RESUMEN
The tumor suppressor p53 prevents cancer development by regulating dozens of target genes with diverse biological functions. Although numerous p53 target genes have been identified to date, the dynamics and function of the regulatory network centered on p53 have not yet been fully elucidated. We herein identified inhibitor of DNA-binding/differentiation-3 (ID3) as a direct p53 target gene. p53 bound the distal promoter of ID3 and positively regulated its transcription. ID3 expression was significantly decreased in clinical lung cancer tissues, and was closely associated with overall survival outcomes in these patients. Functionally, ID3 deficiency promoted the metastatic ability of lung cancer cells through its effects on the transcriptional regulation of CDH1. Furthermore, the ectopic expression of ID3 in p53-knockdown cells restored E-cadherin expression. Collectively, the present results demonstrate that ID3 plays a tumor-suppressive role as a downstream effector of p53 and impedes lung cancer cell metastasis by regulating E-cadherin expression.
Asunto(s)
Neoplasias Pulmonares , Humanos , Cadherinas/genética , Cadherinas/metabolismo , Línea Celular Tumoral , Regulación de la Expresión Génica , Proteínas Inhibidoras de la Diferenciación/genética , Proteínas Inhibidoras de la Diferenciación/metabolismo , Neoplasias Pulmonares/patología , Proteínas de Neoplasias/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Su(var)3-9, Enhancer-of-zeste, and Trithorax (SET) domain-containing protein 8 (SET8) is the sole enzyme that monomethylates Lys-20 of histone H4 (H4K20). SET8 has been implicated in the regulation of multiple biological processes, such as gene transcription, the cell cycle, and senescence. SET8 quickly undergoes ubiquitination and degradation by several E3 ubiquitin ligases; however, the enzyme that deubiquitinates SET8 has not yet been identified. Here we demonstrated that ubiquitin-specific peptidase 17-like family member (USP17) deubiquitinates and therefore stabilizes the SET8 protein. We observed that USP17 interacts with SET8 and removes polyubiquitin chains from SET8. USP17 knockdown not only decreased SET8 protein levels and H4K20 monomethylation but also increased the levels of the cyclin-dependent kinase inhibitor p21. As a consequence, USP17 knockdown suppressed cell proliferation. We noted that USP17 was down-regulated in replicative senescence and that USP17 inhibition alone was sufficient to trigger cellular senescence. These results reveal a regulatory mechanism whereby USP17 prevents cellular senescence by removing ubiquitin marks from and stabilizing SET8 and transcriptionally repressing p21.
Asunto(s)
Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Endopeptidasas/metabolismo , N-Metiltransferasa de Histona-Lisina/metabolismo , Envejecimiento/metabolismo , Animales , Células COS , Ciclo Celular/fisiología , Línea Celular , Proliferación Celular/fisiología , Chlorocebus aethiops , Células HCT116 , Histonas/metabolismo , Humanos , Células MCF-7 , Metiltransferasas/metabolismo , Procesamiento Proteico-Postraduccional , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación/fisiologíaRESUMEN
In response to cellular stresses, activating transcriptional factor 4 (ATF4) regulates the expression of both stress-relieving genes and apoptosis-inducing genes, eliciting cell fate determination. Since pharmacological activation of ATF4 exerts potent anti-tumor effects, modulators of ATF4 activation may have potential in cancer therapy. We herein attempted to identify small molecules that activate ATF4. A cell-based screening to monitor TRB3 promoter activation was performed using crude drugs used in traditional Japanese Kampo medicine. We found that an extract from Sophora flavescens roots exhibited potent TRB3 promoter activation. The activity-guided fractionation revealed that kurarinone was identified as the active ingredient. Intriguingly, ATF4 activation in response to kurarinone required PKR-like endoplasmic reticulum kinase (PERK). Moreover, kurarinone induced the cyclin-dependent kinase inhibitor p21 as well as cytostasis in cancer cells. Importantly, the cytostatic effect of kurarinone was reduced by pharmacological inhibition of PERK. These results indicate that kurarinone triggers ATF4 activation through PERK and exerts cytostatic effects on cancer cells. Taken together, our results suggest that modulation of the PERK-ATF4 pathway with kurarinone has potential as a cancer treatment.
