RESUMO
Mammalian SWI/SNF (BAF) chromatin remodelers play dosage-sensitive roles in many human malignancies and neurologic disorders. The gene encoding the BAF subunit actin-like 6a (ACTL6A) is amplified early in the development of many squamous cell carcinomas (SCCs), but its oncogenic role remains unclear. Here we demonstrate that ACTL6A overexpression leads to its stoichiometric assembly into BAF complexes and drives their interaction and engagement with specific regulatory regions in the genome. In normal epithelial cells, ACTL6A was substoichiometric to other BAF subunits. However, increased ACTL6A levels by ectopic expression or in SCC cells led to near saturation of ACTL6A within BAF complexes. Increased ACTL6A occupancy enhanced polycomb opposition genome-wide to activate SCC genes and facilitated the co-dependent loading of BAF and TEAD-YAP complexes on chromatin. Both mechanisms appeared to be critical and function as a molecular AND gate for SCC initiation and maintenance, thereby explaining the specificity of the role of ACTL6A amplification in SCCs.
Assuntos
Actinas/metabolismo , Carcinoma de Células Escamosas/metabolismo , Montagem e Desmontagem da Cromatina , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Actinas/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Cromatina/genética , Proteínas Cromossômicas não Histona/genética , Proteínas de Ligação a DNA/genética , Epigênese Genética , Amplificação de Genes , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Proteínas do Grupo Polycomb/genética , Ligação Proteica , Fatores de Transcrição de Domínio TEA/genética , Fatores de Transcrição de Domínio TEA/metabolismo , Proteínas de Sinalização YAP/genética , Proteínas de Sinalização YAP/metabolismoRESUMO
The transition of mouse embryonic stem cells (ESCs) between serum/LIF and 2i(MEK and GSK3 kinase inhibitor)/LIF culture conditions serves as a valuable model for exploring the mechanisms underlying ground and confused pluripotent states. Regulatory networks comprising core and ancillary pluripotency factors drive the gene expression programs defining stable naïve pluripotency. In our study, we systematically screened factors essential for ESC pluripotency, identifying TEAD2 as an ancillary factor maintaining ground-state pluripotency in 2i/LIF ESCs and facilitating the transition from serum/LIF to 2i/LIF ESCs. TEAD2 exhibits increased binding to chromatin in 2i/LIF ESCs, targeting active chromatin regions to regulate the expression of 2i-specific genes. In addition, TEAD2 facilitates the expression of 2i-specific genes by mediating enhancer-promoter interactions during the serum/LIF to 2i/LIF transition. Notably, deletion of Tead2 results in reduction of a specific set of enhancer-promoter interactions without significantly affecting binding of chromatin architecture proteins, CCCTC-binding factor (CTCF), and Yin Yang 1 (YY1). In summary, our findings highlight a novel prominent role of TEAD2 in orchestrating higher-order chromatin structures of 2i-specific genes to sustain ground-state pluripotency.
Assuntos
Cromatina , Proteínas de Ligação a DNA , Células-Tronco Pluripotentes , Fatores de Transcrição de Domínio TEA , Animais , Camundongos , Cromatina/metabolismo , Cromatina/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Elementos Facilitadores Genéticos , Células-Tronco Embrionárias Murinas/metabolismo , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Pluripotentes/metabolismo , Células-Tronco Pluripotentes/citologia , Regiões Promotoras Genéticas , Fatores de Transcrição de Domínio TEA/genética , Fatores de Transcrição de Domínio TEA/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genéticaRESUMO
As an output effector of the Hippo signaling pathway, the TEAD transcription factor and co-activator YAP play crucial functions in promoting cell proliferation and organ size. The tumor suppressor NF2 has been shown to activate LATS1/2 kinases and interplay with the Hippo pathway to suppress the YAP-TEAD complex. However, whether and how NF2 could directly regulate TEAD remains unknown. We identified a direct link and physical interaction between NF2 and TEAD4. NF2 interacted with TEAD4 through its FERM domain and C-terminal tail and decreased the protein stability of TEAD4 independently of LATS1/2 and YAP. Furthermore, NF2 inhibited TEAD4 palmitoylation and induced the cytoplasmic translocation of TEAD4, resulting in ubiquitination and dysfunction of TEAD4. Moreover, the interaction with TEAD4 is required for NF2 function to suppress cell proliferation. These findings reveal an unanticipated role of NF2 as a binding partner and inhibitor of the transcription factor TEAD, shedding light on an alternative mechanism of how NF2 functions as a tumor suppressor through the Hippo signaling cascade.
