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Posterior fossa A (PFA) ependymomas are lethal malignancies of the hindbrain in infants and toddlers. Lacking highly recurrent somatic mutations, PFA ependymomas are proposed to be epigenetically driven tumors for which model systems are lacking. Here we demonstrate that PFA ependymomas are maintained under hypoxia, associated with restricted availability of specific metabolites to diminish histone methylation, and increase histone demethylation and acetylation at histone 3 lysine 27 (H3K27). PFA ependymomas initiate from a cell lineage in the first trimester of human development that resides in restricted oxygen. Unlike other ependymomas, transient exposure of PFA cells to ambient oxygen induces irreversible cellular toxicity. PFA tumors exhibit a low basal level of H3K27me3, and, paradoxically, inhibition of H3K27 methylation specifically disrupts PFA tumor growth. Targeting metabolism and/or the epigenome presents a unique opportunity for rational therapy for infants with PFA ependymoma.
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Ependimoma/genética , Ependimoma/metabolismo , Epigenoma/genética , Neoplasias Infratentoriales/genética , Neoplasias Infratentoriales/metabolismo , Animales , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Línea Celular , Proliferación Celular/genética , Metilación de ADN/genética , Epigenómica/métodos , Histonas/genética , Histonas/metabolismo , Humanos , Lactante , Lisina/genética , Lisina/metabolismo , Masculino , Ratones Endogámicos C57BL , Mutación/genéticaRESUMEN
Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype with the worst prognosis and few effective therapies. Here we identified MS023, an inhibitor of type I protein arginine methyltransferases (PRMTs), which has antitumor growth activity in TNBC. Pathway analysis of TNBC cell lines indicates that the activation of interferon responses before and after MS023 treatment is a functional biomarker and determinant of response, and these observations extend to a panel of human-derived organoids. Inhibition of type I PRMT triggers an interferon response through the antiviral defense pathway with the induction of double-stranded RNA, which is derived, at least in part, from inverted repeat Alu elements. Together, our results represent a shift in understanding the antitumor mechanism of type I PRMT inhibitors and provide a rationale and biomarker approach for the clinical development of type I PRMT inhibitors.
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Proteína-Arginina N-Metiltransferasas , Neoplasias de la Mama Triple Negativas , Biomarcadores , Línea Celular Tumoral , Humanos , Interferones/uso terapéutico , Proteína-Arginina N-Metiltransferasas/antagonistas & inhibidores , Proteína-Arginina N-Metiltransferasas/metabolismo , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/metabolismoRESUMEN
Human glioblastomas harbour a subpopulation of glioblastoma stem cells that drive tumorigenesis. However, the origin of intratumoural functional heterogeneity between glioblastoma cells remains poorly understood. Here we study the clonal evolution of barcoded glioblastoma cells in an unbiased way following serial xenotransplantation to define their individual fate behaviours. Independent of an evolving mutational signature, we show that the growth of glioblastoma clones in vivo is consistent with a remarkably neutral process involving a conserved proliferative hierarchy rooted in glioblastoma stem cells. In this model, slow-cycling stem-like cells give rise to a more rapidly cycling progenitor population with extensive self-maintenance capacity, which in turn generates non-proliferative cells. We also identify rare 'outlier' clones that deviate from these dynamics, and further show that chemotherapy facilitates the expansion of pre-existing drug-resistant glioblastoma stem cells. Finally, we show that functionally distinct glioblastoma stem cells can be separately targeted using epigenetic compounds, suggesting new avenues for glioblastoma-targeted therapy.