Asunto(s)
Factor de Transcripción Activador 4/metabolismo , Proteínas de Ciclo Celular/genética , Flavonoides/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Represoras/genética , Sophora/química , eIF-2 Quinasa/metabolismo , Factor de Transcripción Activador 4/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HEK293 , Células HeLa , Humanos , Fosforilación , Regiones Promotoras Genéticas/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/genética , eIF-2 Quinasa/genéticaRESUMEN
BACKGROUND: Plasma low-density lipoprotein (LDL) cholesterol is implicated in abdominal aorta (AA) and aortic dissection (AD); however, its role in the pathogenesis of AA and AD, a disease with a high mortality rate, is unknown. The existing animal models such as apolipoprotein E-deficient (Apoe-/-) mice cannot reproduce all the conditions of AA/AD, including elevated LDL-cholesterol levels and spontaneous atheroma formation; therefore, a more reliable in vivo model is required. Here, we analyzed angiotensin II (Ang II)-induced mice with combined deficiency of the LDL receptor and the catalytic component of the apolipoprotein B-edisome complex (Ldlr-/-/Apobec1-/- [WKO]) to understand AA formation and AD occurrence in relation to plasma lipid composition. METHODS: AAs and ADs were created in 18- to 22- week-old male Apoe-/- and Ldlr-/-/Apobec1-/- mice by Ang II infusion. Immunostaining allowed assessment of smooth muscle cells and mural monocytes/macrophages. RESULTS: Ldlr-/-/Apobec1-/- mice had elevated LDL-cholesterol levels characteristic for human type IIa hyperlipidemia, resulting in atherogenesis, which promoted mortality, AA formation, and AD development. Interestingly, variations in the distribution of atheromas and inflammatory sites between Apoe-/- and Ldlr-/-/Apobec1-/- mice depending on lipid profiles resulted in differences in AA formation and AD occurrence in the thoracic aorta. CONCLUSIONS: Our results indicate the presence of a pathogenic pathway involving serum lipid composition that plays a key role in AA formation and AD occurrence in Ang II-induced mice.
Asunto(s)
Angiotensina II , Aorta Torácica/metabolismo , Aneurisma de la Aorta Torácica/inducido químicamente , LDL-Colesterol/sangre , Hipercolesterolemia/sangre , Desaminasas APOBEC-1/deficiencia , Desaminasas APOBEC-1/genética , Animales , Aorta Torácica/patología , Aneurisma de la Aorta Torácica/sangre , Aneurisma de la Aorta Torácica/patología , Biomarcadores/sangre , Modelos Animales de Enfermedad , Predisposición Genética a la Enfermedad , Hipercolesterolemia/complicaciones , Hipercolesterolemia/genética , Masculino , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados para ApoE , Fenotipo , Placa Aterosclerótica , Receptores de LDL/deficiencia , Receptores de LDL/genética , Factores de Tiempo , Regulación hacia ArribaRESUMEN
Tribbles 1 (TRB1) is one of the mammalian orthologs of Drosophila Tribbles, which regulates development and cell proliferation. TRB1 is suggested to act as a scaffold protein in signaling pathways for important cellular processes. TRB1 has also been identified as a myeloid oncogenic driver and mediates leukemogenesis through the mitogen-activated protein extracellular kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway and CCAAT/enhancer binding protein (C/EBP) transcriptional factors. However, the physiological roles of TRB1 in solid tumors have not been clarified. Here, we show that TRB1 interacts with p53 and suppresses its tumor suppressor activity. TRB1 knockdown enhances transcriptional activity of p53 and decreases cell viability. Interestingly, TRB1 enhances histone deacety lase 1 (HDAC1)-mediated p53 deacetylation and decreases DNA binding of p53. These results suggest that TRB1 is involved in the proliferation of tumor cells by inhibiting the activities of tumor suppressor p53 in solid tumors.
Asunto(s)
Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/metabolismo , Línea Celular Tumoral , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , ADN/metabolismo , Técnicas de Silenciamiento del Gen , Histona Desacetilasa 1/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Interferente Pequeño/genéticaRESUMEN
Tribbles 1 (TRB1), a member of the Tribbles family, is a pseudokinase that is conserved among species and implicated in various human diseases including leukemia, cardiovascular diseases, and metabolic disorders. However, the role of TRB1 in the immune response is not understood. To evaluate this role, we examined regulation of TRB1 expression and the function of TRB1 in interleukin-2 (IL-2) induction in Jurkat cells, a human acute T cell leukemia cell line. We found that TRB1 was strongly induced by phorbol 12-myristate 13-acetate (PMA) and ionomycin in these cells. IL-2 expression was induced in Jurkat cells activated by PMA and ionomycin; however, knockdown of TRB1 resulted in decreased induction of IL-2. TRB1 null Jurkat cells established using the CRISPR/Cas9 system also showed reduction of IL-2 expression on PMA/ionomycin stimulation. TRB1 knockdown also markedly inhibited IL-2 promoter activation. To determine the mechanism of the stimulatory effect on IL-2 induction, we focused on histone deacetylases (HDACs), and found that HDAC1 preferentially interacts with TRB1. TRB1 suppressed the interaction of HDAC1 with nuclear factor of activated T cells 2 (NFAT2), which is a crucial transcription factor for IL-2 induction. These results indicate that TRB1 is a positive regulator of IL-2 induction in activated T cells.
