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Chromosomal rearrangements of the mixed lineage leukemia (MLL) gene occur in â¼10% of B-cell acute lymphoblastic leukemia (B-ALL) and define a group of patients with dismal outcomes. Immunohistochemical staining of bone marrow biopsies from most of these patients revealed aberrant expression of BCL6, a transcription factor that promotes oncogenic B-cell transformation and drug resistance in B-ALL. Our genetic and ChIP-seq (chromatin immunoprecipitation [ChIP] combined with high-throughput sequencing) analyses showed that MLL-AF4 and MLL-ENL fusions directly bound to the BCL6 promoter and up-regulated BCL6 expression. While oncogenic MLL fusions strongly induced aberrant BCL6 expression in B-ALL cells, germline MLL was required to up-regulate Bcl6 in response to physiological stimuli during normal B-cell development. Inducible expression of Bcl6 increased MLL mRNA levels, which was reversed by genetic deletion and pharmacological inhibition of Bcl6, suggesting a positive feedback loop between MLL and BCL6. Highlighting the central role of BCL6 in MLL-rearranged B-ALL, conditional deletion and pharmacological inhibition of BCL6 compromised leukemogenesis in transplant recipient mice and restored sensitivity to vincristine chemotherapy in MLL-rearranged B-ALL patient samples. Oncogenic MLL fusions strongly induced transcriptional activation of the proapoptotic BH3-only molecule BIM, while BCL6 was required to curb MLL-induced expression of BIM. Notably, peptide (RI-BPI) and small molecule (FX1) BCL6 inhibitors derepressed BIM and synergized with the BH3-mimetic ABT-199 in eradicating MLL-rearranged B-ALL cells. These findings uncover MLL-dependent transcriptional activation of BCL6 as a previously unrecognized requirement of malignant transformation by oncogenic MLL fusions and identified BCL6 as a novel target for the treatment of MLL-rearranged B-ALL.
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Regulación Leucémica de la Expresión Génica , Proteína de la Leucemia Mieloide-Linfoide/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/fisiopatología , Proteínas Proto-Oncogénicas c-bcl-6/genética , Proteínas Proto-Oncogénicas c-bcl-6/metabolismo , Animales , Biomarcadores de Tumor/genética , Supervivencia Celular/genética , Células Cultivadas , Eliminación de Gen , Marcación de Gen , Humanos , Ratones , Proteínas de Fusión Oncogénica/genética , Proteínas de Fusión Oncogénica/metabolismo , Pronóstico , Regiones Promotoras Genéticas/genéticaRESUMEN
Knockout of the ubiquitously expressed miRNA-17â¼92 cluster in mice produces a lethal developmental lung defect, skeletal abnormalities, and blocked B lymphopoiesis. A shared target of miR-17â¼92 miRNAs is the pro-apoptotic protein BIM, central to life-death decisions in mammalian cells. To clarify the contribution of miR-17â¼92:Bim interactions to the complex miR-17â¼92 knockout phenotype, we used a system of conditional mutagenesis of the nine Bim 3' UTR miR-17â¼92 seed matches. Blocking miR-17â¼92:Bim interactions early in development phenocopied the lethal lung phenotype of miR-17â¼92 ablation and generated a skeletal kinky tail. In the hematopoietic system, instead of causing the predicted B cell developmental block, it produced a selective inability of B cells to resist cellular stress; and prevented B and T cell hyperplasia caused by Bim haploinsufficiency. Thus, the interaction of miR-17â¼92 with a single target is essential for life, and BIM regulation by miRNAs serves as a rheostat controlling cell survival in specific physiological contexts.
