RESUMEN
Zinc Finger MYND (Myeloid, Nervy, and DEAF-1) type containing 8 (ZMYND8) is a crucial epigenetic regulator that plays a multifaceted role in governing a spectrum of vital cellular processes, encompassing proliferation, apoptosis, migration, tumor suppression, and differentiation. It has emerged as a key player in neuronal differentiation by orchestrating the expression of neuronal lineage-committed genes. The present study uncovers the role of ZMYND8 in regulating the Sonic Hedgehog (SHH) signaling axis, which is crucial for neuronal differentiation. Genetic deletion of ZMYND8 leads to a significant reduction in SHH pathway genes, GLI1, and PTCH1 expression during all-trans-retinoic acid (ATRA)-induced differentiation. ZMYND8 and RNA pol II S5P are found to co-occupy the GLI1 and PTCH1 gene promoters, positively impacting their gene transcription upon ATRA treatment. Interestingly, ZMYND8 is found to counteract the inhibitory effects of Cyclopamine that block the upstream SHH pathway protein SMO, resulting in enhanced neurite formation in neuroblastoma cells following their treatment with ATRA. These results indicate that ZMYND8 is an epigenetic regulator of the SHH signaling pathway and has tremendous therapeutic potential in ATRA-mediated differentiation of neuroblastoma.
Asunto(s)
Diferenciación Celular , Proteínas Hedgehog , Neuroblastoma , Transducción de Señal , Tretinoina , Proteínas Hedgehog/metabolismo , Proteínas Hedgehog/genética , Humanos , Diferenciación Celular/efectos de los fármacos , Tretinoina/farmacología , Transducción de Señal/efectos de los fármacos , Neuroblastoma/metabolismo , Neuroblastoma/patología , Neuroblastoma/genética , Línea Celular Tumoral , Receptor Patched-1/metabolismo , Receptor Patched-1/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Proteína con Dedos de Zinc GLI1/metabolismo , Proteína con Dedos de Zinc GLI1/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Ratones , Animales , Proteínas Supresoras de TumorRESUMEN
Post-translational modification (PTM) is important in controlling many biological processes by changing the structure and function of a protein. Protein methylation is an important PTM, and the role of methyltransferases has been implicated in numerous cellular functions. Protein L-isoaspartyl methyltransferase (PIMT) is ubiquitously expressed in almost all organisms and govern important cellular processes including apoptosis. Among other functions, PIMT has also been identified as a potent oncogene because it destabilizes the structure of the tumor suppressor p53 via methylation at the transactivation domain. In the present study we identified that out of the three methyltransferase inhibitors tested, namely, S-adenosyl-l-homocysteine (AdoHcy), adenosine and adenosine dialdehyde (AdOx), only AdOx augments p53 expression by destabilizing PIMT structure, as evident from far-UV CD. The effect of the inhibitors, AdOx in particular, to the structure of PIMT, and the binding of PIMT to the p53 transactivation domain have been investigated by docking and molecular dynamics simulations. AdOx significantly increases p53 accumulation and nuclear translocation in colon cancer cells, triggering the p53-mediated apoptotic pathway. To better understand the molecular mechanisms underlying p53 accumulation in colon cancer cells, we observed that the level of PIMT is considerably lower in AdOx-treated cells, reducing its association with p53, which stabilized p53. p53 then transactivated BAX, increasing the BAX: BCL-2 ratio and causing colon cancer cell death.
Asunto(s)
Neoplasias del Colon , Neoplasias Colorrectales , Humanos , Proteína D-Aspartato-L-Isoaspartato Metiltransferasa/metabolismo , Proteína D-Aspartato-L-Isoaspartato Metiltransferasa/farmacología , Proteína X Asociada a bcl-2/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Adenosina/farmacología , Apoptosis , Metiltransferasas/metabolismoRESUMEN
Altered metabolism has become an emerging feature of cancer cells impacting their proliferation and metastatic potential in myriad ways. Proliferating heterogeneous tumor cells are surrounded by other resident or infiltrating cells, along with extracellular matrix proteins, and other secretory factors constituting the tumor microenvironment. The diverse cell types of the tumor microenvironment exhibit different molecular signatures that are regulated at their genetic and epigenetic levels. The cancer cells elicit intricate crosstalks with these supporting cells, exchanging essential metabolites which support their anabolic processes and can promote their survival, proliferation, EMT, angiogenesis, metastasis and even therapeutic resistance. In this context, carbohydrate metabolism ensures constant energy supply being a central axis from which other metabolic and biosynthetic pathways including amino acid and lipid metabolism and pentose phosphate pathway are diverged. In contrast to normal cells, increased glycolytic flux is a distinguishing feature of the highly proliferative cancer cells, which supports them to adapt to a hypoxic environment and also protects them from oxidative stress. Such rewired metabolic properties are often a result of epigenetic alterations in the cancer cells, which are mediated by several factors including, DNA, histone and non-histone protein modifications and non-coding RNAs. Conversely, epigenetic landscapes of the cancer cells are also dictated by their diverse metabolomes. Altogether, this metabolic and epigenetic interplay has immense potential for the development of efficient anti-cancer therapeutic strategies. In this book chapter we emphasize upon the significance of reprogrammed carbohydrate metabolism in regulating the tumor microenvironment and cancer progression, with an aim to explore the different metabolic and epigenetic targets for better cancer treatment.