Assuntos
Via de Sinalização Hippo , Neurofibromina 2 , Proteínas Serina-Treonina Quinases , Transdução de Sinais , Fatores de Transcrição de Domínio TEA , Humanos , Proliferação de Células , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Células HEK293 , Lipoilação , Neurofibromina 2/metabolismo , Neurofibromina 2/genética , Ligação Proteica , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Estabilidade Proteica , Fatores de Transcrição de Domínio TEA/metabolismo , Proteínas Supressoras de Tumor , UbiquitinaçãoRESUMO
Transforming growth factor-ß (TGF-ß) and Hippo signaling are two critical pathways engaged in cancer progression by regulating both oncogenes and tumor suppressors, yet how the two pathways coordinately exert their functions in the development of hepatocellular carcinoma (HCC) remains elusive. In this study, we firstly conducted an integrated analysis of public liver cancer databases and our experimental TGF-ß target genes, identifying CYR61 as a pivotal candidate gene relating to HCC development. The expression of CYR61 is downregulated in clinical HCC tissues and cell lines than that in the normal counterparts. Evidence revealed that CYR61 is a direct target gene of TGF-ß in liver cancer cells. In addition, TGF-ß-stimulated Smad2/3 and the Hippo pathway downstream effectors YAP and TEAD4 can form a protein complex on the promoter of CYR61, thereby activating the promoter activity and stimulating CYR61 gene transcription in a collaborative manner. Functionally, depletion of CYR61 enhanced TGF-ß- or YAP-mediated growth and migration of liver cancer cells. Consistently, ectopic expression of CYR61 was capable of impeding TGF-ß- or YAP-induced malignant transformation of HCC cells in vitro and attenuating HCC xenograft growth in nude mice. Finally, transcriptomic analysis indicates that CYR61 can elicit an antitumor program in liver cancer cells. Together, these results add new evidence for the crosstalk between TGF-ß and Hippo signaling and unveil an important tumor suppressor function of CYR61 in liver cancer.
Assuntos
Carcinoma Hepatocelular , Proteína Rica em Cisteína 61 , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas , Fator de Crescimento Transformador beta , Proteínas de Sinalização YAP , Animais , Humanos , Camundongos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Movimento Celular , Proteína Rica em Cisteína 61/metabolismo , Proteína Rica em Cisteína 61/genética , Mineração de Dados , Regulação Neoplásica da Expressão Gênica/genética , Via de Sinalização Hippo , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/genética , Camundongos Nus , Regiões Promotoras Genéticas , Transdução de Sinais/genética , Proteína Smad2/metabolismo , Proteína Smad2/genética , Proteína Smad3/metabolismo , Proteína Smad3/genética , Fatores de Transcrição de Domínio TEA/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/genética , Regulação para Cima , Proteínas de Sinalização YAP/metabolismo , Proteínas de Sinalização YAP/genéticaRESUMO
Transcriptional enhanced associate domain family members 1 to 4 (TEADs) are a family of four transcription factors and the major transcriptional effectors of the Hippo pathway. In order to activate transcription, TEADs rely on interactions with other proteins, such as the transcriptional effectors Yes-associated protein and transcriptional co-activator with PDZ-binding motif. Nuclear protein interactions involving TEADs influence the transcriptional regulation of genes involved in cell growth, tissue homeostasis, and tumorigenesis. Clearly, protein interactions for TEADs are functionally important, but the full repertoire of TEAD interaction partners remains unknown. Here, we employed an affinity purification mass spectrometry approach to identify nuclear interacting partners of TEADs. We performed affinity purification mass spectrometry experiment in parallel in two different cell types and compared a wildtype TEAD bait protein to a nuclear localization sequence mutant that does not localize to the nucleus. We quantified the results using SAINT analysis and found a significant enrichment of proteins linked to DNA damage including X-ray repair cross-complementing protein 5 (XRCC5), X-ray repair cross-complementing protein 6 (XRCC6), poly(ADP-ribose) polymerase 1 (PARP1), and Rap1-interacting factor 1 (RIF1). In cellular assays, we found that TEADs co-localize with DNA damage-induced nuclear foci marked by histone H2AX phosphorylated on S139 (γH2AX) and Rap1-interacting factor 1. We also found that depletion of TEAD proteins makes cells more susceptible to DNA damage by various agents and that depletion of TEADs promotes genomic instability. Additionally, depleting TEADs dampens the efficiency of DNA double-stranded break repair in reporter assays. Our results connect TEADs to DNA damage response processes, positioning DNA damage as an important avenue for further research of TEAD proteins.