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Linfocitos B/citología , Proteína 11 Similar a Bcl2/metabolismo , Supervivencia Celular/genética , Regulación del Desarrollo de la Expresión Génica/genética , Hematopoyesis/genética , MicroARNs/metabolismo , Regiones no Traducidas 3'/genética , Animales , Linfocitos B/patología , Proteína 11 Similar a Bcl2/genética , Técnicas de Inactivación de Genes , Pulmón/embriología , Ratones , MicroARNs/genética , Mutación , Estrés FisiológicoRESUMEN
Kinetochores form the link between chromosomes and microtubules of the mitotic spindle. The heterodecameric Dam1 complex (Dam1c) is a major component of the Saccharomyces cerevisiae outer kinetochore, assembling into 3 MDa-sized microtubule-embracing rings, but how ring assembly is specifically initiated in vivo remains to be understood. Here, we describe a molecular pathway that provides local control of ring assembly during the establishment of sister kinetochore bi-orientation. We show that Dam1c and the general microtubule plus end-associated protein (+TIP) Bim1/EB1 form a stable complex depending on a conserved motif in the Duo1 subunit of Dam1c. EM analyses reveal that Bim1 crosslinks protrusion domains of adjacent Dam1c heterodecamers and promotes the formation of oligomers with defined curvature. Disruption of the Dam1c-Bim1 interaction impairs kinetochore localization of Dam1c in metaphase and delays mitosis. Phosphorylation promotes Dam1c-Bim1 binding by relieving an intramolecular inhibition of the Dam1 C-terminus. In addition, Bim1 recruits Bik1/CLIP-170 to Dam1c and induces formation of full rings even in the absence of microtubules. Our data help to explain how new kinetochore end-on attachments are formed during the process of attachment error correction.
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Cinetocoros/metabolismo , Proteínas de Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Saccharomycetales/fisiología , Segregación Cromosómica , Mitosis/fisiología , Complejos Multiproteicos/metabolismo , Fosforilación , Unión Proteica , Huso Acromático/metabolismoRESUMEN
Certain BH3-only proteins transiently bind and activate Bak and Bax, initiating their oligomerization and the permeabilization of the mitochondrial outer membrane, a pivotal step in the mitochondrial pathway to apoptosis. Here we describe the first crystal structures of an activator BH3 peptide bound to Bak and illustrate their use in the design of BH3 derivatives capable of inhibiting human Bak on mitochondria. These BH3 derivatives compete for the activation site at the canonical groove, are the first engineered inhibitors of Bak activation, and support the role of key conformational transitions associated with Bak activation.
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Apoptosis/efectos de los fármacos , Proteína 11 Similar a Bcl2 , Mitocondrias , Péptidos , Proteína Destructora del Antagonista Homólogo bcl-2 , Animales , Proteína 11 Similar a Bcl2/química , Proteína 11 Similar a Bcl2/farmacología , Línea Celular Transformada , Humanos , Ratones , Mitocondrias/genética , Mitocondrias/metabolismo , Péptidos/química , Péptidos/farmacología , Unión Proteica , Relación Estructura-Actividad , Proteína Destructora del Antagonista Homólogo bcl-2/química , Proteína Destructora del Antagonista Homólogo bcl-2/genética , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismoRESUMEN
The Bcl-2 family protein Bim triggers mitochondrial apoptosis. Bim is expressed in nonapoptotic cells at the mitochondrial outer membrane, where it is activated by largely unknown mechanisms. We found that Bim is regulated by formation of large protein complexes containing dynein light chain 1 (DLC1). Bim rapidly inserted into cardiolipin-containing membranes in vitro and recruited DLC1 to the membrane. Bim binding to DLC1 induced the formation of large Bim complexes on lipid vesicles, on isolated mitochondria, and in intact cells. Native gel electrophoresis and gel filtration showed Bim-containing mitochondrial complexes of several hundred kilodaltons in all cells tested. Bim unable to form complexes was consistently more active than complexed Bim, which correlated with its substantially reduced binding to anti-apoptotic Bcl-2 proteins. At endogenous levels, Bim surprisingly bound only anti-apoptotic Mcl-1 but not Bcl-2 or Bcl-XL, recruiting only Mcl-1 into large complexes. Targeting of DLC1 by RNAi in human cell lines induced disassembly of Bim-Mcl-1 complexes and the proteasomal degradation of Mcl-1 and sensitized the cells to the Bcl-2/Bcl-XL inhibitor ABT-737. Regulation of apoptosis at mitochondria thus extends beyond the interaction of monomers of proapoptotic and anti-apoptotic Bcl-2 family members but involves more complex structures of proteins at the mitochondrial outer membrane, and targeting complexes may be a novel therapeutic strategy.
