RESUMEN
Bioorthogonal catalysis (BC) generates chemical reactions not present in normal physiology for the purpose of disease treatment. Because BC catalytically produces the desired therapy only at the site of disease, it holds the promise of site-specific treatment with little or no systemic exposure or side effects. Transition metals are typically used as catalytic centers in BC; however, solubility and substrate specificity typically necessitate a coordinating enzyme and/or stabilizing superstructure for in vivo application. We report the use of self-assembling, porous exoshells (tESs) to encapsulate and deliver an iron-containing reaction center for the treatment of breast cancer. The catalytic center is paired with indole-3-acetic acid (IAA), a natural product found in edible plants, which undergoes oxidative decarboxylation, via reduction of iron(III) to iron(II), to produce free radicals and bioactive metabolites. The tES encapsulation is critical for endocytic uptake of BC reaction centers and, when followed by administration of IAA, results in apoptosis of MDA-MB-231 triple negative cancer cells and complete regression of in vivo orthotopic xenograft tumors (p < 0.001, n = 8 per group). When Renilla luciferase (rLuc) is substituted for horseradish peroxidase (HRP), whole animal luminometry can be used to monitor in vivo activity.
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Productos Biológicos , Nanopartículas , Neoplasias , Animales , Humanos , Compuestos Férricos , Peroxidasa de Rábano Silvestre/metabolismo , Catálisis , HierroRESUMEN
WW-domain-binding protein 2 (WBP2) is an oncogene that drives breast carcinogenesis through regulating Wnt, estrogen receptor (ER), and Hippo signaling. Recent studies have identified neoteric modes of action of WBP2 other than its widely recognized function as a transcriptional coactivator. Here, we identified a previously unexplored role of WBP2 in inflammatory signaling in breast cancer via an integrated proteogenomic analysis of The Cancer Genome Atlas Breast Invasive Carcinoma (TCGA BRCA) dataset. WBP2 was shown to enhance the migration and invasion in triple-negative breast cancer (TNBC) cells especially under tumor necrosis factor alpha (TNF-α) stimulation. Molecularly, WBP2 potentiates TNF-α-induced nuclear factor kappa B (NF-κB) transcriptional activity and nuclear localization through aggrandizing ubiquitin-mediated proteasomal degradation of its upstream inhibitor, NF-κB inhibitor alpha (NFKBIA; also known as IκBα). We further demonstrate that WBP2 induces mRNA stability of beta-transducin repeat-containing E3 ubiquitin protein ligase (BTRC), which targets IκBα for ubiquitination and degradation. Disruption of IκBα rescued the impaired migratory and invasive phenotypes in WBP2-silenced cells, while loss of BTRC ameliorated WBP2-driven migration and invasion. Clinically, the WBP2-BTRC-IκBα signaling axis correlates with poorer prognosis in breast cancer patients. Our findings reveal a pivotal mechanism of WBP2 in modulating BTRC-IκBα-NF-κB pathway to promote TNBC aggressiveness.
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FN-kappa B/metabolismo , Invasividad Neoplásica , Metástasis de la Neoplasia , ARN Mensajero/genética , Transactivadores/fisiología , Neoplasias de la Mama Triple Negativas/patología , Proteínas con Repetición de beta-Transducina/genética , Línea Celular Tumoral , Variaciones en el Número de Copia de ADN , Femenino , Humanos , Inflamación/metabolismo , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/metabolismoRESUMEN
Cancer is a global health problem. The delineation of molecular mechanisms pertinent to cancer initiation and development has spurred cancer therapy in the form of precision medicine. The Hippo signalling pathway is a tumour suppressor pathway implicated in a multitude of cancers. Elucidation of the Hippo pathway has revealed an increasing number of regulators that are implicated, some being potential therapeutic targets for cancer interventions. WW domain-binding protein 2 (WBP2) is an oncogenic transcriptional co-factor that interacts, amongst others, with two other transcriptional co-activators, YAP and TAZ, in the Hippo pathway. WBP2 was recently discovered to modulate the upstream Hippo signalling components by associating with LATS2 and WWC3. Exacerbating the complexity of the WBP2/Hippo network, WBP2 itself is reciprocally regulated by Hippo-mediated microRNA biogenesis, contributing to a positive feedback loop that further drives carcinogenesis. Here, we summarise the biological mechanisms of WBP2/Hippo reciprocal regulation and propose therapeutic strategies to overcome Hippo defects in cancers through targeting WBP2.