Assuntos
Dano ao DNA , Reparo do DNA , Fatores de Transcrição de Domínio TEA , Humanos , Carcinogênese/metabolismo , Reparo do DNA/fisiologia , Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição de Domínio TEA/metabolismoRESUMO
Podocyte injury and loss are hallmarks of diabetic nephropathy (DN). However, the molecular mechanisms underlying these phenomena remain poorly understood. YAP (Yes-associated protein) is an important transcriptional coactivator that binds with various other transcription factors, including the TEAD family members (nuclear effectors of the Hippo pathway), that regulate cell proliferation, differentiation, and apoptosis. The present study found an increase in YAP phosphorylation at S127 of YAP and a reduction of nuclear YAP localization in podocytes of diabetic mouse and human kidneys, suggesting dysregulation of YAP may play a role in diabetic podocyte injury. Tamoxifen-inducible podocyte-specific Yap gene knockout mice (YappodKO) exhibited accelerated and worsened diabetic kidney injury. YAP inactivation decreased transcription factor WT1 expression with subsequent reduction of Tead1 and other well-known targets of WT1 in diabetic podocytes. Thus, our study not only sheds light on the pathophysiological roles of the Hippo pathway in diabetic podocyte injury but may also lead to the development of new therapeutic strategies to prevent and/or treat DN by targeting the Hippo signaling pathway.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Diabetes Mellitus Experimental , Nefropatias Diabéticas , Camundongos Knockout , Fosfoproteínas , Podócitos , Transdução de Sinais , Fatores de Transcrição , Proteínas WT1 , Proteínas de Sinalização YAP , Podócitos/metabolismo , Podócitos/patologia , Animais , Proteínas WT1/metabolismo , Proteínas WT1/genética , Proteínas de Sinalização YAP/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Nefropatias Diabéticas/patologia , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/etiologia , Nefropatias Diabéticas/genética , Humanos , Fosforilação , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/complicações , Fosfoproteínas/metabolismo , Fosfoproteínas/genética , Fatores de Transcrição de Domínio TEA/metabolismo , Via de Sinalização Hippo , Camundongos , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Masculino , Camundongos Endogâmicos C57BL , Tamoxifeno/farmacologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genéticaRESUMO
A number of studies have confirmed that Yes-associated protein (YAP)/transcriptional co-activator with PDZ-binding motif (TAZ)-transcriptional enhanced associate domain (TEAD) activity is the driver of cancer development. However, the role and mechanism of the YAP/TAZ-TEAD pathway in cervical intraepithelial neoplasia (CIN) remain to be clarified. Therefore, this study was designed to observe the effect of YAP/TAZ-TEAD activity on the development of CIN and provide new ideas for the diagnosis and treatment of CIN. Firstly, cervical tissues were collected from CIN patients in different stages [CIN grade 1 (CIN1) tissue, CIN grade 2/3 (CIN 2/3) and squamous cell carcinoma (SCC)] and healthy volunteers. Next, the expression levels of YAP, TAZ and TEAD in cervical tissues and cells were observed by immunohistochemistry, qRT-PCR and western blot. Besides, Z172 and Z183 cells were transfected with siRNA-YAP/TAZ (si-YAP/TAZ) and YAP/TAZ overexpression vector (YAP-5SA). Also, Z172 cells were co-transfected with YAP-5SA and si-TEAD2/4. Subsequently, the stemness characteristics, glycolysis level and malignant transformation of cells in each group were observed by sphere-formation assay, commercial kit, MTT, Transwell, scratch experiment, xenotransplantation and western blot.The expression of YAP, TAZ and TEAD increased significantly in cervical cancer tissue and cell line at the stage of CIN2/3 and SCC. When YAP/TAZ was knocked down, the stemness characteristics, glycolysis level and malignant transformation of cancer cells were notably inhibited; while activating YAP/TAZ exhibited a completely opposite result. In addition, activating YAP/TAZ and knocking down the TEAD expression at the same time significant weakened the effect of activated YAP/TAZ signal on precancerous cells and reduced inhibitory effect of knocking down TEAD alone. YAP/TAZ-TEAD signal activates the characteristics and Warburg effect of cancer stem cells, thereby promoting the malignant transformation of CIN.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Transformação Celular Neoplásica , Células-Tronco Neoplásicas , Transativadores , Fatores de Transcrição , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional , Displasia do Colo do Útero , Neoplasias do Colo do Útero , Proteínas de Sinalização YAP , Humanos , Feminino , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Sinalização YAP/metabolismo , Proteínas de Sinalização YAP/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/patologia , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Displasia do Colo do Útero/patologia , Displasia do Colo do Útero/genética , Displasia do Colo do Útero/metabolismo , Animais , Transativadores/genética , Transativadores/metabolismo , Fatores de Transcrição de Domínio TEA/metabolismo , Linhagem Celular Tumoral , Camundongos , Efeito Warburg em Oncologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proliferação de Células/genética , Camundongos Nus , Regulação Neoplásica da Expressão Gênica , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologiaRESUMO
BACKGROUND: Non-small cell lung cancer (NSCLC) accounts for more than 80 % of lung cancer (LC) cases, making it the primary cause of cancer-related mortality worldwide. T-box transcription factor 5 (TBX5) is an important regulator of embryonic and organ development and plays a key role in cancer development. Here, our objective was to investigate the involvement of TBX5 in ferroptosis within LC cells and the underlying mechanisms. METHODS: First, TBX5 expression was examined in human LC cells. Next, overexpression of TBX5 and Yes1-associated transcriptional regulator (YAP1) and knockdown of TEA domain 1 (TEAD1) were performed in A549 and NCI-H1703 cells. The proliferation ability of A549 and NCI-H1703 cells, GSH, MDA, ROS, and Fe2+ levels were measured. Co-immunoprecipitation (Co-IP) was performed to verify whether TBX5 protein could bind YAP1. Then TBX5, YAP1, TEAD1, GPX4, p53, FTH1, SLC7A11 and PTGS2 protein levels were assessed. Finally, we verified the effect of TBX5 on ferroptosis in LC cells in vivo. RESULTS: TBX5 expression was down-regulated in LC cells, especially in A549 and NCI-H1703 cells. Overexpression of TBX5 significantly decreased proliferation ability of A549 and NCI-H1703 cells, downregulated GPX4 and GSH levels, and upregulated MDA, ROS, and Fe2+ levels. Co-IP verified that TBX5 protein could bind YAP1. Moreover, oe-YAP1 promoted proliferation ability of A549 and NCI-H1703 cells transfected with Lv-TBX5, upregulated GPX4 and GSH levels and downregulated MDA, ROS, and Fe2+ levels. Additionally, oe-YAP1 promoted FTH1 and SLC7A11 levels and inhibited p53 and PTGS2 levels in A549 and NCI-H1703 cells transfected with Lv-TBX5. However, transfection with si-TEAD1 further reversed these effects. In vivo experiments further validated that TBX5 promoted ferroptosis in LC cells. CONCLUSIONS: TBX5 inhibited the activation of YAP1-TEAD1 pathway to promote ferroptosis in LC cells.
