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The E2F family of transcription factors is crucial for cell cycle progression and cell fate decisions. Although E2Fs have been widely studied in mammals, there have been few studies performed in insects. Here, we determined the function of E2F4 in the silkworm, Bombyx mori. We demonstrate that E2F proteins are highly conserved among species from lower animals to higher mammals. Overexpression of the BmE2F4 gene led to cell cycle arrest in the G1 phase, whereas interfering with the BmE2F4 mRNA led to accumulation of cells in the S phase. These results indicate that BmE2F4 is important in cell cycle regulation. We also demonstrate that the BmE2F4 gene is involved in DNA replication of BmN‐SWU1 cells and DNA synthesis in the silk gland. Furthermore, we identified a protein called Bm14‐3‐3ζ that can interact with BmE2F4 and allow it to localize in the nucleus. Overexpression of the Bm14‐3‐3ζ gene led to cell cycle arrest in the G1 phase, while knocking down the gene increased the proportion of cells in S phase. These findings provide important insights into the function of E2F transcription factors and increase our understanding of their involvement in cell cycle regulation. [ABSTRACT FROM AUTHOR]

E2f4 is a multifunctional transcription factor that is essential for many cellular processes. Although the role of E2f4 during cell cycle progression has been investigated in great detail, less is known about E2f4 during embryonic development. Here, we investigated the role of E2f4 during zebrafish development. Zebrafish e2f4 mutants displayed ectopic otolith formation due to abnormal ciliary beating in the otic vesicle. The beating defects of motile cilia were caused by abnormal expression of ciliary motility genes. The expression of two genes, lrrc23 and ccdc151, were significantly decreased in the absence of E2f4. In addition to that, e2f4 mutants also displayed growth retardation both in the body length and body weight and mostly died at around 6 months old. Although food intake was normal in the mutants, we found that the microvilli of the intestinal epithelia were significantly affected in the mutants. Finally, the intestinal epithelia of e2f4 mutants also displayed reduced cell proliferation, together with an increased level of cell apoptosis. Our data suggested a tissue‐specific role of E2f4 during zebrafish development, which is distinct from the traditional views of this protein as a transcription repressor. [ABSTRACT FROM AUTHOR]

Cervical cancer is one of the major malignancies, the pathophysiology and progression of which remain to be scarcely understood. Long non‐coding RNAs (lncRNAs) have been previously implicated in the progression of cervical cancer. Here, the purpose of this study was to identify whether lncRNA heart‐ and neural crest derivative‐expressed 2‐antisense RNA 1 (HAND2‐AS1) affect the development of cervical cancer through regulation of chromosome 16 open reading frame 74 (C16orf74) by mediating a transcription factor E2F4. RT‐qPCR was performed to determine the expression of HAND2‐AS1 in cervical cancer cells. Then, cervical cancer cells were treated with HAND2‐AS1 or si‐E2F4 RNA, or C16orf74, after which the proliferation, colony formation, migration and invasion were detected. Moreover, the binding between HAND2‐AS1 and E2F4 or between E2F4 and C16orf74 was explored. Finally, the tumorigenesis of cervical cancer cells was measured in nude mice with altered HAND2‐AS1/E2F4/C16orf74 expression. HAND2‐AS1 exhibited poor expression in cervical cancer, and HAND2‐AS1 overexpression suppressed the proliferation, colony formation, migration and invasion of cervical cancer cells. In addition, HAND2‐AS1 was found to recruit transcription factor E2F4 to C16orf74 promoter region and down‐regulate C16orf74 expression. Lastly, HAND2‐AS1/E2F4/C16orf74 modulated the tumorigenesis of cervical cancer in nude mice. In conclusion, this study provided evidence on the inhibitory effect of HAND2‐AS1 on the development of cervical cancer through the suppression of C16orf74 expression by recruiting transcription factor E2F4. This study highlights the potential of lncRNA HAND2‐AS1 as a target in the treatment of cervical cancer. [ABSTRACT FROM AUTHOR]