Asunto(s)
Histona Desacetilasa 1/metabolismo , Interleucina-2/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Activación de Linfocitos/fisiología , Factores de Transcripción NFATC/metabolismo , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Linfocitos T/metabolismo , Ionóforos de Calcio/farmacología , Carcinógenos/farmacología , Humanos , Ionomicina/farmacología , Células Jurkat , Leucemia de Células T/metabolismo , Regiones Promotoras Genéticas , Proteínas Serina-Treonina Quinasas/metabolismo , Acetato de Tetradecanoilforbol/farmacologíaRESUMEN
Cardiac glycosides (CGs) have been used for decades to treat heart failure and arrhythmic diseases. Recent non-clinical and epidemiological findings have suggested that CGs exhibit anti-tumor activities. Therefore, CGs may be repositioned as drugs for the treatment of cancer. A detailed understanding of the anti-cancer mechanisms of CGs is essential for their application to the treatment of targetable cancer types. To elucidate the factors associated with the anti-tumor effects of CGs, we performed transcriptome profiling on human multiple myeloma AMO1 cells treated with periplocin, one of the CGs. Periplocin significantly down-regulated the transcription of MYC (c-Myc), a well-established oncogene. Periplocin also suppressed c-Myc expression at the protein levels. This repression of c-Myc was also observed in several cell lines. To identify target proteins for the inhibition of c-Myc, we generated CG-resistant (C9) cells using a sustained treatment with digoxin. We confirmed that C9 cells acquired resistance to the inhibition of c-Myc expression and cell proliferation by CGs. Moreover, the sequencing of genomic DNA in C9 cells revealed the mutation of D128N in α1-Na/K-ATPase, indicating the target protein. These results suggest that CGs suppress c-Myc expression in cancer cells via α1-Na/K-ATPase, which provides further support for the anti-tumor activities of CGs.
Asunto(s)
Glicósidos Cardíacos , Humanos , Glicósidos Cardíacos/farmacología , Línea Celular , Proliferación Celular , Perfilación de la Expresión Génica , Adenosina TrifosfatasasRESUMEN
Transforming growth factor ß (TGF-ß) is a multifunctional cytokine that induces a diverse set of cellular processes principally through Smad-dependent transcription. Transcriptional responses induced by Smads are tightly regulated by Smad cofactors and histone modifications; however, the underlying mechanisms have not yet been elucidated in detail. We herein report lysine methyltransferase SET8 as a negative regulator of TGF-ß signaling. SET8 physically associates with Smad2/3 and negatively affects transcriptional activation by TGF-ß in a catalytic activity-independent manner. The depletion of SET8 results in an increase in TGF-ß-induced plasminogen activator inhibitor-1 (PAI-1) and p21 expression and enhances the antiproliferative effects of TGF-ß. Mechanistically, SET8 occupies the PAI-1 and p21 promoters, and a treatment with TGF-ß triggers the replacement of the suppressive binding of SET8 with p300 on these promoters, possibly to promote gene transcription. Collectively, the present results reveal a novel role for SET8 in the negative regulation of TGF-ß signaling.
Asunto(s)
Inhibidor 1 de Activador Plasminogénico , Factor de Crecimiento Transformador beta , Factor de Crecimiento Transformador beta/metabolismo , Inhibidor 1 de Activador Plasminogénico/genética , Inhibidor 1 de Activador Plasminogénico/metabolismo , Transducción de Señal/fisiología , Activación Transcripcional , Metilación , Proteína Smad2/genética , Proteína Smad2/metabolismoRESUMEN
Transcriptional coactivator with PDZ-binding motif (TAZ) is a downstream transcriptional regulator of the Hippo pathway that controls cell growth and differentiation. The aberrant activation of TAZ correlates with a poor prognosis in human cancers, such as breast and colon cancers. We previously demonstrated that TAZ inhibited the tumor suppressor functions of p53 and enhanced cell proliferation. Statins, which are used to treat dyslipidemia, have been reported to suppress the activity of TAZ and exert anti-tumor effects. In the present study, we focused on the regulation of p53 functions by TAZ and investigated whether statins modulate these functions via TAZ. The results obtained suggest that statins, such as simvastatin and fluvastatin, activated the transcriptional function of p53 by suppressing TAZ protein expression. Furthermore, co-treatment with simvastatin and anti-tumor agents that cooperatively activate p53 suppressed cancer cell survival. These results indicate a useful mechanism by which statins enhance the effects of anti-tumor agents through the activation of p53 and may represent a novel approach to cancer therapy.