Assuntos
Ferroptose , Neoplasias Pulmonares , Proteínas com Domínio T , Fatores de Transcrição de Domínio TEA , Fatores de Transcrição , Proteínas de Sinalização YAP , Ferroptose/genética , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/genética , Proteínas de Sinalização YAP/metabolismo , Proteínas de Sinalização YAP/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição de Domínio TEA/metabolismo , Proteínas com Domínio T/metabolismo , Proteínas com Domínio T/genética , Animais , Linhagem Celular Tumoral , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Camundongos Nus , Proliferação de Células , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Camundongos , Regulação Neoplásica da Expressão Gênica , Células A549 , Transdução de Sinais , Espécies Reativas de Oxigênio/metabolismoRESUMO
Gastric cancer (GC) exhibits significant heterogeneity and its prognosis remains dismal. Therefore, it is essential to investigate new approaches for diagnosing and treating GC. Desmosome proteins are crucial for the advancement and growth of cancer. Plakophilin-2 (PKP2), a member of the desmosome protein family, frequently exhibits aberrant expression and is strongly associated with many tumor types' progression. In this study, we found upregulation of PKP2 in GC. Further correlation analysis showed a notable association between increased PKP2 expression and both tumor stage and poor prognosis in individuals diagnosed with gastric adenocarcinoma. In addition, our research revealed that the Yes-associated protein1 (YAP1)/TEAD4 complex could stimulate the transcriptional expression of PKP2 in GC. Elevated PKP2 levels facilitate activation of the AKT/mammalian target of rapamycin signaling pathway, thereby promoting the malignant progression of GC. By constructing a mouse model, we ultimately validated the molecular mechanism and function of PKP2 in GC. Taken together, these discoveries suggest that PKP2, as a direct gene target of YAP/TEAD4 regulation, has the potential to be used as an indication of GC progression and prognosis. PKP2 is expected to be a promising therapeutic target for GC.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Proteínas de Ligação a DNA , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Proteínas Musculares , Placofilinas , Neoplasias Gástricas , Fatores de Transcrição de Domínio TEA , Fatores de Transcrição , Proteínas de Sinalização YAP , Humanos , Neoplasias Gástricas/patologia , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Animais , Placofilinas/genética , Placofilinas/metabolismo , Fatores de Transcrição de Domínio TEA/metabolismo , Camundongos , Proteínas de Sinalização YAP/metabolismo , Proteínas de Sinalização YAP/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas Musculares/metabolismo , Proteínas Musculares/genética , Prognóstico , Linhagem Celular Tumoral , Masculino , Proliferação de Células , Transdução de Sinais , Feminino , Camundongos Nus , Adenocarcinoma/patologia , Adenocarcinoma/metabolismo , Adenocarcinoma/genéticaRESUMO
In pancreatic ductal adenocarcinomas (PDAC), profound hypoxia plays key roles in regulating cancer cell behavior, including proliferation, migration, and resistance to therapies. The initial part of this research highlights the important role played by long noncoding RNA (lncRNA) MKLN1-AS, which is controlled by hypoxia-inducible factor-1 alpha (HIF-1α), in the progression of PDAC. Human samples of PDAC showed a notable increase in MKLN1-AS expression, which was linked to a worse outcome. Forced expression of MKLN1-AS greatly reduced the inhibitory impact on the growth and spread of PDAC cells caused by HIF-1α depletion. Experiments on mechanisms showed that HIF-1α influences the expression of MKLN1-AS by directly attaching to a hypoxia response element in the promoter region of MKLN1-AS.MKLN1-AS acts as a competitive endogenous RNA (ceRNA) by binding to miR-185-5p, resulting in the regulation of TEAD1 expression and promoting cell proliferation, migration, and tumor growth. TEAD1 subsequently enhances the development of PDAC. Our study results suggest that MKLN1-AS could serve as a promising target for treatment and a valuable indicator for predicting outcomes in PDAC. PDAC is associated with low oxygen levels, and the long non-coding RNA MKLN1-AS interacts with TEAD1 in this context.
