl-2, EMA, ß-catenin, E-cadherin, PAX8, and MIB1 in thymomas.

The expression of immunohistochemical markers has been extensively investigated in thymomas to assist in the differential diagnosis. We have studied six select markers to determine their utility in the evaluation of these tumors. A series of 126 thymomas including 33 type A, 27 type AB, 20 type B1, 22 type B2, and 24 type B3, were examined utilizing a tissue microarray (TMA) technique with antibodies to e-cadherin, ß-catenin, PAX8, bcl-2, EMA, and MIB-1. Keratin AE1/AE3 and p63 were used for quality control. A significant finding was strong and consistent positivity for bcl-2 in type A (90%) and type AB (88.8%) thymoma, while 100% of B1, B2, and B3 were negative. The distribution of e-cadherin and ß-catenin was not useful for differential diagnosis. E-cadherin and ß-catenin were expressed in a high proportion of all the tumors (92-100%), except for B2 thymoma which showed only 45% expression. A significant increase in the expression of the MIB-1 proliferation marker (mean: 12.8% nuclear positivity) was also observed in B3 thymoma compared with the other histologic types. Statistical significance was confirmed using Kruskal's non-parameterized test for distribution. EMA was generally negative except for spindle cells in the fibrous septa in types A and AB thymoma. PAX8 showed less consistent nuclear staining than p63 and was only widely expressed in 55.7% of cases. Bcl-2 may serve as a useful marker to separate spindle cell thymomas (Type A and AB) from the other types, and the MIB1 proliferation index may be of use to differentiate type B2 from type B3 thymoma.

Circular RNA ZNF609 functions as a competing endogenous RNA in regulating E2F transcription factor 6 through competitively binding to microRNA-197-3p to promote the progression of cervical cancer progression.

Countless studies have demonstrated that Circular RNAs (circRNAs) exert vital effects in regulating tumorigenesis of various cancers. CircRNA ZNF609 (circ-ZNF609) has been reported as an oncogene in various human cancers. Nevertheless, its regulating effect in cervical cancer (CC) remains to be further explored. RT-qPCR was adopted to measure circ-ZNF609, miR-197-3p and E2F6 levels. CC cell proliferation, migration and invasion were analyzed via CCK-8 and transwell assays. Dual-luciferase reporter assay was adopted to confirm the interaction between miR-197-3p and circ-ZNF609 or E2F6. In the present study, it was found that circ-ZNF609 was elevated in CC tissues and cell lines, and circ-ZNF609 deletion repressed cell viability, migration and invasion in CC. Moreover, circ-ZNF609 was identified to negatively regulate miR-197-3p expression in CC cells. The inhibition of miR-197-3p abrogated the inhibitory effect on CC cell proliferation, migration and invasion induced by circ-ZNF609 knockdown. Additionally, we further demonstrated that circ-ZNF609 upregulated E2F6 by interacting with miR-197-3p. Finally, rescue assays indicated that E2F6 overexpression upended the suppression of CC progression induced by circ-ZNF609 deletion. In conclusion, circ-ZNF609 promoted CC progression through modulating the miR-197-3p/E2F6 axis as an oncogene. This finding offers a unique insight into CC molecular mechanism and suggests a potential target for CC therapy.

Epstein-Barr Virus Facilitates Expression of KLF14 by Regulating the Cooperative Binding of the E2F-Rb-HDAC Complex in Latent Infection.

Epstein-Barr virus (EBV) was discovered as the first human tumor virus more than 50 years ago. EBV infects more than 90% of the human population worldwide and is associated with numerous hematologic malignancies and epithelial malignancies. EBV establishes latent infection in B cells, which is the typical program seen in lymphomagenesis. Understanding EBV-mediated transcription regulatory networks is one of the current challenges that will uncover new insights into the mechanism of viral-mediated lymphomagenesis. Here, we describe the regulatory profiles of several cellular factors (E2F6, E2F1, Rb, HDAC1, and HDAC2) together with EBV latent nuclear antigens using next-generation sequencing (NGS) analysis. Our results show that the E2F-Rb-HDAC complex exhibits similar distributions in genomic regions of EBV-positive cells and is associated with oncogenic super-enhancers involving long-range regulatory regions. Furthermore, EBV latent antigens cooperatively hijack this complex to bind at KLFs gene loci and facilitate KLF14 gene expression in lymphoblastoid cell lines (LCLs). These results demonstrate that EBV latent antigens can function as master regulators of this multisubunit repressor complex (E2F-Rb-HDAC) to reverse its suppressive activities and facilitate downstream gene expression that can contribute to viral-induced lymphomagenesis. These results provide novel insights into targets for the development of new therapeutic interventions for treating EBV-associated lymphomas.IMPORTANCE Epstein-Barr virus (EBV), as the first human tumor virus, infects more than 90% of the human population worldwide and is associated with numerous human cancers. Exploring EBV-mediated transcription regulatory networks is critical to understand viral-associated lymphomagenesis. However, the detailed mechanism is not fully explored. Now we describe the regulatory profiles of the E2F-Rb-HDAC complex together with EBV latent antigens, and we found that EBV latent antigens cooperatively facilitate KLF14 expression by antagonizing this multisubunit repressor complex in EBV-positive cells. This provides potential therapeutic targets for the treatment of EBV-associated cancers.