Asunto(s)
Neoplasias , Microambiente Tumoral , Humanos , Neoplasias/tratamiento farmacológico , Glucólisis/fisiología , Metabolismo de los Hidratos de Carbono , Histonas/metabolismoRESUMEN
Hyperthermia inhibits DNA double-strand break (DSB) repair that utilizes homologous recombination (HR) pathway by a poorly defined mechanism(s); however, the mechanisms for this inhibition remain unclear. Here we report that hyperthermia decreases H4K16 acetylation (H4K16ac), an epigenetic modification essential for genome stability and transcription. Heat-induced reduction in H4K16ac was detected in humans, Drosophila, and yeast, indicating that this is a highly conserved response. The examination of histone deacetylase recruitment to chromatin after heat-shock identified SIRT1 as the major deacetylase subsequently enriched at gene-rich regions. Heat-induced SIRT1 recruitment was antagonized by chromatin remodeler SMARCAD1 depletion and, like hyperthermia, the depletion of the SMARCAD1 or combination of the two impaired DNA end resection and increased replication stress. Altered repair protein recruitment was associated with heat-shock-induced γ-H2AX chromatin changes and DSB repair processing. These results support a novel mechanism whereby hyperthermia impacts chromatin organization owing to H4K16ac deacetylation, negatively affecting the HR-dependent DSB repair.
RESUMEN
Anoikis resistance is an essential property of cancer cells that allow the extra-cellular matrix-detached cells to survive in a suspended state in body fluid in order to metastasize and invade to distant organs. It is known that integrins play an important role in anoikis resistance, but detailed mechanisms are not well understood. Here we report that highly metastatic colon cancer cells showed a higher degree of anoikis resistance than the normal intestinal epithelial cells. These anoikis-resistant cancer cells express high-levels of integrin-α2, ß1, and activated EGFR in the anchorage-independent state than the anchorage-dependent state. In contrast, normal intestinal epithelial cells failed to elevate these proteins. Interestingly, a higher co-association of EGFR with integrin-α2ß1/-α5ß1 was observed on the surface of anoikis-resistant cells. Thus, in the absence of extra-cellular matrix, integrins in association with EGFR activates downstream effectors ERK and AKT and suppress Caspase-3 activation to induce anoikis resistance as was confirmed from the gene-ablation and pharmacological inhibitor studies. Interestingly, these anoikis-resistant cancer cells express high-level of cancer stem cell signatures (CD24, CD44, CD133, EpCAM) and pluripotent stem cell markers (OCT-4, SOX-2, Nanog) as well as drug-resistant pumps (ABCG2, MDR1, MRP1). Altogether, our findings unravel the interplay between integrin-α2ß1/-α5ß1 and EGFR in anoikis resistance and suggest that the resistant cells are cancer initiating or cancer stem cells, which may serve as a promising target to combat metastasis of cancer.