Assuntos
Carcinoma Ductal Pancreático , Movimento Celular , Proliferação de Células , Proteínas de Ligação a DNA , Regulação Neoplásica da Expressão Gênica , Subunidade alfa do Fator 1 Induzível por Hipóxia , MicroRNAs , Neoplasias Pancreáticas , RNA Longo não Codificante , Fatores de Transcrição de Domínio TEA , Animais , Humanos , Camundongos , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Camundongos Nus , MicroRNAs/genética , MicroRNAs/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Transdução de Sinais/genética , Fatores de Transcrição de Domínio TEA/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genéticaRESUMO
Schwannoma tumours typically arise on the eighth cranial nerve and are mostly caused by loss of the tumour suppressor Merlin (NF2). There are no approved chemotherapies for these tumours and the surgical removal of the tumour carries a high risk of damage to the eighth or other close cranial nerve tissue. New treatments for schwannoma and other NF2-null tumours such as meningioma are urgently required. Using a combination of human primary tumour cells and mouse models of schwannoma, we have examined the role of the Hippo signalling pathway in driving tumour cell growth. Using both genetic ablation of the Hippo effectors YAP and TAZ as well as novel TEAD palmitoylation inhibitors, we show that Hippo signalling may be successfully targeted in vitro and in vivo to both block and, remarkably, regress schwannoma tumour growth. In particular, successful use of TEAD palmitoylation inhibitors in a preclinical mouse model of schwannoma points to their potential future clinical use. We also identify the cancer stem cell marker aldehyde dehydrogenase 1A1 (ALDH1A1) as a Hippo signalling target, driven by the TAZ protein in human and mouse NF2-null schwannoma cells, as well as in NF2-null meningioma cells, and examine the potential future role of this new target in halting schwannoma and meningioma tumour growth.
Assuntos
Neoplasias Meníngeas , Meningioma , Neurilemoma , Animais , Humanos , Camundongos , Proliferação de Células , Neurilemoma/genética , Neurilemoma/patologia , Neurofibromina 2/genética , Neurofibromina 2/metabolismo , Proteínas de Sinalização YAP/metabolismo , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional/metabolismo , Fatores de Transcrição de Domínio TEA/metabolismoRESUMO
INTRODUCTION: This study attempted to investigate how proprotein convertase subtilisin/kexin type 9 (PCSK9) influences the stemness of stomach adenocarcinoma (STAD) cells. METHODS: CCK-8 and sphere-formation assays were used to detect cell viability and stemness. qRT-PCR and Western blot were used to detect PCSK9 and TEAD4 expression. The binding relationship was verified by dual-luciferase and chromatin immunoprecipitation assays. The effect of TEAD4 activating PCSK9 on the stemness of STAD cells was detected by bioinformatics, BODIPY 493/503, Oil red O, Western blot, and kits. In vivo experiments verified the role of the TEAD4/PCSK9 axis in tumor formation in nude mice. RESULTS: PCSK9 and TEAD4 were highly expressed in STAD. PCSK9 was enriched in the fatty acid metabolism (FAM) pathway. PCSK9 activated the fatty acid metabolism and promoted the proliferation and stemness of STAD cells. TEAD4 as a transcription factor upstream of PCSK9, cell experiments revealed that knockdown of PCSK9 inhibited STAD cell stemness, whereas further addition of fatty acid inhibitors could attenuate the promoting effect on STAD cell stemness brought by STAD overexpression. Rescue experiments showed overexpressed PCSK9 exerted an inhibitory effect on the stemness of STAD cells brought by TEAD4 knockdown. The hypothesis that TEAD4/PCSK9 axis can promote STAD cell growth was confirmed by in vivo experiments. CONCLUSION: Transcription factor TEAD4 could activate PCSK9 to promote the stemness of STAD cells through FAM. These results added weight to the assumption that TEAD4/PCSK9 axis has the potential to be the therapeutic target that inhibits cancer stem cell in STAD.