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Vía de Señalización Hippo , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Transactivadores/metabolismo , Variaciones en el Número de Copia de ADN/genética , Humanos , Modelos Biológicos , Medicina de PrecisiónRESUMEN
WBP2 is an emerging oncoprotein with diverse functions in breast tumorigenesis via regulating Wnt, epidermal growth factor receptor, estrogen receptor, and Hippo. Recently, evidence shows that WBP2 is tightly regulated by the components of the miRNA biogenesis machinery such as DGCR8 and Dicer via producing both WBP2's 3'UTR and coding DNA sequence-targeting miRNAs. This led us to hypothesize that WBP2 could provide a feedback loop to the biogenesis of its key upstream regulators by regulating the microprocessor complex activity. Indeed, WBP2 suppressed microprocessor activity by blocking the processing of pri-miRNAs to pre-miRNAs. WBP2 negatively regulated the assembly of the microprocessor complex via physical interactions with its components. Meta-analyses suggest that microprocessor complex components, in particular DGCR8, DDX5, and DEAD-Box Helicase17 (DDX17), have tumor-suppressive properties. 2D and 3D in vitro proliferation assays revealed that WBP2 blocked the tumor-suppressive properties of DGCR8, a key component of the microprocessor complex. In conclusion, WBP2 is a novel regulator of miRNA biogenesis that is a known dysregulated pathway in breast tumorigenesis. The reregulation of miRNA biogenesis machinery via targeting WBP2 protein may have implications in breast cancer therapy.
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Neoplasias de la Mama/genética , MicroARNs/biosíntesis , Transactivadores/metabolismo , Neoplasias de la Mama/metabolismo , Carcinogénesis/genética , Línea Celular Tumoral , ARN Helicasas DEAD-box/metabolismo , Femenino , Humanos , MicroARNs/antagonistas & inhibidores , Proteínas de Unión al ARN/metabolismo , Ribonucleasa III/metabolismo , Transactivadores/fisiologíaRESUMEN
Dysregulation of signaling pathways is responsible for many human diseases. The lack of understanding of the molecular etiology of gastric cancer (GC) poses a substantial challenge to the development of effective cancer therapy. To better understand the molecular mechanisms underlying the pathogenesis of GC, which will facilitate the identification and development of effective therapeutic approaches to improve patient outcomes, mass spectrometry-based phosphoproteomics analysis was performed to map the global molecular changes in GC. A total of 530 proteins with altered phosphorylation levels were detected across a panel of 15 normal and GC cell lines. WW domain-binding protein 2 (WBP2) was validated to be upregulated in a subset of GC cell lines. WBP2 is overexpressed in 61% cases of GC compared to non-cancer tissues and high WBP2 expression correlates with poor clinical outcomes. WBP2 was found to be required for GC cell migration but is dispensable for cell growth and proliferation. WBP2 knockdown increased p-LATS2 with a concomitant increase in p-YAP, resulting in the cytoplasmic retention of YAP and ultimately the inhibition of YAP/TEAD activity and downregulation of TEAD target genes--CTGF and CYR61. Importantly, the loss of LATS2 reversed the activation of Hippo pathway caused by WBP2 knockdown, indicating that WBP2 acts through LATS2 to exert its function on the Hippo pathway. Moreover, WBP2 interacted with LATS2 to inhibit its phosphorylation and activity. In conclusion, our study established a pivotal role for WBP2 in the promotion of GC cell migration via a novel mechanism that inactivates the Hippo pathway transducer LATS2.