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Apoptosis/genética , Proteína 11 Similar a Bcl2/metabolismo , Dineínas/metabolismo , Mitocondrias/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Animales , Proteína 11 Similar a Bcl2/genética , Células CACO-2 , Línea Celular Tumoral , Regulación de la Expresión Génica , Células HeLa , Humanos , Células MCF-7 , Ratones , Unión Proteica , Multimerización de Proteína/genética , Estabilidad Proteica , Interferencia de ARN , Proteína X Asociada a bcl-2/genéticaRESUMEN
INTRODUCTION: Estrogen receptor (ER) positive patients compromise about 70% of breast cancers. Tamoxifen, an antagonist of ERα66 (the classic ER), is the most effective and the standard first-line drug. However, its efficacy is limited by the development of acquired resistance. METHODS: A specific inhibitor of Hsp70-Bim protein-protein interaction (PPI), S1g-2, together with an inhibitor of Hsp70-Bag3 PPI, MKT-077 and an ATP-competitive inhibitor VER155008, were used as chemical tools. Cell viability assays, co-immunoprecipitation and gene knockdown were used to investigate the role of Hsp70 in tamoxifen resistance. A xenograft model was established in which tamoxifen-resistant breast cancer (MCF-7/TAM-R) cells maintained in the presence of 5 µM tamoxifen were subcutaneously inoculated. The anti-tumor efficiency of S1g-2 was measured after a daily injection of 0.8 mg/kg for 14 days. RESULTS: It was revealed that Hsp70-Bim PPI protects ERα-positive breast cancer from tamoxifen-induced apoptosis through binding and stabilizing ERα36, rather than ERα66, resulting in sustained EGFR mRNA and protein expression. Disruption of Hsp70-Bim PPI and downregulation of ERα36 expression in tumor samples are consistent with the in vitro functions of S1g-2, resulting in about a three-fold reduction in tumor volume. CONCLUSIONS: The in vivo activity and safety of S1g-2 illustrated that it is a potential strategy for Hsp70-Bim disruption to overcome tamoxifen-resistant ER-positive breast cancer.
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Neoplasias de la Mama , Tamoxifeno , Humanos , Femenino , Tamoxifeno/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proteína 11 Similar a Bcl2/genética , Proteína 11 Similar a Bcl2/metabolismo , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Receptor alfa de Estrógeno/genética , Receptor alfa de Estrógeno/metabolismo , Regulación Neoplásica de la Expresión GénicaRESUMEN
Viral myocarditis (VMC) is the major reason for sudden cardiac death among both children and young adults. Of these, coxsackievirus B3 (CVB3) is the most common causative agent of myocarditis. Recently, the role of signaling pathways in the pathogenesis of VMC has been evaluated in several studies, which has provided a new perspective on identifying potential therapeutic targets for this hitherto incurable disease. In the present study, in vivo and in vitro experiments showed that CVB3 infection leads to increased Bim expression and triggers apoptosis. In addition, by knocking down Bim using RNAi, we further confirmed the biological function of Bim in apoptosis induced by CVB3 infection. We additionally found that Bim and forkhead box O1 class (FOXO1) inhibition significantly increased the viability of CVB3-infected cells while blocking viral replication and viral release. Moreover, CVB3-induced Bim expression was directly dependent on FOXO1 acetylation, which is catalyzed by the co-regulation of CBP and SirTs. Furthermore, the acetylation of FOXO1 was an important step in Bim activation and apoptosis induced by CVB3 infection. The findings of this study suggest that CVB3 infection induces apoptosis through the FOXO1 acetylation-Bim pathway, thus providing new insights for developing potential therapeutic targets for enteroviral myocarditis.