LncRNA TMPO-AS1 promotes LCN2 transcriptional activity and exerts oncogenic functions in ovarian cancer.

Ovarian cancer remains the sixth most frequently occurring cancer in women worldwide. Long noncoding RNAs (lncRNAs) are capable of regulating gene expression, and thus, participating in a wide range of biological functions and disease processes including cancer development. Our work suggests that lncRNA TMPO antisense RNA 1 (TMPO-AS1) represents an oncogenic lncRNA in ovarian cancer and presents a novel mechanism involving transcription factor E2F transcription factor 6 (E2F6) and lipocalin-2 (LCN2). We identified upregulated lncRNA TMPO-AS1 in ovarian cancer tissues and cells. siRNA-mediated silencing of lncRNA TMPO-AS1 restrained the aggressiveness of ovarian cancer cells and their pro-angiogenic ability, and reduced the expression of LCN2. LncRNA TMPO-AS1 was found to interact with E2F6, a transcriptional repressor that could bind to the promoter region of LCN2 gene. In addition, silencing of E2F6 or overexpression of LCN2 restored the aggressiveness of ovarian cancer cells and their pro-angiogenic ability following siRNA-mediated silencing of lncRNA TMPO-AS1. Taken together, we demonstrated lncRNA TMPO-AS1 could potentially promote LCN2 transcriptional activity by binding to transcription factor E2F6, and thus, stimulated the progression of ovarian cancer. These findings underscore a possible alternative therapeutic strategy for ovarian cancer treatment.

CREB acts as a common transcription factor for major epigenetic repressors; DNMT3B, EZH2, CUL4B and E2F6.

CREB signaling is known for several decades, but how it regulates both positive and negative regulators of cell proliferation is not well understood. On the other hand functions of major epigenetic repressors such as DNMT3B, EZH2 and CUL4B for their repressive epigenetic modifications on chromatin have also been well studied. However, there is very limited information available on how these repressors are regulated at their transcriptional level. Here, using computational tools and molecular techniques including site directed mutagenesis, promoter reporter assay, chromatin immunoprecipitation (ChIP), we identified that CREB acts as a common transcription factor for DNMT3B, EZH2, CUL4B and E2F6. ChIP assay revealed that pCREB binds to promoters of these repressors at CREs and induce their transcription. As expected, the expression of these repressors and their associated repressive marks particularly H3K27me3 and H2AK119ub are increased and decreased upon CREB overexpression and knock-down conditions respectively in the cancer cells indicating that CREB regulates the functions of these repressors by activating their transcription. Since CREB and these epigenetic repressors are overexpressed in various cancer types, our findings showed the molecular relationship between them and indicate that CREB is an important therapeutic target for cancer therapy.

E2F6-Mediated Downregulation of MIR22HG Facilitates the Progression of Laryngocarcinoma by Targeting the miR-5000-3p/FBXW7 Axis.

Recently, abundant evidence has clarified that long noncoding RNAs (lncRNAs) play an oncogenic or anticancer role in the tumorigenesis and development of diverse human cancers. Described as a crucial regulator in some cancers, MIR22HG has not yet been studied in laryngocarcinoma and therefore the underlying regulatory role of MIR22HG in laryngocarcinoma is worth detecting. In this study, MIR22HG expression in laryngocarcinoma cells was confirmed to be downregulated, and upregulated MIR22HG expression led to suppressive effects on laryngocarcinoma cell proliferation and migration. Molecular mechanism assays revealed that MIR22HG sponges miR-5000-3p in laryngocarcinoma cells. Besides, decreased expression of miR-5000-3p suppressed laryngocarcinoma cell proliferation and migration. Moreover, the FBXW7 gene was reported to be a downstream target gene of miR-5000-3p in laryngocarcinoma cells. More importantly, rescue assays verified that FBXW7 depletion or miR-5000-3p upregulation countervailed the repressive effects of MIR22HG overexpression on laryngocarcinoma progression. In addition, E2F6 was proved to be capable of inhibiting MIR22HG transcription in laryngocarcinoma cells. To sum up, E2F6-induced downregulation of MIR22HG promotes laryngocarcinoma progression through the miR-5000-3p/FBXW7 axis.