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Movimiento Celular/fisiología , Proteínas Serina-Treonina Quinasas/metabolismo , Neoplasias Gástricas/metabolismo , Transactivadores/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Proliferación Celular/fisiología , Humanos , Immunoblotting , Inmunohistoquímica , Proteínas Serina-Treonina Quinasas/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Transducción de Señal/genética , Transducción de Señal/fisiología , Neoplasias Gástricas/genética , Transactivadores/genética , Proteínas Supresoras de Tumor/genéticaRESUMEN
WBP2 transcription coactivator is an emerging oncoprotein and a key node of convergence between EGF and Wnt signaling pathways. Understanding how WBP2 is regulated has important implications for cancer therapy. WBP2 is tightly controlled by post-translational modifications, including phosphorylation and ubiquitination, leading to changes in subcellular localization, protein-protein interactions, and protein turnover. As the function of WBP2 is intricately linked to YAP and TAZ, we hypothesize that WBP2 is negatively regulated by the Hippo tumor suppressor pathway. Indeed, MST is demonstrated to negatively regulate WBP2 expression in a kinase-dependent but LATS-independent manner. This was observed in the majority of the breast cancer cell lines tested. The effect of MST was enhanced by SAV and concomitant with the inhibition of the transcription co-activation, in vitro and in vivo tumorigenesis activities of WBP2, resulting in good prognosis in xenografts. Downregulation of WBP2 by MST involved miRNA but not proteasomal or lysosomal degradation. Our data support the existence of a novel MST-Dicer signaling axis, which in turn regulates both WBP2 CDS- and UTR-targeting miRNAs expression, including miR-23a. MiR-23a targets the 3'UTR of WBP2 mRNA directly. Significant inverse relationships between WBP2 and MST or miR23a expression levels in clinical specimens were observed. In conclusion, WBP2 is a target of the Hippo/MST kinase; MST is identified as yet another rheostat in the regulation of WBP2 and its oncogenic function. The findings have implications in targeted therapeutics and precision medicine for breast cancer.
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Neoplasias de la Mama/metabolismo , ARN Helicasas DEAD-box/metabolismo , Ribonucleasa III/metabolismo , Transactivadores/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Línea Celular Tumoral , ARN Helicasas DEAD-box/genética , Expresión Génica/genética , Regulación Neoplásica de la Expresión Génica/genética , Vía de Señalización Hippo , Humanos , Quinasas Quinasa Quinasa PAM/metabolismo , Quinasas Quinasa Quinasa PAM/fisiología , Células MCF-7 , MicroARNs/genética , MicroARNs/metabolismo , Proteínas Oncogénicas/genética , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/fisiología , Ribonucleasa III/genética , Transducción de Señal/genética , Transactivadores/fisiología , Factores de Transcripción/metabolismo , Vía de Señalización WntRESUMEN
WW domain-binding protein 2 (WBP2) is an emerging oncoprotein. Over the past decade, WBP2 surfaced as a key node connecting key signaling pathways associated with ER/PR, EGFR, PI3K, Hippo, and Wnt in cancer. In addition to the oncogenic functions of WBP2, this review discusses the latest research regarding the multilevel regulation and modes of action of WBP2 and how they can be exploited for molecular medicine. In translational research, evidence supports the role of WBP2 as a biomarker for early detection, prognosis, and companion diagnostics in breast cancer. Finally, we envision new trends in WBP2 research in the space of molecular etiology of cancer, targeted therapeutics, and precision medicine.