Asunto(s)
Anoicis , Neoplasias del Colon/metabolismo , Integrina alfa2beta1/metabolismo , Células Madre Neoplásicas/metabolismo , Antígeno AC133/metabolismo , Subfamilia B de Transportador de Casetes de Unión a ATP/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/metabolismo , Anoicis/genética , Antígeno CD24/metabolismo , Línea Celular Tumoral , Neoplasias del Colon/genética , Células Epiteliales/metabolismo , Receptores ErbB/genética , Receptores ErbB/metabolismo , Humanos , Receptores de Hialuranos/metabolismo , Integrina alfa2beta1/genética , Sistema de Señalización de MAP Quinasas/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Proteína Homeótica Nanog/metabolismo , Proteínas de Neoplasias/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Factores de Transcripción SOXB1/metabolismoRESUMEN
T-regulatory cells are an upsurge in the tumor microenvironment and induce immune-evasion. CD4+ Treg cells are well characterized whereas the role of CD8+ Tregs in cancer has recently started to crease attention. Here, we report an augmentation CD8+FOXP3+ Tregs in breast tumor microenvironment. FOXP3, the lineage-specific transcription factor, is a dominant regulator of Treg cell development and function. FOXP3 is induced preferentially by divergent signaling in CD4+ Treg cells. But how FOXP3 is induced and maintained in tumor-CD8+ Tregs is the Cinderella of the investigation. We observed that RUNX3, a CD8+ lineage-specific transcription factor, binds at the FOXP3-promoter to induce its transcription. In addition to promoter activation, involvement of cis-elements CNS1 and CNS2 in the transcriptional regulation of FOXP3 was also evident in these cells. SMAD3 binds to CNS1 region and acts as transcription inducer, whereas GATA3 plays a temporal role in the FOXP3 transcription by differential chromatin modification in CNS regions. In CNS1 region, GATA3 acts as a repressor for FOXP3 in naïve CD8+ T cells. Whereas in CD8+ Tregs, GATA3 binds directly at CNS2 region and persuaded the maintenance of FOXP3. Therefore, the intervention of these concerted transcriptional machinery may have a therapeutic potential in immunotherapy of cancer.
Asunto(s)
Linfocitos T CD8-positivos/metabolismo , ADN Intergénico/genética , Factores de Transcripción Forkhead/genética , Regulación de la Expresión Génica , Regiones Promotoras Genéticas , Linfocitos T Reguladores/metabolismo , Transcripción Genética , Adolescente , Adulto , Animales , Secuencia de Bases , Neoplasias de la Mama/inmunología , Neoplasias de la Mama/patología , Secuencia Conservada/genética , Subunidad alfa 3 del Factor de Unión al Sitio Principal/metabolismo , Femenino , Factores de Transcripción Forkhead/metabolismo , Factor de Transcripción GATA3/metabolismo , Humanos , Terapia de Inmunosupresión , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Ratones Endogámicos BALB C , Persona de Mediana Edad , Modelos Genéticos , Proteína smad3/metabolismo , Microambiente Tumoral , Adulto JovenRESUMEN
Tumor suppressor p53 preserves the genomic integrity by restricting anomaly at the gene level. The hotspots for mutation in half of all colon cancers reside in p53. Hence, in a p53-mutated cellular milieu targeting cancer cells may be achievable by targeting the paralogue(s) of p53. Here we have shown the effectiveness of crocetin, a dietary component, in inducing apoptosis of colon cancer cells with varying p53 status. In wild-type p53-expressing cancer cells, p53 in one hand transactivates BAX and in parallel up-regulates p53-induced death domain protein (PIDD) that in turn cleaves and activates BID through caspase-2. Both BAX and t-BID converge at mitochondria to alter the transmembrane potential thereby leading to caspase-9 and caspase-3-mediated apoptosis. In contrast, in functional p53-impaired cells, this phytochemical exploits p53-paralogue p73, which up-regulates FAS to cleave BID through FAS-FADD-caspase-8-pathway. These findings not only underline the phenomenon of functional switch-over from p53 to p73 in p53-impaired condition, but also validate p73 as a promising and potential target for cancer therapy in absence of functional p53.
Asunto(s)
Carotenoides/uso terapéutico , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/metabolismo , Proteínas Adaptadoras de Señalización del Receptor del Dominio de Muerte/metabolismo , Proteína de Dominio de Muerte Asociada a Fas/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Antineoplásicos Fitogénicos/uso terapéutico , Apoptosis/efectos de los fármacos , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/genética , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/metabolismo , Caspasa 2/metabolismo , Caspasa 8/metabolismo , Neoplasias Colorrectales/patología , Cisteína Endopeptidasas/metabolismo , Proteínas Adaptadoras de Señalización del Receptor del Dominio de Muerte/genética , Proteína de Dominio de Muerte Asociada a Fas/genética , Células HCT116 , Células HT29 , Humanos , Mutación , Transducción de Señal/efectos de los fármacos , Proteína Tumoral p73/metabolismo , Proteína p53 Supresora de Tumor/genética , Vitamina A/análogos & derivados , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismo , Receptor fas/metabolismoRESUMEN
p53 preserves genomic integrity by restricting anomaly at the gene level. Till date, limited information is available for cytosol to nuclear shuttling of p53; except microtubule-based trafficking route, which utilizes minus-end directed motor dynein. The present study suggests that monomeric actin (G-actin) guides p53 traffic towards the nucleus. Histidine-tag pull-down assay using purified p53(1-393)-His and G-actin confirms direct physical association between p53 and monomeric G-actin. Co-immunoprecipitation data supports the same. Confocal imaging explores intense perinuclear colocalization between p53 and G-actin. To address atomistic details of the complex, constraint-based docked model of p53:G-actin complex was generated based on crystal structures. MD simulation reveals that p53 DNA-binding domain arrests very well the G-actin protein. Docking benchmark studies have been carried out for a known crystal structure, 1YCS (complex between p53DBD and BP2), which validates the docking protocol we adopted. Co-immunoprecipitation study using "hot-spot" p53 mutants suggested reduced G-actin association with cancer-associated p53 conformational mutants (R175H and R249S). Considering these findings, we hypothesized that point mutation in p53 structure, which diminishes p53:G-actin complexation results in mutant p53 altered subcellular localization. Our model suggests p53Arg249 form polar-contact with Arg357 of G-actin, which upon mutation, destabilizes p53:G-actin interaction and results in cytoplasmic retention of p53R249S.