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Apoptosis , Proteína 11 Similar a Bcl2 , Infecciones por Coxsackievirus , Enterovirus Humano B , Proteína Forkhead Box O1 , Miocarditis , Miocitos Cardíacos , Proteína 11 Similar a Bcl2/metabolismo , Proteína 11 Similar a Bcl2/genética , Apoptosis/genética , Miocitos Cardíacos/virología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O1/genética , Animales , Miocarditis/virología , Miocarditis/metabolismo , Miocarditis/genética , Miocarditis/patología , Enterovirus Humano B/fisiología , Infecciones por Coxsackievirus/genética , Infecciones por Coxsackievirus/virología , Infecciones por Coxsackievirus/metabolismo , Infecciones por Coxsackievirus/patología , Acetilación , Humanos , Masculino , Ratones , Transducción de Señal , RatasRESUMEN
BACKGROUND: Breast cancer is one of the common malignancies in women. Evidence has demonstrated that FBXO45 plays a pivotal role in oncogenesis and progression. However, the role of FBXO45 in breast tumorigenesis remains elusive. Exploration of the regulatory mechanisms of FBXO45 in breast cancer development is pivotal for potential therapeutic interventions in patients with breast cancer. METHODS: Hence, we used numerous approaches to explore the functions of FBXO45 and its underlaying mechanisms in breast cancer pathogenesis, including CCK-8 assay, EdU assay, colony formation analysis, apoptosis assay, RT-PCR, Western blotting, immunoprecipitation, ubiquitination assay, and cycloheximide chase assay. RESULTS: We found that downregulation of FBXO45 inhibited cell proliferation, while upregulation of FBXO45 elevated cell proliferation in breast cancer. Silencing of FBXO45 induced cell apoptosis, whereas overexpression of FBXO45 inhibited cell apoptosis in breast cancer. Moreover, FBXO45 interacted with BIM and regulated its ubiquitination and degradation. Furthermore, knockdown of FBXO45 inhibited cell proliferation via regulation of BIM pathway. Notably, overexpression of FBXO45 facilitated tumor growth in mice. Strikingly, FBXO45 expression was associated with poor survival of breast cancer patients. CONCLUSION: Our study could provide the rational for targeting FBXO45 to obtain benefit for breast cancer patients. Altogether, modulating FBXO45/Bim axis could be a promising strategy for breast cancer therapy.
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Apoptosis , Proteína 11 Similar a Bcl2 , Neoplasias de la Mama , Proliferación Celular , Progresión de la Enfermedad , Proteínas F-Box , Ubiquitinación , Humanos , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/genética , Femenino , Animales , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Ratones , Proteína 11 Similar a Bcl2/metabolismo , Proteína 11 Similar a Bcl2/genética , Línea Celular Tumoral , Proteolisis , Regulación Neoplásica de la Expresión Génica , Ratones DesnudosRESUMEN
Many reports have shown that stabilization of secondary structure by stapling functional peptides enhances the intracellular bioactivity. However, no report has discussed the correlation between stabilization and biological activity based on the configuration of amino acid residues used as anchors for stapling. To clarify this, we investigated the helix content and apoptotic efficiency of an apoptosis-inducing peptide, Bim, and four stapled Bim peptides containing stapling-related Cys residues introduced with different configurations within the sequence. The results demonstrated that the configuration of Cys residues in stapled Bim peptides affected the secondary structure and intracellular activity of the peptides, and furthermore, there was a correlation between these latter two variables.
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Inadequate highway design is a leading cause of traffic accidents, underscoring the importance of adhering to guidelines and regulations for highway design. These standards exist to safeguard road users by addressing crucial factors, like road geometry, signage, and lane markings. Thus, emphasis is placed on computational methods that can optimize towards higher levels of safety, capacity, efficiency, and sustainability in highway designs. Building Information Modeling (BIM) enhances this process by creating a digital model with physical and operational attributes. In this study, a user-friendly, logic-based language is utilized to encode rules for designing highway passing lanes by which designs are automatically evaluated and generated in the BIM-kit software toolkit. This approach is applied to 16 real-world passing lanes in Alberta, showcasing its utility in transportation. The analysis reveals significant enhancements, with rule compliance increasing from 61.82% to 91.31% after employing generative design techniques. These findings underscore the significance of generative design in transportation, offering engineers an efficient tool to create innovative, compliant solutions for highway projects.
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Defect inspection of existing buildings is receiving increasing attention for digitalization transfer in the construction industry. The development of drone technology and artificial intelligence has provided powerful tools for defect inspection of buildings. However, integrating defect inspection information detected from UAV images into semantically rich building information modeling (BIM) is still challenging work due to the low defect detection accuracy and the coordinate difference between UAV images and BIM models. In this paper, a deep learning-based method coupled with transfer learning is used to detect defects accurately; and a texture mapping-based defect parameter extraction method is proposed to achieve the mapping from the image U-V coordinate system to the BIM project coordinate system. The defects are projected onto the surface of the BIM model to enrich a surface defect-extended BIM (SDE-BIM). The proposed method was validated in a defect information modeling experiment involving the No. 36 teaching building of Nantong University. The results demonstrate that the methods are widely applicable to various building inspection tasks.