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Neoplasias de la Mama/metabolismo , Proteínas Oncogénicas/metabolismo , Transactivadores/metabolismo , Neoplasias de la Mama/diagnóstico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Femenino , Humanos , Proteínas Oncogénicas/genética , Transactivadores/genéticaRESUMEN
PURPOSE: Trastuzumab-based chemotherapy has shown remarkable clinical benefits for patients with HER2-positive breast cancer. However, treatment regimens involving trastuzumab had little or no effect for a subset of patients. Preliminary studies revealed WW-binding protein 2 (WBP2), an oncogenic transcription coactivator, to be coamplified with HER2 in 36% of HER2-positive breast cancers. We hypothesize that WBP2 regulates and correlates with the response of HER2-positive breast cancer to trastuzumab. EXPERIMENTAL DESIGN: The coexpression of WBP2 and HER2 in breast tumors was validated using IHC. The role and mechanism of WBP2 in regulating breast cancer response to trastuzumab was elucidated using in vitro, patient-derived xenograft and murine xenograft models. A multicenter retrospective study involving 143 patients given neoadjuvant trastuzumab-based chemotherapy was conducted to determine whether WBP2 expression correlates with pathologic complete response (pCR). RESULTS: Elevated expression of WBP2 significantly enhanced breast cancer's response to trastuzumab by augmenting trastuzumab-induced HER2 downregulation and cell-cycle arrest via inhibition of cyclin D expression. High level of WBP2 correlated with better pCR (67.19%) compared with low WBP2 level (26.58%). The highest response was observed in subgroups of patients with high WBP2-expressing tumors also aged below 50 years (77.78%) or were premenopausal in status (73.33%). Retrospectively, WBP2 demonstrated sensitivity of 80% to 81% and specificity of 76.5% to 80% in discriminating between patients showing pCR and non-pCR. CONCLUSIONS: WBP2 expression correlates with the response of HER2-positive breast cancer to trastuzumab-based neoadjuvant chemotherapy.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Regulación Neoplásica de la Expresión Génica , Receptor ErbB-2/genética , Transactivadores/genética , Adulto , Anciano , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Modelos Animales de Enfermedad , Femenino , Amplificación de Genes , Redes Reguladoras de Genes , Humanos , Persona de Mediana Edad , Terapia Neoadyuvante , Clasificación del Tumor , Estadificación de Neoplasias , Fosforilación , Receptor ErbB-2/metabolismo , Estudios Retrospectivos , Transducción de Señal/efectos de los fármacos , Trastuzumab/administración & dosificación , Resultado del Tratamiento , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The transcriptional coactivator WW domain-binding protein 2 (WBP2) is an emerging oncogene and serves as a node between the signaling protein Wnt and other signaling molecules and pathways, including epidermal growth factor receptor, estrogen receptor/progesterone receptor, and the Hippo pathway. The upstream regulation of WBP2 is well-studied, but its downstream activity remains unclear. Here, we elucidated WBP2's role in triple-negative breast cancer (TNBC), in which Wnt signaling is predominantly activated. Using RNAi coupled with RNA-Seq and MS analyses to identify Wnt/WBP2- and WBP2-dependent targets in MDA-MB-231 TNBC cells, we found that WBP2 is required for the expression of a core set of genes in Wnt signaling. These included AXIN2, which was essential for Wnt/WBP2-mediated breast cancer growth and migration. WBP2 also regulated a much larger set of genes and proteins independently of Wnt, revealing that WBP2 primes cells to Wnt activity by up-regulating G protein pathway suppressor 1 (GPS1) and TRAF2- and NCK-interacting kinase (TNIK). GPS1 activated the c-Jun N-terminal kinase (JNK)/Jun pathway, resulting in a positive feedback loop with TNIK that mediated Wnt-induced AXIN2 expression. WBP2 promoted TNBC growth by integrating JNK with Wnt signaling, and its expression profoundly influenced the sensitivity of TNBC to JNK/TNIK inhibitors. In conclusion, WBP2 links JNK to Wnt signaling in TNBC. GPS1 and TNIK are constituents of a WBP2-initiated cascade that primes responses to Wnt ligands and are also important for TNBC biology. We propose that WBP2 is a potential drug target for JNK/TNIK-based precision medicine for managing TNBC.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias de la Mama Triple Negativas/metabolismo , Vía de Señalización Wnt , Proteínas Adaptadoras Transductoras de Señales/genética , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HEK293 , Células HeLa , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas JNK Activadas por Mitógenos/genética , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , MAP Quinasa Quinasa 4/antagonistas & inhibidores , MAP Quinasa Quinasa 4/genética , MAP Quinasa Quinasa 4/metabolismo , Células MCF-7 , Proteínas de Neoplasias/genética , Inhibidores de Proteínas Quinasas/farmacología , Transactivadores , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/patologíaRESUMEN