LncRNA GHET1 promotes cervical cancer progression through regulating AKT/mTOR and Wnt/ß-catenin signaling pathways.

Cervical cancer (CC) is a prevalent gynecological cancer, and the patients with CC usually suffer from dismal prognosis. Long non-coding RNAs (lncRNAs) are demonstrated to serve as promising biological targets in human cancers. Gastric carcinoma proliferation enhancing transcript 1 (GHET1) has been revealed to function as an oncogene in several cancers, but it has never been investigated in CC. We proposed to examine the biological role of GHET1 in CC and the underlying mechanism and validated the up-regulated expression of GHET1 in CC cell lines. Loss-of-function assays demonstrated that down-regulation of GHET1 inhibited cell growth, migration and epithelial-to-mesenchymal transition (EMT) in CC. Furthermore, we validated that GHET1 down-regulation could inactivate AKT/mTOR and Wnt/ß-catenin pathways, and that respective activation of these two pathways abrogated the inhibitive effect of GHET1 knockdown on CC cell growth, migration and EMT. Moreover, we unfolded a preliminary investigation on the modulation of GHET1 on AKT/mTOR and Wnt/ß-catenin pathways. We found that GHET1 stabilized E2F6 mRNA through interacting with IGF2BP2, so as to regulate the activity of AKT/mTOR and Wnt/ß-catenin pathways. Rescue assays also proved that GHET1 regulated these two pathways and CC cell growth, migration and EMT through E2F6. In conclusion, we revealed that down-regulation of GHET1 suppresses cervical cancer progression through regulating AKT/mTOR and Wnt/ß-catenin signaling pathways, indicating GHET1 as a promising molecular biomarker for CC treatment improvement.

E2F6-mediated lncRNA CASC2 down-regulation predicts poor prognosis and promotes progression in gastric carcinoma.

AIMS: To investigate the potential biological role of E2F6 and its underlying molecular mechanism in gastric carcinoma (GC) progression. MAIN METHODS: The expressions of cancer susceptibility candidate 2 (CASC2), E2F6 and matrix metalloprotein-2 (MMP-2) were measured by quantitative real-time polymerase chain reaction and western blotting. The inhibitory effect of E2F6 on CASC2 was evaluated using luciferase reporter assay. Cell growth was assessed by colony formation assay and cell counting kit-8. Cell invasion and apoptosis were measured by transwell assay and flow cytometry assay, respectively. In vivo tumorigenicity was assessed by tumor xenografts in nude mice. KEY FINDINGS: Our data revealed that CASC2 was downregulated while E2F6 was upregulated in GC tissues and cell lines. Remarkably, lower expression of CASC2 was associated with poor survival in GC patients. E2F6 inhibited the expression of CASC2. Subsequently, reliable data showed that downregulation of E2F6 suppressed the proliferation and invasion, and promoted the apoptosis of GC cells. Furthermore, downregulation of E2F6 decreased the expression of MMP-2 and increased the activity of caspase-3. However, these changes triggered by E2F6 knockdown could be reversed by inhibition of CASC2. Moreover, we also proved that downregulation of CASC2 reverses the effect of E2F6 knockdown on tumor growth in vivo. SIGNIFICANCE: Our data demonstrated that E2F6 could regulate the proliferation, invasion and apoptosis of GC cells via inhibiting the expression of CASC2, suggesting that E2F6/CASC2 axis is another regulator of GC progression.

Could EMA and cytokeratin 7 be useful in distinguishing tricholemmal carcinoma from clear-cell squamous cell carcinoma? A case series from our department and a brief review of the literature.

Tricholemmal carcinoma is a malignant cutaneous adnexal tumor showing outer root sheath differentiation, thought to be the malignant counterpart of trichilemmoma. Although the real existence of tricholemmal carcinoma continues to be a matter of debate, it has been introduced in the recently published 4th edition of World Health Organization classification of skin tumors. Herein, we evaluated whether immunohistochemistry (EMA, CK7, CK5/14, p63, p16, and Ber-EP4) supports tricholemmal carcinoma as a separate entity and whether it could be useful in this differential diagnosis. A total of 9 cases, 3 tricholemmal carcinomas and 6 clear-cell squamous cell carcinomas were evaluated on the basis of histological criteria suggested by the WHO. In our opinion, although these results need to be validated in larger series, they support tricholemmal carcinoma as a separate entity and suggest an immunohistochemical profile (clear-cell squamous cell carcinomas: EMA diffusely positive, CK7 negative; tricholemmal carcinoma: EMA negative, CK7 patchy or moderately positive) that could be useful for this differential diagnosis.