Asunto(s)
Actinas/metabolismo , Núcleo Celular/metabolismo , Proteínas Mutantes/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Línea Celular Tumoral , Reactivos de Enlaces Cruzados/metabolismo , Doxorrubicina/farmacología , Simulación de Dinámica Molecular , Polimerizacion/efectos de los fármacos , Unión Proteica/efectos de los fármacos , Multimerización de Proteína/efectos de los fármacos , Transporte de Proteínas/efectos de los fármacos , Conejos , Fracciones Subcelulares/metabolismoRESUMEN
BACKGROUND: Cancer metastasis is one of the most common causes of treatment failure and death in cancer patients. It has been acknowledged that aberrant activation of epithelial-to-mesenchymal transition (EMT) program, endows cancer cells with metastatic competence for which E-cadherin switch is a well-established hallmark. Suppression of E-cadherin by its transcriptional repressor Slug is thus a determining factor for EMT. Here, we aimed at discerning (i) the molecular mechanisms that regulate Slug/E-cadherin axis in oncogenic K-ras-expressing non-small cell lung carcinoma (NSCLC) cells, and (ii) the effect of aspirin in modulating the same. METHODS: The migratory behaviour of NSCLC cell line A549 were deciphered by wound healing assay. Further assessment of the molecular mechanisms was done by western blotting, RT-PCR, confocal microscopy, chromatin immunoprecipitation and small interfering RNA (siRNA)-mediated gene silencing. RESULTS: Here we report that in oncogenic K-ras-expressing A549 cells, Ras/ERK downstream Elk-1 forms p-Elk-1-p300 complex that being directly recruited to SLUG promoter acetylates the same to ensure p65NFκB binding for transcriptional up-regulation of Slug, a transcriptional repressor of E-cadherin. Aspirin inhibits EMT and decelerates the migratory potential of A549 cells by down-regulating Slug and thereby up-regulating E-cadherin. Aspirin impedes activation and nuclear translocation of p65NFκB, essential for this transcription factor being available for SLUG promoter binding. As a consequence, Slug transcription is down-regulated relieving A549 cells from Slug-mediated repression of E-cadherin transcription, thereby diminishing the metastatic potential of these oncogenic Ras-expressing NSCLC cells. CONCLUSIONS: Cumulatively, these results signify a crucial role of the anti-inflammatory agent aspirin as a novel negative regulator of epithelial-to-mesenchymal transition thereby suggesting its candidature as a promising tool for deterring metastasis of highly invasive K-ras-expressing NSCLC cells.
Asunto(s)
Aspirina/administración & dosificación , Cadherinas/biosíntesis , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Proteínas Proto-Oncogénicas p21(ras)/biosíntesis , Factores de Transcripción/biosíntesis , Cadherinas/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Transición Epitelial-Mesenquimal/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Regiones Promotoras Genéticas/efectos de los fármacos , Proteínas Proto-Oncogénicas p21(ras)/genética , Factores de Transcripción de la Familia Snail , Factor de Transcripción ReIA/biosíntesis , Factor de Transcripción ReIA/genética , Factores de Transcripción/genéticaRESUMEN
Adverse side effects of chemotherapy during cancer treatment have shifted considerable focus towards therapies that are not only targeted but are also devoid of toxic side effects. We evaluated the antitumorigenic activity of sulphur, and delineated the molecular mechanisms underlying sulphur-induced apoptosis in non-small cell lung carcinoma (NSCLC) cells. A search for the underlying mechanism revealed that the choice between the two cellular processes, NFκBp65-mediated survival and p53-mediated apoptosis, was decided by the competition for a limited pool of transcriptional coactivator protein p300 in NSCLC cells. In contrast, sulphur inhibited otherwise upregulated survival signaling in NSCLC cells by perturbing the nuclear translocation of p65NFκB, its association with p300 histone acetylase, and subsequent transcription of Bcl-2. Under such anti-survival condition, induction of p53-p300 cross-talk enhanced the transcriptional activity of p53 and intrinsic mitochondrial death cascade. Overall, the findings of this preclinical study clearly delineated the molecular mechanism underlying the apoptogenic effect of the non-toxic homeopathic remedy, sulphur, in NSCLC cells.