WW domain binding protein 2 (WBP2), a transcriptional coactivator, plays a vital role in breast tumorigenesis. It positively regulates estrogen receptor, Hippo, and Wnt pathways, which subsequently enhance the transcription of downstream target genes contributing to cancer. Understanding the regulation of the expression and activity of WBP2 oncoprotein has implication in cancer therapy. We have previously reported that WBP2 is regulated at the post-translational and post-transcriptional levels. However, its regulation at the transcriptional level is not known. In this study, the minimal promoter region of WBP2 that is critical for its transcription was identified. The E-box motif in the WBP2 promoter was demonstrated to be essential for its transcription. The E-box binding protein upstream stimulatory factor 1 (USF-1) was discovered to be a key transcription factor for WBP2 by yeast one-hybrid analysis and was validated through reporter and chromatin immunoprecipitation assays and tandem mass spectrometry, which also suggested that USF-1 acts by regulating a network of genes, in addition to WBP2, associated with cell movement, proliferation, cell-cycle, and survival cellular processes. USF-1 is overexpressed in majority of the breast cancer cell lines and tissues tested, and has profound effects on cancer cell proliferation. USF-1-mediated transcription of WBP2 was demonstrated to be inducible by insulin, which led to AKT-mediated phosphorylation of USF-1 that modulated its ability to bind to the WBP2 promoter and activate its transcription. This study sheds new light onto the regulation of the WBP2 oncogene at the transcriptional level by a novel oncogenic transcription factor, USF-1. USF-1 is a potential drug target for treatment of WBP2-positive breast cancer.-Ramos, A., Miow, Q. H., Liang, X., Lin, Q. S., Putti, T. C., Lim, Y. P. Phosphorylation of E-box binding USF-1 by PI3K/AKT enhances its transcriptional activation of the WBP2 oncogene in breast cancer cells.
RESUMEN
Long intergenic noncoding RNAs (lincRNAs) are not fully characterized in disease, although many are involved in controlling differentiation. We discovered and sequenced a novel 4.4 kb human lincRNA called linc-9432 in pterygium, an ocular disease characterized by a wedge lesion. This transcript inhibited differentiation-induced cell death, promoted expression of stem cell markers, and decreased expression of epithelial and mesenchymal differentiation markers. This lincRNA regulated 30 differentiation-related genes in transcriptome analysis and 17/30 gene products were known to be directly associated in a network. When the lincRNA was silenced with pooled siRNA, the levels of these transcripts decreased in accordance with their predicted binding affinity for the lincRNA. TBC1D8B had the strongest affinity, interacting in-vitro at positions 269-292 within TBC1D8B.
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Antígenos de Diferenciación/biosíntesis , Diferenciación Celular , Fibroblastos/metabolismo , Regulación de la Expresión Génica , Pterigion/metabolismo , ARN Largo no Codificante/biosíntesis , Antígenos de Diferenciación/genética , Células Cultivadas , Femenino , Fibroblastos/patología , Perfilación de la Expresión Génica , Humanos , Masculino , Pterigion/genética , Pterigion/patología , ARN Largo no Codificante/genéticaRESUMEN
S100P is known to affect tumor development and metastasis of various cancers, but its role in endometrial cancer is unclear. We reported that S100P expression was dramatically elevated in both endometrial squamous cell carcinoma and adenosquamous carcinoma, but not in adenocarcinoma and normal endometrial samples. Moreover, we revealed an oncogenic role of S100P promoting cell proliferation, invasion, and migration while reducing apoptosis, possibly via its upregulation and/or activation of receptors of advanced glycation end products and consequently the oncogenic PI3K-AKT and MAPK pathways. Therefore, S100P might be a specific biomarker and a potential drug target for squamous cell and adenosquamous carcinoma subtypes of endometrial cancer.