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The growing interest in building data management, especially the building information model (BIM), has significantly influenced urban management, materials supply chain analysis, documentation, and storage. However, the integration of BIM into 3D GIS tools is becoming more common, showing progress beyond the traditional problem. To address this, this study proposes data transformation methods involving mapping between three domains: industry foundation classes (IFC), city geometry markup language (CityGML), and web ontology framework (OWL)/resource description framework (RDF). Initially, IFC data are converted to CityGML format using the feature manipulation engine (FME) at CityGML standard's levels of detail 4 (LOD4) to enhance BIM data interoperability. Subsequently, CityGML is converted to the OWL/RDF diagram format to validate the proposed BIM conversion process. To ensure integration between BIM and GIS, geometric data and information are visualized through Cesium Ion web services and Unreal Engine. Additionally, an RDF graph is applied to analyze the association between the semantic mapping of the CityGML standard, with Neo4j (a graph database management system) utilized for visualization. The study's results demonstrate that the proposed data transformation methods significantly improve the interoperability and visualization of 3D city models, facilitating better urban management and planning.
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Structural health monitoring (SHM) is critical for ensuring the safety of infrastructure such as bridges. This article presents a digital twin solution for the SHM of railway bridges using low-cost wireless accelerometers and machine learning (ML). The system architecture combines on-premises edge computing and cloud analytics to enable efficient real-time monitoring and complete storage of relevant time-history datasets. After train crossings, the accelerometers stream raw vibration data, which are processed in the frequency domain and analyzed using machine learning to detect anomalies that indicate potential structural issues. The digital twin approach is demonstrated on an in-service railway bridge for which vibration data were collected over two years under normal operating conditions. By learning allowable ranges for vibration patterns, the digital twin model identifies abnormal spectral peaks that indicate potential changes in structural integrity. The long-term pilot proves that this affordable SHM system can provide automated and real-time warnings of bridge damage and also supports the use of in-house-designed sensors with lower cost and edge computing capabilities such as those used in the demonstration. The successful on-premises-cloud hybrid implementation provides a cost effective and scalable model for expanding monitoring to thousands of railway bridges, democratizing SHM to improve safety by avoiding catastrophic failures.
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Anguillid eel populations are under threat globally. A particularly vulnerable life-cycle stage is the migration of mature adult eels downstream from freshwater habitats through estuaries into the sea to spawn. This study investigated the factors associated with downstream migration of the short-finned eel Anguilla australis (Richardson 1841) from a coastal wetland (Lake Condah) in south-east Australia, using acoustic telemetry. Migration was associated with time of the year, higher water level and river flows, decreasing water temperature, and darker moon phases. Larger individuals and those in better condition were more likely to migrate from the wetland. Downstream migration peaked in spring, in contrast to the typical autumn migration period for other temperate anguillids. Variable responses, in comparison to other studies, highlight how migration cues may not be universal. In south-east Australia, short-finned eels may have evolved to migrate in multiple phases by first migrating to the estuary during typical seasonal spring flow pulses (e.g., to avoid being stranded in upland reaches during dry summer periods) and then migrating into the ocean in autumn. More research is needed to unravel these processes and causes, especially considering that the relationship between migration and hydrology may be complex and confounded (e.g., by human-induced disruptions to migratory pathways).
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Anguilla , Migración Animal , Estaciones del Año , Humedales , Animales , Anguilla/fisiología , Telemetría , Temperatura , FemeninoRESUMEN
Glaucoma is one of the leading causes of acquired blindness and characterized by retinal ganglion cell (RGC) death. MicroRNAs are small noncoding RNAs that degrade their target mRNAs. Apoptosis is one of the common mechanisms leading to neuronal death in many neurodegenerative diseases, including glaucoma. In the present study, we identified microRNAs that modulate RGC death caused by the intravitreal injection of N-methyl-d-aspartic acid (NMDA). We found an upregulation of miR-29b and downregulation of miR-124 in the retina of the NMDA-injected eyes. The intravitreal injection of an miR-29b inhibitor 18 h before NMDA injection reduced RGC death and the downregulation of myeloid cell leukemia 1 (MCL-1), an anti-apoptotic factor, induced by intravitreal NMDA. The intravitreal injection of an miR-124 mimic 18 h before NMDA injection also reduced RGC death and the upregulation of B-cell/chronic lymphocytic leukemia lymphoma 2 (bcl-2)-associated X protein (Bax) and bcl-2 interacting protein (Bim), pro-apoptotic factors, induced by intravitreal NMDA. These data suggest that expressional changes in microRNA are involved in the excitotoxicity of RGCs, and that complement and/or inhibition of microRNA may be a potential therapeutic approach for the diseases related to the excitotoxicity of RGCs, such as glaucoma and retinal central artery occlusion.