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Diferenciación Celular , Linaje de la Célula , Rastreo Celular , Glioblastoma/patología , Células Madre Neoplásicas/patología , Animales , Diferenciación Celular/efectos de los fármacos , Linaje de la Célula/efectos de los fármacos , Proliferación Celular , Células Clonales/efectos de los fármacos , Células Clonales/patología , Epigénesis Genética , Femenino , Glioblastoma/tratamiento farmacológico , Xenoinjertos , Humanos , Ratones , Invasividad Neoplásica , Trasplante de Neoplasias , Células Madre Neoplásicas/efectos de los fármacos , Fenotipo , Procesos EstocásticosRESUMEN
Pre-mRNA alternative splicing is modified in cancer, but the origin and specificity of these changes remain unclear. Here, we probed ovarian tumors to identify cancer-associated splicing isoforms and define the mechanism by which splicing is modified in cancer cells. Using high-throughput quantitative PCR, we monitored the expression of splice variants in laser-dissected tissues from ovarian tumors. Surprisingly, changes in alternative splicing were not limited to the tumor tissues but were also found in the tumor microenvironment. Changes in the tumor-associated splicing events were found to be regulated by splicing factors that are differentially expressed in cancer tissues. Overall, â¼20% of the alternative splicing events affected by the down-regulation of the splicing factors QKI and RBFOX2 were altered in the microenvironment of ovarian tumors. Together, our results indicate that the tumor microenvironment undergoes specific changes in alternative splicing orchestrated by a limited number of splicing factors.
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Empalme Alternativo , Neoplasias Ováricas/metabolismo , ARN Mensajero/genética , Línea Celular Tumoral , Células Epiteliales/metabolismo , Femenino , Expresión Génica , Humanos , Captura por Microdisección con Láser , Especificidad de Órganos , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Sitios de Empalme de ARN , Factores de Empalme de ARN , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/fisiología , Proteínas Represoras/fisiología , Células del Estroma/metabolismo , Microambiente TumoralRESUMEN
Chemical probes and chemogenomic compounds are valuable tools to link gene to phenotype, explore human biology, and uncover novel targets for precision medicine. The mission of the Target 2035 initiative is to discover chemical tools for all human proteins by the year 2035. Here, we draw a landscape of the current chemical coverage of human biological pathways. Although available chemical tools target only 3% of the human proteome, they already cover 53% of human biological pathways and represent a versatile toolkit to dissect a vast portion of human biology. Pathways targeted by existing drugs might be enriched in unknown but valid drug targets and could be prioritized in future Target 2035 efforts.
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Cilia are cellular signaling hubs. Given that human kinases are central regulators of signaling, it is not surprising that kinases are key players in cilia biology. In fact, many kinases modulate ciliogenesis, which is the generation of cilia, and distinct ciliary pathways. Several of these kinases are understudied with few publications dedicated to the interrogation of their function. Recent efforts to develop chemical probes for members of the cyclin-dependent kinase like (CDKL), never in mitosis gene A (NIMA) related kinase (NEK), and tau tubulin kinase (TTBK) families either have delivered or are working toward delivery of high-quality chemical tools to characterize the roles that specific kinases play in ciliary processes. A better understanding of ciliary kinases may shed light on whether modulation of these targets will slow or halt disease onset or progression. For example, both understudied human kinases and some that are more well-studied play important ciliary roles in neurons and have been implicated in neurodevelopmental, neurodegenerative, and other neurological diseases. Similarly, subsets of human ciliary kinases are associated with cancer and oncological pathways. Finally, a group of genetic disorders characterized by defects in cilia called ciliopathies have associated gene mutations that impact kinase activity and function. This review highlights both progress related to the understanding of ciliary kinases as well as in chemical inhibitor development for a subset of these kinases. We emphasize known roles of ciliary kinases in diseases of the brain and malignancies and focus on a subset of poorly characterized kinases that regulate ciliary biology.
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Huntingtin protein, mutated in Huntington's disease, is implicated in nucleic acid-mediated processes, yet the evidence for direct huntingtin-nucleic acid interaction is limited. Here, we show wild-type and mutant huntingtin copurify with nucleic acids, primarily RNA, and interact directly with G-rich RNAs in in vitro assays. Huntingtin RNA-immunoprecipitation sequencing from patient-derived fibroblasts and neuronal progenitor cells expressing wild-type and mutant huntingtin revealed long noncoding RNA NEAT1 as a significantly enriched transcript. Altered NEAT1 levels were evident in Huntington's disease cells and postmortem brain tissues, and huntingtin knockdown decreased NEAT1 levels. Huntingtin colocalized with NEAT1 in paraspeckles, and we identified a high-affinity RNA motif preferred by huntingtin. This study highlights NEAT1 as a huntingtin interactor, demonstrating huntingtin's involvement in RNA-mediated functions and paraspeckle regulation.