RESUMEN
The tumor suppressor p53 is a transcription factor that regulates the expression of dozens of target genes and diverse physiological processes. To precisely regulate the p53 network, p53 undergoes various post-translational modifications and alters the selectivity of target genes. Acetylation plays an essential role in cell fate determination through the activation of p53. Although the acetylation of p53 has been examined, the underlying regulatory mechanisms remain unclear and, thus, have attracted the interest of researchers. We herein discuss the role of acetylation in the p53 pathway, with a focus on p53 acetyltransferases and deacetylases. We also review recent findings on the regulators of these enzymes to understand the mode of p53 acetylation from a broader perspective.
Asunto(s)
Procesamiento Proteico-Postraduccional , Proteína p53 Supresora de Tumor , Proteína p53 Supresora de Tumor/metabolismo , Acetilación , Acetiltransferasas/metabolismo , Factores de Transcripción/metabolismoRESUMEN
The c-Myc oncoprotein is frequently overexpressed in human cancers and is essential for cancer cell proliferation. The dysregulation of ubiquitin-proteasome-mediated degradation is one of the contributing factors to the upregulated expression of c-Myc in human cancers. We herein identified USP17 as a novel deubiquitinating enzyme that regulates c-Myc levels and controls cell proliferation and glycolysis. The overexpression of USP17 stabilized the c-Myc protein by promoting its deubiquitination. In contrast, the knockdown of USP17 promoted c-Myc degradation and reduced c-Myc levels. The knockdown of USP17 also suppressed cell proliferation and glycolysis. Collectively, the present results reveal a novel role for USP17 in the regulation of c-Myc stability and suggest its potential as a therapeutic target for cancer treatment.
Asunto(s)
Endopeptidasas/genética , Glucólisis/genética , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Animales , Células COS , Línea Celular Tumoral , Proliferación Celular/genética , Chlorocebus aethiops , Endopeptidasas/metabolismo , Células Epiteliales/metabolismo , Células Epiteliales/patología , Regulación Neoplásica de la Expresión Génica , Glucosa/metabolismo , Humanos , Ácido Láctico/metabolismo , Proteolisis , Proteínas Proto-Oncogénicas c-myc/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Transducción de SeñalRESUMEN
The unfolded protein response (UPR) controls protein homeostasis through transcriptional and translational regulation. However, dysregulated UPR signaling has been associated with the pathogenesis of many human diseases. Therefore, the compounds modulating UPR may provide molecular insights for these pathologies in the context of UPR. Here, we screened small-molecule compounds that suppress UPR, using a library of Myanmar wild plant extracts. The screening system to track X-box binding protein 1 (XBP1) splicing activity revealed that the ethanol extract of the Periploca calophylla stem inhibited the inositol-requiring enzyme 1 (IRE1)-XBP1 pathway. We isolated and identified periplocin as a potent inhibitor of the IRE1-XBP1 axis. Periplocin also suppressed other UPR axes, protein kinase R-like endoplasmic reticulum kinase (PERK), and activating transcription factor 6 (ATF6). Examining the structure-activity relationship of periplocin revealed that cardiac glycosides also inhibited UPR. Moreover, periplocin suppressed the constitutive activation of XBP1 and exerted cytotoxic effects in the human multiple myeloma cell lines, AMO1 and RPMI8226. These results reveal a novel suppressive effect of periplocin or the other cardiac glycosides on UPR regulation, suggesting that these compounds will contribute to our understanding of the pathological or physiological importance of UPR.