Asunto(s)
Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Proteína p300 Asociada a E1A/metabolismo , Neoplasias Pulmonares/metabolismo , FN-kappa B/metabolismo , Azufre/farmacología , Proteína p53 Supresora de Tumor/metabolismo , Apoptosis , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Evaluación Preclínica de Medicamentos , Proteína p300 Asociada a E1A/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , FN-kappa B/genética , Transducción de Señal/efectos de los fármacos , Proteína p53 Supresora de Tumor/genéticaRESUMEN
INTRODUCTION: The existence of cancer stem cells (CSCs) has been associated with tumor initiation, therapy resistance, tumor relapse, angiogenesis, and metastasis. Curcumin, a plant ployphenol, has several anti-tumor effects and has been shown to target CSCs. Here, we aimed at evaluating (i) the mechanisms underlying the aggravated migration potential of breast CSCs (bCSCs) and (ii) the effects of curcumin in modulating the same. METHODS: The migratory behavior of MCF-7 bCSCs was assessed by using cell adhesion, spreading, transwell migration, and three-dimensional invasion assays. Stem cell characteristics were studied by using flow cytometry. The effects of curcumin on bCSCs were deciphered by cell viability assay, Western blotting, confocal microscopy, and small interfering RNA (siRNA)-mediated gene silencing. Evaluations of samples of patients with breast cancer were performed by using immunohistochemistry and flow cytometry. RESULTS: Here, we report that bCSCs are endowed with aggravated migration property due to the inherent suppression of the tumor suppressor, E-cadherin, which is restored by curcumin. A search for the underlying mechanism revealed that, in bCSCs, higher nuclear translocation of beta-catenin (i) decreases E-cadherin/beta-catenin complex formation and membrane retention of beta-catenin, (ii) upregulates the expression of its epithelial-mesenchymal transition (EMT)-promoting target genes (including Slug), and thereby (iii) downregulates E-cadherin transcription to subsequently promote EMT and migration of these bCSCs. In contrast, curcumin inhibits beta-catenin nuclear translocation, thus impeding trans-activation of Slug. As a consequence, E-cadherin expression is restored, thereby increasing E-cadherin/beta-catenin complex formation and cytosolic retention of more beta-catenin to finally suppress EMT and migration of bCSCs. CONCLUSIONS: Cumulatively, our findings disclose that curcumin inhibits bCSC migration by amplifying E-cadherin/beta-catenin negative feedback loop.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Cadherinas/metabolismo , Movimiento Celular/efectos de los fármacos , Curcumina/farmacología , Células Madre Neoplásicas/efectos de los fármacos , beta Catenina/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Retroalimentación Fisiológica/efectos de los fármacos , Femenino , Humanos , Células MCF-7 , Células Madre Neoplásicas/patologíaRESUMEN
Lung cancer is the leading cause of cancer-related deaths worldwide. Despite decades of research, the treatment options for lung cancer patients remain inadequate, either to offer a cure or even a substantial survival advantage owing to its intrinsic resistance to chemotherapy. Our results propose the effectiveness of capsaicin in down-regulating VEGF expression in non-small cell lung carcinoma (NSCLC) cells in hypoxic environment. Capsaicin-treatment re-activated p53-SMAR1 positive feed-back loop in these cells to persuade p53-mediated HIF-1α degradation and SMAR1-induced repression of Cox-2 expression that restrained HIF-1α nuclear localization. Such signal-modulations consequently down regulated VEGF expression to thwart endothelial cell migration and network formation, pre-requisites of angiogenesis in tumor micro-environment. The above results advocate the candidature of capsaicin in exclusively targeting angiogenesis by down-regulating VEGF in tumor cells to achieve more efficient and cogent therapy of resistant NSCLC.