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Proteína Huntingtina , Enfermedad de Huntington , ARN Largo no Codificante , Proteínas de Unión al ARN , Humanos , Proteína Huntingtina/metabolismo , Proteína Huntingtina/genética , ARN Largo no Codificante/metabolismo , ARN Largo no Codificante/genética , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/genética , Enfermedad de Huntington/patología , Unión Proteica , Fibroblastos/metabolismo , MutaciónRESUMEN
For most of our 25,000 genes, the removal of introns by pre-messenger RNA (pre-mRNA) splicing represents an essential step toward the production of functional messenger RNAs (mRNAs). Alternative splicing of a single pre-mRNA results in the production of different mRNAs. Although complex organisms use alternative splicing to expand protein function and phenotypic diversity, patterns of alternative splicing are often altered in cancer cells. Alternative splicing contributes to tumorigenesis by producing splice isoforms that can stimulate cell proliferation and cell migration or induce resistance to apoptosis and anticancer agents. Cancer-specific changes in splicing profiles can occur through mutations that are affecting splice sites and splicing control elements, and also by alterations in the expression of proteins that control splicing decisions. Recent progress in global approaches that interrogate splicing diversity should help to obtain specific splicing signatures for cancer types. The development of innovative approaches for annotating and reprogramming splicing events will more fully establish the essential contribution of alternative splicing to the biology of cancer and will hopefully provide novel targets and anticancer strategies. Metazoan genes are usually made up of several exons interrupted by introns. The introns are removed from the pre-mRNA by RNA splicing. In conjunction with other maturation steps, such as capping and polyadenylation, the spliced mRNA is then transported to the cytoplasm to be translated into a functional protein. The basic mechanism of splicing requires accurate recognition of each extremity of each intron by the spliceosome. Introns are identified by the binding of U1 snRNP to the 5' splice site and the U2AF65/U2AF35 complex to the 3' splice site. Following these interactions, other proteins and snRNPs are recruited to generate the complete spliceosomal complex needed to excise the intron. While many introns are constitutively removed by the spliceosome, other splice junctions are not used systematically, generating the phenomenon of alternative splicing. Alternative splicing is therefore the process by which a single species of pre-mRNA can be matured to produce different mRNA molecules (Fig. 1). Depending on the number and types of alternative splicing events, a pre-mRNA can generate from two to several thousands different mRNAs leading to the production of a corresponding number of proteins. It is now believed that the expression of at least 70 % of human genes is subjected to alternative splicing, implying an enormous contribution to proteomic diversity, and by extension, to the development and the evolution of complex animals. Defects in splicing have been associated with human diseases (Caceres and Kornblihtt, Trends Genet 18(4):186-93, 2002, Cartegni et al., Nat Rev Genet 3(4):285-98, 2002, Pagani and Baralle, Nat Rev Genet 5(5):389-96, 2004), including cancer (Brinkman, Clin Biochem 37(7):584-94, 2004, Venables, Bioessays 28(4):378-86, 2006, Srebrow and Kornblihtt, J Cell Sci 119(Pt 13):2635-2641, 2006, Revil et al., Bull Cancer 93(9):909-919, 2006, Venables, Transworld Res Network, 2006, Pajares et al., Lancet Oncol 8(4):349-57, 2007, Skotheim and Nees, Int J Biochem Cell Biol 39:1432-1449, 2007). Numerous studies have now confirmed the existence of specific differences in the alternative splicing profiles between normal and cancer tissues. Although there are a few cases where specific mutations are the primary cause for these changes, global alterations in alternative splicing in cancer cells may be primarily derived from changes in the expression of RNA-binding proteins that control splice site selection. Overall, these cancer-specific differences in alternative splicing offer an immense potential to improve the diagnosis and the prognosis of cancer. This review will focus on the functional impact of cancer-associated alternative splicing variants, the molecular determinants that alter the splicing decisions in cancer cells, and future therapeutic strategies.