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Proteínas de Unión al Calcio/genética , Carcinoma Adenoescamoso/genética , Carcinoma de Células Escamosas/genética , Neoplasias Endometriales/genética , Expresión Génica , Proteínas de Neoplasias/genética , Apoptosis/efectos de los fármacos , Apoptosis/genética , Biomarcadores de Tumor , Proteínas de Unión al Calcio/metabolismo , Carcinoma Adenoescamoso/metabolismo , Carcinoma Adenoescamoso/patología , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Supervivencia Celular/efectos de los fármacos , Cisplatino/farmacología , Neoplasias Endometriales/metabolismo , Neoplasias Endometriales/patología , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Proteínas de Neoplasias/metabolismo , Transducción de Señal , TransfecciónRESUMEN
Pterygium is a triangular shaped ocular fibrous surface lesion growing from conjunctiva towards central cornea, causing ocular irritation, astigmatism, and visual disturbance. The condition is characterized by epithelial proliferation, fibrovascular growth, chronic inflammation, and prominent extracellular matrix remodeling. Studies have suggested that aberrant extracellular proteins secreted by fibroblasts lead to abnormal matrix production and tissue invasion contributing to the development of the disease. In this study, secreted proteins collected from paired pterygium and conjunctival fibroblasts in vitro were identified and quantified by LC-MS iTRAQ-based analysis, in which 433 proteins common to all samples were identified. Among these proteins, 48.0% (208) were classified as classically secreted proteins, 17.1% (74) were exported out of the cells via non-classical secretion pathways, and 31.2% (135) were exosome proteins. A minority (3.7%) was not previously known to be secreted, or might be contaminants. 31 and 27 proteins were found up- or down-regulated in the conditioned media of pterygium fibroblasts relative to the media of control cells, respectively. Molecular function analysis showed that these proteins either belonged to catalytic proteins, structural molecules or were involved with receptor activities and protein binding. Further pathway analysis revealed that these proteins were involved in ECM-receptor interaction, focal adhesion, cancer-related, p53 signaling, complement and coagulation, and TGF-beta signaling pathways. These molecules identified may serve as extracellular ligands to activate intracellular pathways, possibly serving as potential therapeutic targets.
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Conjuntiva/metabolismo , Proteínas del Ojo/metabolismo , Proteómica/métodos , Pterigion/metabolismo , Anciano , Western Blotting , Adhesión Celular , Células Cultivadas , Conjuntiva/patología , Medios de Cultivo Condicionados , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Masculino , Espectrometría de Masas , Persona de Mediana Edad , Pterigion/patología , ARN Mensajero , Transducción de SeñalRESUMEN
Cross-talk between the Hippo and Wnt pathways has been implicated recently in breast cancer development, but key intersections have yet to be fully defined. Here we report that WBP2, a transcription coactivator that binds the Hippo pathway transcription factor YAP/TAZ, contributes to Wnt signaling and breast cancer pathogenesis. Clinically, overexpression of WBP2 in breast cancer specimens correlated with malignant progression and poor patient survival. In breast cancer cells, nuclear entry and interaction of WBP2 with ß-catenin was stimulated by Wnt3A, thereby activating TCF-mediated transcription and driving malignant invasive character. Mechanistic investigations showed WBP2 levels were controlled by the E3 ligase ITCH, which bound and target WBP2 for ubiquitin-dependent proteasomal degradation. Accordingly, ITCH silencing could elevate WBP2 levels. Wnt signaling upregulated WBP2 by disrupting ITCH-WBP2 interactions via EGFR-mediated tyrosine phosphorylation of WBP2 and TAZ/YAP competitive binding. Conversely, ITCH-mediated downregulation of WBP2 inhibited TCF/ß-catenin transcription, in vitro transformation, and in vivo tumorigenesis. We identified somatic mutations in ITCH, which impaired its ability to degrade WBP2 and to block its function in cancer, even while retaining binding capacity to WBP2. Thus, the Wnt pathway appeared to engage WBP2 primarily by affecting its protein stability. Our findings show how WBP2/ITCH signaling functions to link the intricate Wnt and Hippo signaling networks in breast cancer. Cancer Res; 76(21); 6278-89. ©2016 AACR.