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Glaucoma , MicroARNs , Oclusión de la Arteria Retiniana , Animales , Ratones , N-Metilaspartato , Muerte Celular , Apoptosis/genética , Retina , MicroARNs/genética , Glaucoma/genética , Proteínas Proto-Oncogénicas c-bcl-2/genéticaRESUMEN
Cervical squamous cell carcinoma (CSCC) is one of the leading causes of cancer death in women worldwide. Patients with advanced cervical carcinoma always have a poor prognosis once resistant to cisplatin due to the lack of effective treatment. It is urgent to investigate the molecular mechanisms of cisplatin resistance. Circular RNAs (circRNAs) are known to exert their regulatory functions in a series of malignancies. However, their effects on CSCC remain to be elucidated. Here, we found that cytoplasmic circARHGAP5, derived from second and third exons of the ARHGAP5 gene, was downregulated in cisplatin-resistant tissues compared with normal cervix tissues and untreated cervical cancer tissues. In addition, experiments from overexpression/knockdown cell lines revealed that circARHGAP5 could inhibit cisplatin-mediated cell apoptosis in CSCC cells both in vitro and in vivo. Mechanistically, circARHGAP5 interacted with AU-rich element RNA-binding protein (AUF1) directly. Overexpression of AUF1 could also inhibit cell apoptosis mediated by cisplatin. Furthermore, we detected the potential targets of AUF1 related to the apoptotic pathway and found that bcl-2-like protein 11 (BIM) was not only negatively regulated by AUF1 but positively regulated by circARHGAP5, which indicated that BIM mRNA might be degraded by AUF1 and thereby inhibited tumor cell apoptosis. Collectively, our data indicated that circARHGAP5 directly bound to AUF1 and prevented AUF1 from interacting with BIM mRNA, thereby playing a pivotal role in cisplatin resistance in CSCC. Our study provides insights into overcoming cancer resistance to cisplatin treatment.
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Carcinoma de Células Escamosas , Ribonucleoproteína Nuclear Heterogénea D0 , ARN Circular , Neoplasias del Cuello Uterino , Femenino , Humanos , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Línea Celular Tumoral , Proliferación Celular , Cisplatino/farmacología , Proteínas Activadoras de GTPasa/genética , Ribonucleoproteína Nuclear Heterogénea D0/metabolismo , ARN Circular/genética , ARN Mensajero/metabolismo , Neoplasias del Cuello Uterino/tratamiento farmacológico , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patologíaRESUMEN
Dysregulated Wnt signaling is associated with malignant oncogenic transformation, especially in colon cancer. Recently, numerous drugs have been developed based on tumorigenesis biomarkers, thus having high potential as drug targets. Likewise, WNT/ß-catenin pathway members are attractive therapeutic targets for colon cancer and are currently in various stages of development. However, although inhibitors of proteins regulating the WNT/ß-catenin signaling pathway have been extensively studied, they have yet to be clinically approved, and the underlying molecular mechanism(s) of their anticancer effects remain poorly understood. Herein, we show that a novel WNT/ß-catenin inhibitor, DGG-300273, inhibits colon cancer cell growth in a Wnt-dependent manner due to upregulation of the BCL2-family protein Bim and caspase-dependent apoptotic cell death. Additionally, DGG-300273-mediated cell death occurs by increased reactive oxygen species (ROS), as shown by abrogation of apoptotic cell death and ROS production following pretreatment with the antioxidant N-acetylcysteine. These results suggest that DGG-300273 represents a promising investigational drug for the treatment of Wnt-associated cancer, thus warranting further characterization and study.