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Proteómica , Precursores del ARN , Empalme Alternativo , Animales , Humanos , Mutación , Empalme del ARN , ARN MensajeroRESUMEN
Cancer metastasis is a complex cascade that involves the activation of cancer cell migration and invasion of the extracellular space. Cancer-associated fibroblasts (CAFs) are known inducers of cancer cell invasion. However, current in vitro invasion assays such as the Boyden chamber assay are cumbersome and low throughput. Therefore, there is an urgent need for new ex vivo, surrogate invasion assays that can faithfully recapitulate the cancer cell invasion process in vitro and are amenable to large-scale screening of small-molecule libraries in a high-throughput fashion. Here, we describe a well-established high-throughput three-dimensional (3D) spheroid invasion assay as a powerful tool to identify novel molecular targets that can potentially mediate CAF-dependent cancer cell invasion.
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Ensayos Analíticos de Alto Rendimiento , Bibliotecas de Moléculas Pequeñas , Humanos , Ensayos Analíticos de Alto Rendimiento/métodos , Bibliotecas de Moléculas Pequeñas/farmacología , Esferoides Celulares , Movimiento Celular , Invasividad Neoplásica/prevención & control , Línea Celular TumoralRESUMEN
Biallelic inactivation of the tumour suppressor gene Von Hippel-Lindau (VHL) occurs in the vast majority of clear cell renal cell carcinoma (ccRCC) instances, disrupting cellular oxygen-sensing mechanisms to yield a state of persistent pseudo-hypoxia, defined as a continued hypoxic response despite the presence of adequate oxygen levels. However, loss of VHL alone is often insufficient to drive oncogenesis. Results from genomic studies have shown that co-deletions of VHL with one (or more) of three genes encoding proteins involved in chromatin modification and remodelling, polybromo-1 gene (PBRM1), BRCA1-associated protein 1 (BAP1) and SET domain-containing 2 (SETD2), are common and important co-drivers of tumorigenesis. These genes are all located near VHL on chromosome 3p and are often altered following cytogenetic rearrangements that lead to 3p loss and precede the establishment of ccRCC. These three proteins have multiple roles in the regulation of crucial cancer-related pathways, including protection of genomic stability, antagonism of polycomb group (PcG) complexes to maintain a permissive transcriptional landscape in physiological conditions, and regulation of genes that mediate responses to immune checkpoint inhibitor therapy. An improved understanding of these mechanisms will bring new insights regarding cellular drivers of ccRCC growth and therapy response and, ultimately, will support the development of novel translational therapeutics.
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Carcinoma de Células Renales , Neoplasias Renales , Humanos , Carcinogénesis/genética , Carcinoma de Células Renales/patología , Transformación Celular Neoplásica/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Renales/patología , Mutación , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Ubiquitina Tiolesterasa/genética , Ubiquitina Tiolesterasa/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genéticaRESUMEN
Transmembrane protein 106B (TMEM106B), a protein that is localized to the lysosome, is genetically linked to many neurodegenerative diseases and forms fibrils in diseased brains. The reproducibility of TMEM106B research would be enhanced if the community had access to well-characterized anti-TMEM106B antibodies. In this study, we characterized six commercially available TMEM106B antibodies for their performance in Western blot, immunoprecipitation, and immunofluorescence, using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. We identified many high-performing antibodies and encourage readers to use this report as a guide to select the most appropriate antibody for their specific needs.
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Proteínas de la Membrana , Proteínas del Tejido Nervioso , Proteínas de la Membrana/metabolismo , Proteínas del Tejido Nervioso/genética , Reproducibilidad de los Resultados , Western Blotting , Técnica del Anticuerpo Fluorescente , InmunoprecipitaciónRESUMEN
Primary cilia direct cellular signaling events during brain development and neuronal differentiation. The primary cilium is a dynamic organelle formed in a multistep process termed ciliogenesis that is tightly coordinated with the cell cycle. Genetic alterations, such as ciliary gene mutations, and epigenetic alterations, such as post-translational modifications and RNA processing of cilia related factors, give rise to human neuronal disorders and brain tumors such as glioblastoma and medulloblastoma. This review discusses the important role of genetics/epigenetics, as well as RNA processing and post-translational modifications in primary cilia function during brain development and cancer formation. We summarize mouse and human studies of ciliogenesis and primary cilia activity in the brain, and detail how cilia maintain neuronal progenitor populations and coordinate neuronal differentiation during development, as well as how cilia control different signaling pathways such as WNT, Sonic Hedgehog (SHH) and PDGF that are critical for neurogenesis. Moreover, we describe how post-translational modifications alter cilia formation and activity during development and carcinogenesis, and the impact of missplicing of ciliary genes leading to ciliopathies and cell cycle alterations. Finally, cilia genetic and epigenetic studies bring to light cellular and molecular mechanisms that underlie neurodevelopmental disorders and brain tumors.