Asunto(s)
Glicósidos Cardíacos/farmacología , Saponinas/farmacología , Respuesta de Proteína Desplegada/efectos de los fármacos , Línea Celular , Línea Celular Tumoral , Retículo Endoplásmico/efectos de los fármacos , Retículo Endoplásmico/metabolismo , Células HEK293 , Humanos , Periploca/química , Extractos Vegetales/farmacología , Empalme del ARN/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Proteína 1 de Unión a la X-Box/metabolismoRESUMEN
Transcriptional coactivator with a PDZ-binding motif (TAZ) is one of the mammalian orthologs of Drosophila Yorkie, a transcriptional coactivator of the Hippo pathway. TAZ has been suggested to function as a regulator that modulates the expression of cell proliferation and anti-apoptotic genes in order to stimulate cell proliferation. TAZ has also been associated with a poor prognosis in several cancers, including breast cancer. However, the physiological role of TAZ in tumorigenesis remains unclear. We herein demonstrated that TAZ negatively regulated the activity of the tumor suppressor p53. The overexpression of TAZ down-regulated p53 transcriptional activity and its downstream gene expression. In contrast, TAZ knockdown up-regulated p21 expression induced by p53 activation. Regarding the underlying mechanism, TAZ inhibited the interaction between p53 and p300 and suppressed the p300-mediated acetylation of p53. Furthermore, TAZ knockdown induced cellular senescence in a p53-dependent manner. These results suggest that TAZ negatively regulates the tumor suppressor functions of p53 and attenuates p53-mediated cellular senescence.
Asunto(s)
Senescencia Celular , Transactivadores/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Acetilación , Línea Celular , ADN/metabolismo , Proteína p300 Asociada a E1A/metabolismo , Fibroblastos/metabolismo , Humanos , Modelos Biológicos , Unión Proteica , Transcripción Genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZ , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Kurarinone, a flavonoid isolated from the roots of Sophora flavescens, was suggested to exert potent antioxidant and immunosuppressive effects. However, the underlying mechanisms remain unclear. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key transcription factor that regulates the antioxidant defense system with anti-inflammatory activity. In the present study, we demonstrated that kurarinone activated Nrf2 and increased the expression of antioxidant enzymes, including heme oxygenase-1 (HO-1). Mechanistically, kurarinone downregulated the expression of kelch-like ECH-associated protein 1 (KEAP1), subsequently leading to the activation of Nrf2. Kurarinone also inhibited the expression of the inflammatory cytokine, interleukin (IL)-1ß, and inducible nitric oxide synthase (iNos) in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. The overexpression of HO-1 suppressed the LPS-induced production of inflammatory mediators in RAW264.7 cells, and the immunosuppressive effects of kurarinone were partially inhibited by a treatment with Tin Protomorphyrin IX (TinPPIX), an inhibitor of HO-1. These results indicate that kurarinone activates the KEAP1/Nrf2 pathway to induce HO-1 expression, thereby exerting immunosuppressive effects.
RESUMEN
A murine genetic model of LDL-cholesterol- (LDL-C-) driven atherosclerosis, based on complete deficiencies of both the LDL-receptor (Ldlr-/-) and key catalytic component of an apolipoprotein B-edisome complex (Apobec1-/-), which converts apoB-100 to apoB-48, has been extensively characterized. These gene deficiencies allow high levels of apoB-100 to be present and inefficiently cleared, thus leading to very high levels of LDL-C in mice on a normal diet. Many key features of atherosclerotic plaques observed in human familial hypercholesterolemia are found in these mice as they are allowed to age through 72 weeks. The general characteristics include the presence of high levels of LDL-C in plasma and macrophage-related fatty streak formation in the aortic tree, which progressively worsens with age. More specifically, plaque found in the aortic sinuses contains a lipid core with relatively high numbers of macrophages and a smooth muscle cell α-actin- and collagen-containing cap, which thins with age. These critical features of plaque progression suggest that the Ldlr-/-/Apobec1-/- mouse line presents a superior model of LDL-C-driven atherosclerosis.
Asunto(s)
Aterosclerosis/metabolismo , Modelos Animales de Enfermedad , Hipercolesterolemia/complicaciones , Desaminasas APOBEC-1/genética , Animales , Aterosclerosis/etiología , Humanos , Hiperlipoproteinemia Tipo II , Masculino , Ratones , Ratones Noqueados , Receptores de LDLRESUMEN
Tribbles-related protein (TRB) family members are the mammalian orthologs of Drosophila tribbles. Tribbles was originally identified as a cell cycle regulator during Drosophila development. Tribbles genes are evolutionary conserved, and three TRB genes (TRB1, TRB2 and TRB3) have been identified in mammals. TRBs are considered pseudokinases because they lack an ATP binding site or one of the conserved catalytic motifs essential for kinase activity. Instead, TRBs play important roles in various cellular processes as scaffolds or adaptors to promote the degradation of target proteins and to regulate several key signaling pathways. Recent research has focused on the role of TRBs in tumorigenesis and neoplastic progression. In this review, we focus on the physiological roles of TRB family members in tumorigenesis through the regulation of the ubiquitin-proteasome system and discuss TRBs as biomarkers or potential therapeutic targets in cancer.