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Proteínas Adaptadoras Transductoras de Señales/fisiología , Neoplasias de la Mama/patología , Proteínas Nucleares/metabolismo , Proteínas Represoras/fisiología , Factores de Transcripción/metabolismo , Ubiquitina-Proteína Ligasas/fisiología , Vía de Señalización Wnt/fisiología , Aciltransferasas , Animales , Proteínas de Ciclo Celular , Línea Celular Tumoral , Receptores ErbB/fisiología , Femenino , Humanos , Ratones , Complejo de la Endopetidasa Proteasomal/fisiología , Transactivadores , Proteínas Supresoras de Tumor/fisiología , Proteína Wnt3A/fisiologíaRESUMEN
Metastasis is a significant health issue. The standard mode of care is combination of chemotherapy and targeted therapeutics but the 5-year survival rate remains low. New/better drug targets that can improve outcomes of patients with metastatic disease are needed. Metastasis is a complex process, with each step conferred by a set of genetic aberrations. Mapping the molecular changes associated with metastasis improves our understanding of the etiology of this disease and contributes to the pipeline of targeted therapeutics. Here, phosphoproteomics of a xenograft-derived in vitro model comprising 4 isogenic cell lines with increasing metastatic potential implicated Transient Receptor Potential Vanilloid subtype 4 in breast cancer metastasis. TRPV4 mRNA levels in breast, gastric and ovarian cancers correlated with poor clinical outcomes, suggesting a wide role of TRPV4 in human epithelial cancers. TRPV4 was shown to be required for breast cancer cell invasion and transendothelial migration but not growth/proliferation. Knockdown of Trpv4 significantly reduced the number of metastatic nodules in mouse xenografts leaving the size unaffected. Overexpression of TRPV4 promoted breast cancer cell softness, blebbing, and actin reorganization. The findings provide new insights into the role of TRPV4 in cancer extravasation putatively by reducing cell rigidity through controlling the cytoskeleton at the cell cortex.
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Citoesqueleto de Actina/metabolismo , Neoplasias de la Mama/patología , Canales Catiónicos TRPV/metabolismo , Animales , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/mortalidad , Calcio/metabolismo , Línea Celular Tumoral , Movimiento Celular , Supervivencia sin Enfermedad , Femenino , Humanos , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/secundario , Células MCF-7 , Ratones , Fosfopéptidos/análisis , Interferencia de ARN , ARN Mensajero/metabolismo , ARN Interferente Pequeño/metabolismo , Canales Catiónicos TRPV/antagonistas & inhibidores , Canales Catiónicos TRPV/genética , Migración Transendotelial y Transepitelial , Trasplante Heterólogo , Regulación hacia ArribaRESUMEN
Incomplete knowledge of the mechanisms at work continues to hamper efforts to maximize reprogramming efficiency. Here, we present a systematic genome-wide RNAi screen to determine the global regulators during the early stages of human reprogramming. Our screen identifies functional repressors and effectors that act to impede or promote the reprogramming process. Repressors and effectors form close interacting networks in pathways, including RNA processing, G protein signaling, protein ubiquitination, and chromatin modification. Combinatorial knockdown of five repressors (SMAD3, ZMYM2, SFRS11, SAE1, and ESET) synergistically resulted in â¼85% TRA-1-60-positive cells. Removal of the novel splicing factor SFRS11 during reprogramming is accompanied by rapid acquisition of pluripotency-specific spliced forms. Mechanistically, SFRS11 regulates exon skipping and mutually exclusive splicing of transcripts in genes involved in cell differentiation, mRNA splicing, and chromatin modification. Our study provides insights into the reprogramming process, which comprises comprehensive and multi-layered transcriptional, splicing, and epigenetic machineries.
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Reprogramación Celular/genética , Interferencia de ARN , Células Cultivadas , Técnicas de Silenciamiento del Gen , Pruebas Genéticas , Genoma Humano , Humanos , Cinética , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Empalme del ARN/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño/metabolismo , Proteínas Represoras/metabolismo , Factores de Empalme Serina-Arginina/metabolismoRESUMEN
Most recently approved anti-cancer drugs by the US FDA are targeted therapeutic agents and this represents an important trend for future anticancer therapy. Unlike conventional chemotherapy that rarely considers individual differences, it is crucial for targeted therapies to identify the beneficial subgroup of patients for the treatment. Currently, genomics and transcriptomics are the major 'omic' analytics used in studies of drug response prediction. However, proteomic profiling excels both in its advantages of directly detecting an instantaneous dynamic of the whole proteome, which contains most current diagnostic markers and therapeutic targets. Moreover, proteomic profiling improves understanding of the mechanism for drug resistance and helps finding optimal combination therapy. This article reviews the recent success of applications of proteomic analytics in predicting the response to targeted anticancer therapeutics, and discusses the potential avenues and pitfalls of proteomic platforms and techniques used most in the field.