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A subset of stem-like cells in glioblastoma (GBM; GSC) underlies tumor propagation, therapeutic resistance, and tumor recurrence. Immune evasion is critical for GSCs to carry out these functions. However, the molecular mechanisms employed by GSCs to escape antitumor immunity remain largely unknown. The reprogramming transcription factors Oct4 and Sox2 function as core multipotency factors and play an essential role in the formation and maintenance of GSCs, but the roles of these transcription factors in GSC immune escape have not been well explored. Here we examine how Oct4/Sox2 coexpression contributes to the immunosuppressive phenotype of GSCs. Combined transcription profiling and functional studies of Oct4/Sox2 coexpressing GSCs and differentiated GBM cells demonstrated that Oct4 and Sox2 cooperatively induce an immunosuppressive transcriptome consisting of multiple immunosuppressive checkpoints (i.e., PD-L1, CD70, A2aR, TDO) and dysregulation of cytokines and chemokines that are associated with an immunosuppressive tumor microenvironment. Mechanistically, induction and function of BRD/H3k27Ac-dependent immunosuppressive genes played a role in the immunosuppressive phenotype of GSCs. Pan-BET bromodomain inhibitors (e.g., JQ1) and shBRD4 constructs significantly inhibited the immunosuppressive transcriptome and immunosuppressive biological responses induced by Oct4/Sox2. Our findings identify targetable mechanisms by which tumor-propagating GSCs contribute to the immunosuppressive microenvironment in GBM. SIGNIFICANCE: This report identifies mechanisms by which the reprogramming transcription factors Oct4 and Sox2 function to drive the immunomodulatory transcriptome of GSCs and contribute to the immunosuppressive microenvironment in GBM.
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Proteínas de Ciclo Celular/metabolismo , Tolerancia Inmunológica , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Factores de Transcripción SOXB1/metabolismo , Factores de Transcripción/metabolismo , Transcriptoma/inmunología , Animales , Apoptosis/genética , Neoplasias Encefálicas , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Proteínas de Ciclo Celular/genética , Movimiento Celular/genética , Femenino , Glioblastoma , Humanos , Células Jurkat , Ratones , Ratones Endogámicos C57BL , Células Madre Neoplásicas , Factor 3 de Transcripción de Unión a Octámeros/genética , Factores de Transcripción SOXB1/genética , Células THP-1 , Factores de Transcripción/genética , Transfección , Transgenes , Carga Tumoral/genética , Microambiente Tumoral/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Glioblastoma (GBM) is a deadly cancer in which cancer stem cells (CSCs) sustain tumor growth and contribute to therapeutic resistance. Protein arginine methyltransferase 5 (PRMT5) has recently emerged as a promising target in GBM. Using two orthogonal-acting inhibitors of PRMT5 (GSK591 or LLY-283), we show that pharmacological inhibition of PRMT5 suppresses the growth of a cohort of 46 patient-derived GBM stem cell cultures, with the proneural subtype showing greater sensitivity. We show that PRMT5 inhibition causes widespread disruption of splicing across the transcriptome, particularly affecting cell cycle gene products. We identify a GBM splicing signature that correlates with the degree of response to PRMT5 inhibition. Importantly, we demonstrate that LLY-283 is brain-penetrant and significantly prolongs the survival of mice with orthotopic patient-derived xenografts. Collectively, our findings provide a rationale for the clinical development of brain penetrant PRMT5 inhibitors as treatment for GBM.