Esophageal cancer represents the eighth most frequently occurring cancer, as well as the sixth most widespread cause of cancer‐related deaths. In recent years, accumulating evidence has implicated long non‐coding RNAs in the progression of esophageal squamous cell carcinoma (ESCC). The aim of the present study was to investigate the potential involvement and underlying mechanisms of LINC00337 in ESCC. Expression patterns of LINC00337 and targeting protein for Xenopus kinesin‐like protein 2 (TPX2) in ESCC tissues and cells were detected using RT‐qPCR and immunohistochemical staining. Next, the interactions among LINC00337, E2F4, and TPX2 were identified using chromatin immunoprecipitation, dual‐luciferase reporter, and RNA‐binding protein immunoprecipitation assays, suggesting that LINC00337 could recruit E2F4 to enhance the transcription of TPX2. Thereafter, the regulatory roles of LINC00337 and TPX2 in ESCC were analyzed by altering the expression of LINC00337 or TPX2 in ESCC cells following treatment with cisplatin (DDP). The levels of autophagy‐related proteins Beclin1 and LC3II/LC3I, viability, autophagy, apoptosis, and chemoresistance of ESCC cells to DDP were measured following transfection treatment with different plasmids. Additionally, the role of the LINC00337/E2F4/TPX2 axis was assessed in vivo by measuring tumor formation in nude mice. The results demonstrated that LINC00337 upregulated TPX2, consequently leading to elevated levels of Beclin1 and LC3II/LC3I, promoted cell viability and autophagy, while inhibiting apoptosis and chemosensitivity to DDP in ESCC. In sum, the current study evidenced that the overexpression of LINC00337 could potentially enhance ESCC cell autophagy and chemoresistance to DDP via the upregulation of TPX2 by recruiting E2F4. Thus, LINC00337 may serve as a potential candidate for the treatment of ESCC. [ABSTRACT FROM AUTHOR]

Acute myeloid leukaemia (AML) is an aggressive and mostly incurable haematological malignancy with frequent relapse after an initial response to standard chemotherapy. Therefore, novel therapies are urgently required to improve AML clinical outcome. Here, we aim to study the dysregulation of a particular transcription factor, E2F4, and its role in the progression of AML. In this study, human clinical data from the Gene Expression Profiling Interactive Analysis (GEPIA) revealed that increased E2F4 expression was associated with poor prognosis in AML patients. Moreover, the experimental results showed that E2F4 was aberrantly overexpressed in human AML patients and cell lines. Depletion of E2F4 inhibited the proliferation, induced the differentiation and suppressed the growth of AML cells in a nude mouse model. By contrast, overexpression of E2F4 promoted the proliferation and inhibited the differentiation of AML cells in vitro. Additionally, E2F4 expression not only is positively correlated with EZH2 but also can bind to EZH2. RNA microarray results also showed that E2F4 can regulate MAPK signalling pathway. EZH2 can reverse the inhibitory effect of E2F4 silencing on MAPK signaling pathway. In summary, our data suggest that E2F4 may be a potential therapeutic target for AML therapy. [ABSTRACT FROM AUTHOR]

The transcription factor E2F4 is a key determinant of cell differentiation and cell‐cycle progression, but its function and regulatory mechanism are not completely understood. Here, we report that E2F4 acts as a positive regulator of the biosynthesis of milk components and proliferation of bovine mammary epithelial cells (BMECs). Overexpression of E2F4 in BMECs resulted in the upregulation of β‐casein, triglyceride, and lactose levels and increased cell proliferation, whereas E2F4 knockdown by small interfering RNA had the opposite effects. We further detected that overexpression of E2F4 significantly increased the messenger RNA expression of mTOR, SREBP‐1c, and Cyclin D1, and increased protein levels of SREBP‐1c, and Cyclin D1, and the ratio of p‐mTOR/mTOR, whereas E2F4 knockdown had the opposite effects. E2F4 was almost entirely located in the nucleus, and we further identified, via ChIP‐qPCR analysis, that mTOR, SREBP‐1c, and Cyclin D1 were E2F4 target genes, and exogenous administration of methionine, leucine, β‐estradiol, and prolactin markedly increased the protein levels of E2F4 and its binding to the promoters of these three genes. In summary, our data reveal that E2F4 responds to extracellular stimuli and regulates the expression of mTOR, SREBP‐1c, and Cyclin D1 for milk biosynthesis and proliferation of BMECs. [ABSTRACT FROM AUTHOR]