Asunto(s)
Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/metabolismo , Terapia Molecular Dirigida/métodos , Neoplasias/tratamiento farmacológico , Proteoma/metabolismo , Proteómica/métodos , Animales , Perfilación de la Expresión Génica/métodos , Humanos , Neoplasias/diagnóstico , Proteoma/genéticaRESUMEN
Gastric cancer is the second leading cause of cancer-related deaths worldwide. Gastric cancer is often detected at a late stage when treatment is difficult. Biomarkers for early detection and drug targets for gastric cancer therapy are critical for effective management of gastric cancer. Secreted proteins not only play integral roles in cancer progression and metastasis, they are also easily accessible. Secreted proteins within the tumor microenvironment are therefore an attractive source of biomarkers and drug targets. In this study, iTRAQ-based liquid chromatography/tandem mass spectrometry was used for comparative profiling of the secretomes of 11 gastric cancer cell lines versus a normal gastric epithelial cell line. Of the close to 800 proteins detected, about 600 proteins were detected to display differential expression in one or more gastric cancer cell lines compared to normal cells. These differentially expressed proteins predominantly have binding or enzymatic activities and are largely associated with cellular and metabolic processes. Overexpression of ARPC4 was validated in gastric cell lines and its novel function in gastric cancer cell migration and invasion demonstrated in vitro. The findings support the notion of ARPC4 as a potential biomarker/drug target for metastatic gastric cancer.
Asunto(s)
Proteínas de Neoplasias/metabolismo , Proteoma/metabolismo , Neoplasias Gástricas/metabolismo , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Movimiento Celular/fisiología , Perfilación de la Expresión Génica/métodos , Humanos , Invasividad Neoplásica/patología , Proteómica/métodos , Microambiente Tumoral/fisiologíaRESUMEN
Ubiquitin modification of the TGF-ß pathway components is emerging as a key mechanism of TGF-ß pathway regulation. To limit TGF-ß responses, TGF-ß signaling is regulated through a negative feedback loop whereby the E3 ligase SMURF2 targets the TGF-ß receptor (TßR) complex for ubiquitin-mediated degradation. Counteracting this process, a number of deubiquitinating (DUBs) enzymes have recently been identified that deubiquitinate and stabilize the TßR. However the precise mechanism by which these DUBs act on TßR function remains poorly defined. Here, we demonstrate that apart from targeting the TßR complex directly, USP15 also deubiquitinates SMURF2 resulting in enhanced TßR stability and downstream pathway activation. Through proteomic analysis, we show that USP15 modulates the ubiquitination of Lys734, a residue required for SMURF2 catalytic activity. Our results show that SMURF2 is a critical target of USP15 in the TGF-ß pathway and may also explain how USP15 and SMURF2 target multiple complementary protein complexes in other pathways.
Asunto(s)
Ubiquitina-Proteína Ligasas/metabolismo , Proteasas Ubiquitina-Específicas/fisiología , Ubiquitinación , Secuencia de Aminoácidos , Movimiento Celular , Células HEK293 , Humanos , Cinética , Lisina/metabolismo , Estructura Terciaria de Proteína , Proteolisis , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
An iTRAQ-based tandem mass spectrometry approach was employed to relatively quantify proteins in the membrane proteome of eleven gastric cancer cell lines relative to a denominator non-cancer gastric epithelial cell line HFE145. Of the 882 proteins detected, 57 proteins were found to be upregulated with > 1.3-fold change in at least 6 of the 11 cell lines. Bioinformatics analysis revealed that these proteins are significantly associated with cancer, cell growth and proliferation, death, survival and cell movement. The catalogue of membrane proteins presented that are potential regulators/effectors of gastric cancer progression has implications in cancer therapy. DLAT, a subunit of the pyruvate dehydrogenase complex, was selected as a candidate protein for further studies as its function in gastric cancer has yet to be established. SiRNA studies supported a role of DLAT in gastric cancer cell proliferation and carbohydrate metabolism, reprogramming of which is a hallmark of cancer. Our study contributes to recent interest and discussion in cancer energetics and related phenomena such as the Warburg and Reverse Warburg effects. Future mechanistic studies should lead to the elucidation of the mode of action of DLAT in human gastric cancer and establish DLAT as a viable drug target.