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Antineoplásicos/farmacología , Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Proteína-Arginina N-Metiltransferasas/metabolismo , Animales , Apoptosis , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Ciclo Celular , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Descubrimiento de Drogas , Epigenómica , Femenino , Regulación Neoplásica de la Expresión Génica , Glioblastoma/tratamiento farmacológico , Glioblastoma/genética , Glioblastoma/patología , Humanos , Ratones , Células Madre Neoplásicas/metabolismo , Proteína-Arginina N-Metiltransferasas/efectos de los fármacos , Proteína-Arginina N-Metiltransferasas/genética , Empalme del ARN , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Triple negative breast cancer (TNBC) is a deadly form of breast cancer due to the development of resistance to chemotherapy affecting over 30% of patients. New therapeutics and companion biomarkers are urgently needed. Recognizing the elevated expression of glucose transporter 1 (GLUT1, encoded by SLC2A1) and associated metabolic dependencies in TNBC, we investigated the vulnerability of TNBC cell lines and patient-derived samples to GLUT1 inhibition. We report that genetic or pharmacological inhibition of GLUT1 with BAY-876 impairs the growth of a subset of TNBC cells displaying high glycolytic and lower oxidative phosphorylation (OXPHOS) rates. Pathway enrichment analysis of gene expression data suggests that the functionality of the E2F pathway may reflect to some extent OXPHOS activity. Furthermore, the protein levels of retinoblastoma tumor suppressor (RB1) strongly correlate with the degree of sensitivity to GLUT1 inhibition in TNBC, where RB1-negative cells are insensitive to GLUT1 inhibition. Collectively, our results highlight a strong and targetable RB1-GLUT1 metabolic axis in TNBC and warrant clinical evaluation of GLUT1 inhibition in TNBC patients stratified according to RB1 protein expression levels.
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Transportador de Glucosa de Tipo 1/antagonistas & inhibidores , Transportador de Glucosa de Tipo 1/metabolismo , Proteínas de Unión a Retinoblastoma/metabolismo , Neoplasias de la Mama Triple Negativas/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Biomarcadores de Tumor , Neoplasias de la Mama/metabolismo , Ciclo Celular , Línea Celular Tumoral , Proliferación Celular , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Transportador de Glucosa de Tipo 1/genética , Humanos , Ratones , Fosforilación Oxidativa , Proteómica , Pirazoles/farmacología , Piridinas/farmacología , Quinolinas , ARN Mensajero/metabolismo , Neoplasias de la Mama Triple Negativas/genética , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
The Cdx1 gene product is essential for normal anterior-posterior vertebral patterning. Expression of Cdx1 is regulated by several pathways implicated in anterior-posterior patterning events, including retinoid and Wnt signaling. We have previously shown that retinoic acid plays a key role in early stages of Cdx1 expression at embryonic day 7.5 (E7.5), while both Wnt3a signaling and an autoregulatory loop, dependent on Cdx1 itself, are involved in later stages of expression (E8.5 to E9.5). This autoregulation is reflected by the ability of Cdx1 to affect expression from proximal Cdx1 promoter sequences in tissue culture. However, this region is devoid of a demonstrable Cdx response element(s). We have now found that Cdx1 and LEF1, a nuclear effector of Wnt signaling, synergize to induce expression from the Cdx1 promoter through previously documented LEF/T-cell factor response elements. We also found a direct physical interaction between the homeodomain of Cdx1 and the B box of LEF1, suggesting a basis for this synergy. Consistent with these observations, analysis of Cdx1 Wnt3a(vt) compound mutants demonstrated that Wnt and Cdx1 converged on Cdx1 expression and vertebral patterning in vivo. Further data suggest that Cdx-high-mobility group box interactions might be involved in a number of additional pathways.