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Neoplasias del Colon , beta Catenina , Humanos , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Vía de Señalización WntRESUMEN
Chronic non-healing wounds, a prevalent complication of diabetes, are associated with increased mortality in diabetic patients. Excessive accumulation of M1 macrophages in diabetic wounds promotes inflammation and results in dysregulated tissue repair. Adipose tissue macrophages (ATMs) derived from healthy lean donors have the ability to improve glucose tolerance and insulin sensitivity, as well as modulate inflammation. MicroRNAs (miRs), which can be packaged into exosomes (Exos) and secreted from cells, serve as essential regulators of macrophage polarization. Here, we revealed that ATMs isolated from lean mice secrete miRs-containing Exos, which modulate macrophage polarization and promote rapid diabetic wound healing when administered to diabetes-prone db/db mice. The miRs sequence of tissue samples from wounds treated with Exos secreted by lean ATMs (ExosLean) revealed that miR-222-3p was up-regulated. Further analyses showed that inhibiting miR-222-3p using a miR inhibitor impaired the macrophage-reprogramming effect of ExosLean. In the excisional skin wound mouse model, locally inhibiting miR-222-3p disrupted healing dynamics and failed to modulate macrophage polarization. Mechanistic studies revealed a connection between miR-222-3p, Bcl2l11/Bim, an inflammatory response effector, macrophage polarization, and diabetic wound healing. In summary, ExosLean act as positive regulators of macrophage polarization by regulating miR levels in wounds and accelerating wound healing, and thus have important implications for wound management in diabetes.
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Diabetes Mellitus , Exosomas , MicroARNs , Ratones , Animales , Tejido Adiposo , MicroARNs/genética , MicroARNs/farmacología , Inflamación , Macrófagos , Cicatrización de HeridasRESUMEN
BACKGROUND: For cancer therapy, the identification of both selective autophagy targets and small molecules that specifically regulate autophagy is greatly needed. Heat shock protein 70 (Hsp70) is a recently discovered BH3 receptor that forms a proteinâprotein interaction (PPI) with Bcl-2-interacting mediator of cell death (Bim). Herein, a specific inhibitor of the Hsp70-Bim PPI, S1g-2, and its analog S1, which is a Bcl-2-Bim disruptor, were used as chemical tools to explore the role of Hsp70-Bim PPI in regulating mitophagy. METHODS: Co-immunoprecipitation and immunofluorescence assays were used to determine protein interactions and colocalization patterns. Organelle purification and immunodetection of LC3-II/LC3-I on mitochondria, endoplasmic reticulum (ER) and Golgi were applied to identify specific types of autophagy. Cell-based and in vitro ubiquitination studies were used to study the role of the Hsp70-Bim PPI in parkin-mediated ubiquitination of outer mitochondrial membrane 20 (TOMM20). RESULTS: We found that after the establishment of their PPI, Hsp70 and Bim form a complex with parkin and TOMM20, which in turn facilitates parkin translocation to mitochondria, TOMM20 ubiquitination and mitophagic flux independent of Bax/Bak. Moreover, S1g-2 selectively inhibits stress-induced mitophagy without interfering with basal autophagy. CONCLUSIONS: The findings highlight the dual protective function of the Hsp70-Bim PPI in regulating both mitophagy and apoptosis. S1g-2 is thus a newly discovered antitumor drug candidate that drives both mitophagy and cell death via apoptosis.
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Proteínas HSP70 de Choque Térmico , Mitofagia , Proteínas HSP70 de Choque Térmico/metabolismo , Autofagia/fisiología , Mitocondrias/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Receptores de Superficie Celular/metabolismoRESUMEN
The amyloid precursor protein (APP) is a cell surface protein of uncertain function that is notable for being the parent protein of beta-amyloid. Research around this protein has focussed heavily on the link to Alzheimer's disease and neurodegeneration. However, there is increasing evidence that APP may be linked to neuronal loss through mechanisms independent of beta-amyloid. FoxO3a is a transcription factor associated with neuronal longevity and apoptosis. In neurons, FoxO3a is associated with cell death through pathways that include BIM, a BCL-2 family member. In this study we have shown that APP overexpression increased the cellular levels and activity of FoxO3a. This increased expression and activity is not a result of decreased phosphorylation but is more likely a result of increased nuclear stability due to increased levels of FANCD2, a binding partner of FoxO3a. The changes caused by APP overexpression were shown to be due to the AICD fragment of APP possibly directly inducing transcription increase in FANCD2. These findings strengthen the link between APP metabolism and FoxO3a neuronal activity. This link may be crucial in better understanding the cellular role of APP and its link to neurodegeneration and aging.