Assuntos
Adenocarcinoma , Proliferação de Células , Proteínas de Ligação a DNA , Ácidos Graxos , Camundongos Nus , Proteínas Musculares , Células-Tronco Neoplásicas , Pró-Proteína Convertase 9 , Neoplasias Gástricas , Fatores de Transcrição de Domínio TEA , Fatores de Transcrição , Neoplasias Gástricas/patologia , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/genética , Fatores de Transcrição de Domínio TEA/metabolismo , Animais , Humanos , Proteínas Musculares/metabolismo , Proteínas Musculares/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Adenocarcinoma/patologia , Adenocarcinoma/metabolismo , Adenocarcinoma/genética , Linhagem Celular Tumoral , Camundongos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Ácidos Graxos/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Proliferação de Células/genética , Pró-Proteína Convertase 9/metabolismo , Pró-Proteína Convertase 9/genética , Regulação Neoplásica da Expressão GênicaRESUMO
The transcriptional coactivator YAP is emerging as a master regulator of cell growth. In the liver, YAP activity is linked to hepatomegaly, regeneration, dedifferentiation, and aggressive tumor growth. Here we present genomic studies to address how YAP may elicit such profound biological changes in murine models. YAP bound the genome in a TEAD-dependent manner, either at loci constitutively occupied by TEAD or by pioneering enhancers, which comprised a fraction of HNF4a/FOXA-bound embryonic enhancers active during embryonic development but silent in the adult. YAP triggered transcription on promoters by recruiting BRD4, enhancing H3K122 acetylation, and promoting RNApol2 loading and pause-release. YAP also repressed HNF4a target genes by binding to their promoters and enhancers, thus preventing RNApol2 pause-release. YAP activation led to the induction of hepatocyte proliferation, accompanied by tissue remodeling, characterized by polarized macrophages, exhausted T-lymphocytes and dedifferentiation of endothelial cells into proliferative progenitors. Overall, these analyses suggest that YAP is a master regulator of liver function that reshapes the enhancer landscape to control transcription of genes involved in metabolism, proliferation, and inflammation, subverts lineage specification programs by antagonizing HNF4a and modulating the immune infiltrate and the vascular architecture of the liver.
Assuntos
Fígado , Fatores de Transcrição de Domínio TEA , Proteínas de Sinalização YAP , Animais , Células Endoteliais/metabolismo , Elementos Facilitadores Genéticos , Fator 4 Nuclear de Hepatócito/metabolismo , Fígado/metabolismo , Macrófagos , Camundongos , Proteínas Nucleares/metabolismo , Regiões Promotoras Genéticas , RNA Polimerase II/metabolismo , Linfócitos T , Fatores de Transcrição de Domínio TEA/metabolismo , Fatores de Transcrição , Transcrição Gênica , Proteínas de Sinalização YAP/genética , Proteínas de Sinalização YAP/metabolismoRESUMO
UDP-glucose pyrophosphorylase 2 (UGP2), the enzyme that synthesizes uridine diphosphate (UDP)-glucose, rests at the convergence of multiple metabolic pathways, however, the role of UGP2 in tumor maintenance and cancer metabolism remains unclear. Here, we identify an important role for UGP2 in the maintenance of pancreatic ductal adenocarcinoma (PDAC) growth in both in vitro and in vivo tumor models. We found that transcription of UGP2 is directly regulated by the Yes-associated protein 1 (YAP)-TEA domain transcription factor (TEAD) complex, identifying UGP2 as a bona fide YAP target gene. Loss of UGP2 leads to decreased intracellular glycogen levels and defects in N-glycosylation targets that are important for the survival of PDACs, including the epidermal growth factor receptor (EGFR). These critical roles of UGP2 in cancer maintenance, metabolism, and protein glycosylation may offer insights into therapeutic options for otherwise intractable PDACs.
Assuntos
Carcinoma Ductal Pancreático/enzimologia , Regulação Enzimológica da Expressão Gênica/fisiologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Glicogênio/biossíntese , Neoplasias Pancreáticas/enzimologia , UTP-Glucose-1-Fosfato Uridililtransferase/metabolismo , Animais , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Técnicas de Silenciamento de Genes , Glicosilação , Humanos , Camundongos , Camundongos Nus , Neoplasias Experimentais , Neoplasias Pancreáticas/patologia , Fatores de Transcrição de Domínio TEA/genética , Fatores de Transcrição de Domínio TEA/metabolismo , UTP-Glucose-1-Fosfato Uridililtransferase/genética , Proteínas de Sinalização YAP/genética , Proteínas de Sinalização YAP/metabolismoRESUMO
BACKGROUND AND AIMS: Peroxisome proliferator-activated receptor α (PPARα, NR1C1) is a ligand-activated nuclear receptor involved in the regulation of lipid catabolism and energy homeostasis. PPARα activation induces hepatomegaly and plays an important role in liver regeneration, but the underlying mechanisms remain unclear. APPROACH AND RESULTS: In this study, the effect of PPARα activation on liver enlargement and regeneration was investigated in several strains of genetically modified mice. PPARα activation by the specific agonist WY-14643 significantly induced hepatomegaly and accelerated liver regeneration after 70% partial hepatectomy (PHx) in wild-type mice and Pparafl/fl mice, while these effects were abolished in hepatocyte-specific Ppara-deficient (PparaΔHep ) mice. Moreover, PPARα activation promoted hepatocyte hypertrophy around the central vein area and hepatocyte proliferation around the portal vein area. Mechanistically, PPARα activation regulated expression of yes-associated protein (YAP) and its downstream targets (connective tissue growth factor, cysteine-rich angiogenic inducer 61, and ankyrin repeat domain 1) as well as proliferation-related proteins (cyclins A1, D1, and E1). Binding of YAP with the PPARα E domain was critical for the interaction between YAP and PPARα. PPARα activation further induced nuclear translocation of YAP. Disruption of the YAP-transcriptional enhancer factor domain family member (TEAD) association significantly suppressed PPARα-induced hepatomegaly and hepatocyte enlargement and proliferation. In addition, PPARα failed to induce hepatomegaly in adeno-associated virus-Yap short hairpin RNA-treated mice and liver-specific Yap-deficient mice. Blockade of YAP signaling abolished PPARα-induced hepatocyte hypertrophy around the central vein area and hepatocyte proliferation around the portal vein area. CONCLUSIONS: This study revealed a function of PPARα in regulating liver size and liver regeneration through activation of the YAP-TEAD signaling pathway. These findings have implications for understanding the physiological functions of PPARα and suggest its potential for manipulation of liver size and liver regeneration.