Bone‐marrow‐derived mesenchymal stem cells (MSCs) have great potential in transplantation medicine due to their multiple advantages. However, the controlled differentiation of MSCs is one of the key aspects of effective clinical transplantation. Growing evidence suggests that the cell cycle plays an important role in regulating differentiation, while p130 and E2F4 are key to cell cycle checkpoints. The aim of the study is to evaluate the effects and mechanism of p130/E2F4 on the multidifferentiation of MSCs. Our data showed that the transduction efficiencies of p130 or E2F4 mediated by lentiviral vectors were 80.3%–84.4%. p130 and E2F4 mRNA expression was significantly higher in MSC‐p130 and MSC‐E2F4 cells than in MSC normal control (NC) cells. Similar results were also observed for p130 and E2F4 protein expression. After osteogenic or adipogenic differentiation, the G1 phase was significantly delayed in the MSC‐p130 and MSC‐E2F4 groups compared with that in the MSC‐NC group. However, the G1 phase in the MSC‐p130 and MSC‐E2F4 groups did the opposite after chondrogenic differentiation. Moreover, overexpressing p130 or E2F4 significantly improved osteogenic differentiation while inhibiting adipogenic and chondrogenic differentiation of mouse MSCs (mMSCs). Moreover, overexpressing p130 or E2F4 significantly improved migration but not proliferation of mMSCs. Our data suggest that cell cycle regulation may be involved in p130/E2F4‐mediated changes in the multipotential abilities of bone‐marrow‐derived mMSCs. The controlled differentiation of mesenchymal stem cells (MSCs) is one of the key aspects of effective clinical transplantation. Growing evidence suggests that the cell cycle plays an important role in regulating differentiation, while p130 and E2F4 are key to cell cycle checkpoints. Our data suggest that cell cycle regulation may be involved in p130/E2F4‐mediated changes in the multipotential abilities of bone‐marrow‐derived mouse MSCs (mMSCs). [ABSTRACT FROM AUTHOR]

The retinoblastoma (pRb)-retated p130 pocket protein is a regulator of cell growth and differentiation, and a candidate tumour suppressor. Both pRb and p130 operate through interactions with cellular proteins, including the E2F transcription factors. White such interactions are controlled by phosphorylation of multiple sites of pRb, regulation of p130 remains poorly understood. We now identify 22 in vivo phosphorylation sites of p130, targeted by diverse kinases, and present evidence for three cyclin-dependent kinase 4(6) [Cdk4(6)] specific phosphorylations, which appear critical for controlling the growth-restraining activity of p130. When expressed in U2OS cells, the phosphorylation-deficient mutant p130ΔCdk4, in which the Cdk4 specific sites were mutated to alanine residues, imposed a more sustained G1 arrest than a constitutively active pRbΔCdk, known to repress all cellular E2F activity. Experiments using p130ΔCdk4 and another phosphorylation-deficient mutant, p130PM19A, with 19 phosphorylation sites mutated, revealed that the p130-imposed G1 block reflects cooperative growth-suppressive effects of phosphorylation-regulated E2F binding and phosphorylation-independent sequestration of cyclin E(A)-Cdk2 through the N-terminal cyclin binding motif of p130. [ABSTRACT FROM AUTHOR]