Assuntos
Hepatomegalia/genética , Regeneração Hepática/genética , PPAR alfa/metabolismo , Fatores de Transcrição de Domínio TEA/metabolismo , Proteínas de Sinalização YAP/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Hepatectomia/efeitos adversos , Hepatócitos/patologia , Hepatomegalia/patologia , Humanos , Fígado/patologia , Fígado/cirurgia , Regeneração Hepática/efeitos dos fármacos , Masculino , Camundongos , Camundongos Transgênicos , PPAR alfa/agonistas , Pirimidinas/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Proteínas de Sinalização YAP/genéticaRESUMO
TEAD is a transcription factor responsible for the output of the tumor suppressor Hippo pathway. The transcriptional activity of TEAD requires molecular interaction with its transcriptional coactivator, YAP. Aberrant activation of TEAD is deeply involved in tumorigenesis and is associated with poor prognosis, suggesting that inhibitors targeting the YAP-TEAD system are promising as antitumor agents. In this study, we identified NPD689, an analog of the natural product alkaloid emetine, as an inhibitor of the YAP-TEAD interaction. NPD689 suppressed the transcriptional activity of TEAD and reduced the viability of human malignant pleural mesothelioma and non-small cell lung cancer cells but not the viability of normal human mesothelial cells. Our results suggest that NPD689 is not only a new useful chemical tool for elucidating the biological role of the YAP-TEAD system but also has potential as a starting compound for developing a cancer therapeutic agent that targets the YAP-TEAD interaction.
Assuntos
Antineoplásicos , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Antineoplásicos/farmacologia , Emetina , Neoplasias Pulmonares/patologia , Fatores de Transcrição/metabolismo , Proteínas de Sinalização YAP , Fatores de Transcrição de Domínio TEA/metabolismoRESUMO
The molecular mechanisms that drive the granulosa cells' (GC) differentiation into a more estrogenic phenotype during follicular divergence and establishment of follicle dominance have not been completely elucidated. The main Hippo signaling effector, YAP, has, however, emerged as a potential key player to explain such complex processes. Studies using rat and bovine GC demonstrate that, in conditions where the expression of the classic YAP-TEAD target gene tissue growth factor (CTGF) is augmented, CYP19A1 expression and activity and, consequently, estradiol (E2) secretion are reduced. These findings led us to hypothesize that, during ovarian follicular divergence in cattle, FSH downregulates YAP-TEAD-dependent transcriptional activity in GC to allow the future dominant follicle to exert its augmented estrogenic capacity. To address this, we performed a series of experiments employing distinct bovine models. Our in vitro and ex vivo experiments indicated that indeed FSH downregulates, in a concentration-dependent manner, mRNA levels not only for CTGF but also for the other classic YAP-TEAD transcriptional target genes ANKRD1 and CYR61 by a mechanism that involves increased YAP phosphorylation. To better elucidate the functional importance of such FSH-induced YAP activity regulation, we then cultured GC in the presence of verteporfin (VP) or peptide 17 (P17), two pharmacological inhibitors known to interfere with YAP binding to TEADs. The results showed that both VP and P17 increased CYP19A1 basal mRNA levels in a concentration-dependent manner. Most interestingly, by using GC samples obtained in vivo from dominant vs. subordinate follicles, we found that mRNA levels for CTGF, CYR61, and ANKRD1 are higher in subordinate follicles following the follicular divergence. Taken together, our novel results demonstrate that YAP transcriptional activity is regulated in bovine granulosa cells to allow the increased estrogenic capacity of the selected dominant follicle.
Assuntos
Hormônio Foliculoestimulante , Folículo Ovariano , Animais , Bovinos/genética , Bovinos/metabolismo , Feminino , Ratos , Estrona/metabolismo , Hormônio Foliculoestimulante/farmacologia , Hormônio Foliculoestimulante/metabolismo , Células da Granulosa/metabolismo , Folículo Ovariano/metabolismo , RNA Mensageiro/metabolismo , Verteporfina , Fatores de Transcrição de Domínio TEA/metabolismo , Proteínas de Sinalização YAP/metabolismoRESUMO
We have previously demonstrated that the transcription co-factor yes-associated protein 1 (YAP1) promotes vascular smooth muscle cell (VSMC) de-differentiation. Yet, the role and underlying mechanisms of YAP1 in neointima formation in vivo remain unclear. The goal of this study was to investigate the role of VSMC-expressed YAP1 in vascular injury-induced VSMC proliferation and delineate the mechanisms underlying its action. Experiments employing gain- or loss-of-function of YAP1 demonstrated that YAP1 promotes human VSMC proliferation. Mechanistically, we identified platelet-derived growth factor receptor beta (PDGFRB) as a novel YAP1 target gene that confers the YAP1-dependent hyper-proliferative effects in VSMCs. Furthermore, we identified TEA domain transcription factor 1 (TEAD1) as a key transcription factor that mediates YAP1-dependent PDGFRß expression. ChIP assays demonstrated that TEAD1 is enriched at a PDGFRB gene enhancer. Luciferase reporter assays further demonstrated that YAP1 and TEAD1 co-operatively activate the PDGFRB enhancer. Consistent with these observations, we found that YAP1 expression is upregulated after arterial injury and correlates with PDGFRß expression and VSMC proliferation in vivo. Using a novel inducible SM-specific Yap1 knockout mouse model, we found that the specific deletion of Yap1 in adult VSMCs is sufficient to attenuate arterial injury-induced neointima formation, largely due to inhibited PDGFRß expression and VSMC proliferation. Our study unravels a novel mechanism by which YAP1/TEAD1 promote VSMC proliferation via transcriptional induction of PDGFRß, thereby enhancing PDGF-BB downstream signaling and promoting neointima formation.
Assuntos
Regulação da Expressão Gênica , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Neointima/metabolismo , Receptor beta de Fator de Crescimento Derivado de Plaquetas/genética , Fatores de Transcrição de Domínio TEA/genética , Proteínas de Sinalização YAP/genética , Animais , Becaplermina/metabolismo , Proliferação de Células , Elementos Facilitadores Genéticos , Feminino , Camundongos , Modelos Biológicos , Regiões Promotoras Genéticas , Ligação Proteica , Receptor beta de Fator de Crescimento Derivado de Plaquetas/metabolismo , Transdução de Sinais , Fatores de Transcrição de Domínio TEA/metabolismo , Ativação Transcricional , Proteínas de Sinalização YAP/metabolismoRESUMO
Re-activation of fetal haemoglobin (HbF) has been proved to be an effective strategy for the treatment of ß-haemoglobinopathies. In this study, we identified TEA domain transcription factor 4 (TEAD4) as a new potential regulator of HbF by integrating public data sets with quantitative polymerase chain reaction analysis in ß-thalassaemia patients. Significant negative correlation was observed between the expression of TEAD4 and HbF levels in ß-thalassaemia patients. Functional validations of TEAD4 inhibition in both ß-thalassaemia CD34+ cells and HUDEP-2 cells indicated that depletion of TEAD4 led to a significant increase of HbF. Finally, we identified a binding motif of TEAD4 on γ-globin gene promoters; its disruption consistently led to de-repression of HbF. Taken together, these results demonstrate that TEAD4 could act as a transcriptional inhibitor of the γ-globin gene through direct binding on its promoter. Our findings demonstrate a novel role of TEAD4 on the regulation of HbF, which may benefit patients with ß-haemoglobinopathies.
Assuntos
Hemoglobina Fetal/genética , Regiões Promotoras Genéticas , Fatores de Transcrição de Domínio TEA/metabolismo , gama-Globinas/genética , Linhagem Celular , Células Eritroides/metabolismo , Regulação da Expressão Gênica , Humanos , Ligação Proteica , Talassemia beta/genética , Talassemia beta/metabolismoRESUMO
BACKGROUND: Frizzled (FZD) proteins function as receptors for WNT ligands. Members in FZD family including FZD2, FZD4, FZD7, FZD8 and FZD10 have been demonstrated to mediate cancer cell epithelial-mesenchymal transition (EMT). METHODS: CCLE and TCGA databases were interrogated to reveal the association of FZD5 with EMT. EMT was analyzed by investigating the alterations in CDH1 (E-cadherin), VIM (Vimentin) and ZEB1 expression, cell migration and cell morphology. Transcriptional modulation was determined by ChIP in combination with Real-time PCR. Survival was analyzed by Kaplan-Meier method. RESULTS: In contrast to other FZDs, FZD5 was identified to prevent EMT in gastric cancer. FZD5 maintains epithelial-like phenotype and is negatively modulated by transcription factors SNAI2 and TEAD1. Epithelial-specific factor ELF3 is a downstream effecter, and protein kinase C (PKC) links FZD5 to ELF3. ELF3 represses ZEB1 expression, further guarding against EMT. Moreover, FZD5 signaling requires its co-receptor LRP5 and WNT7B is a putative ligand for FZD5. FZD5 and ELF3 are associated with longer survival, whereas SNAI2 and TEAD1 are associated with shorter survival. CONCLUSIONS: Taken together, FZD5-ELF3 signaling blocks EMT, and plays a potential tumor-suppressing role in gastric cancer. Video Abstract.