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Proteínas de Unión al ADN/metabolismo , Proteínas de Homeodominio/metabolismo , ARN Mensajero/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética/fisiología , Animales , Regulación de la Expresión Génica/fisiología , Proteínas HMGB , Técnicas In Vitro , Factor de Unión 1 al Potenciador Linfoide , Ratones , Proteínas Nucleares/metabolismo , Regiones Promotoras Genéticas , Estructura Terciaria de Proteína , Factores de Transcripción SOXB1RESUMEN
Glioblastomas exhibit a hierarchical cellular organization, suggesting that they are driven by neoplastic stem cells that retain partial yet abnormal differentiation potential. Here, we show that a large subset of patient-derived glioblastoma stem cells (GSCs) express high levels of Achaete-scute homolog 1 (ASCL1), a proneural transcription factor involved in normal neurogenesis. ASCL1hi GSCs exhibit a latent capacity for terminal neuronal differentiation in response to inhibition of Notch signaling, whereas ASCL1lo GSCs do not. Increasing ASCL1 levels in ASCL1lo GSCs restores neuronal lineage potential, promotes terminal differentiation, and attenuates tumorigenicity. ASCL1 mediates these effects by functioning as a pioneer factor at closed chromatin, opening new sites to activate a neurogenic gene expression program. Directing GSCs toward terminal differentiation may provide therapeutic applications for a subset of GBM patients and strongly supports efforts to restore differentiation potential in GBM and other cancers.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neoplasias Encefálicas/patología , Carcinogénesis/patología , Linaje de la Célula , Cromatina/metabolismo , Glioblastoma/patología , Neuronas/patología , Secuencia de Bases , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Neoplasias Encefálicas/genética , Carcinogénesis/genética , Diferenciación Celular/genética , Progresión de la Enfermedad , Elementos de Facilitación Genéticos/genética , Regulación Neoplásica de la Expresión Génica , Glioblastoma/genética , Humanos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Neuronas/metabolismo , Regiones Promotoras Genéticas/genética , Unión Proteica , Análisis de Secuencia de ARN , Regulación hacia Arriba/genéticaRESUMEN
The Drosophila neuralized (neur) gene belongs to the neurogenic group of genes involved in regulating cell-cell interactions required for neural precursor development. neur mutant phenotypes include strong overcommitment to neural fates at the expense of epidermal fates. The human neuralized homolog (NEURL) has been recently determined and found to map to chromosome 10q25.1 within the region frequently deleted in malignant astrocytomas. Because of its potential importance in developmental processes, we analyzed the structure of the mouse homolog, Neurl, and its expression pattern in embryonic tissues. Neurl activity is detected from early developmental stages in several tissues and organs including neural tissues, limbs, the skeletal system, sense organs and internal organs undergoing epithelial-mesenchymal interactions. Neurl encodes a polypeptide associated with the plasma membrane but also detected in the cytoplasm. Similarly to the Drosophila gene, mammalian neuralized may code for an important regulatory factor.
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Proteínas del Tejido Nervioso/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Células COS , Plexo Coroideo/química , Plexo Coroideo/embriología , Mapeo Cromosómico , Clonación Molecular , Regulación del Desarrollo de la Expresión Génica , Hibridación in Situ , Hígado/química , Hígado/embriología , Pulmón/química , Pulmón/embriología , Mesencéfalo/química , Mesencéfalo/embriología , Ratones , Ratones Endogámicos BALB C , Microscopía Confocal , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/análisis , Proteínas del Tejido Nervioso/metabolismo , Alineación de Secuencia , Homología de Secuencia de Ácido Nucleico , TransfecciónRESUMEN
The Structural Genomics Consortium (SGC) and its clinical, industry and disease-foundation partners are launching open-source preclinical translational medicine studies.
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Línea Celular , Evaluación Preclínica de Medicamentos/métodos , Cultivo Primario de Células , Humanos , Pacientes , Asociación entre el Sector Público-Privado , Investigación Biomédica TraslacionalRESUMEN
Reprogramming somatic cells into induced pluripotent stem cells (iPSCs) has provided huge insight into the pathways, mechanisms and transcription factors that control differentiation. Here we use high-throughput RT-PCR technology to take a snapshot of splicing changes in the full spectrum of high- and low-expressed genes during induction of fibroblasts, from several donors, into iPSCs and their subsequent redifferentiation. We uncover a programme of concerted alternative splicing changes involved in late mesoderm differentiation and controlled by key splicing regulators MBNL1 and RBFOX2. These critical splicing adjustments arise early in vertebrate evolution and remain fixed in at least 10 genes (including PLOD2, CLSTN1, ATP2A1, PALM, ITGA6, KIF13A, FMNL3, PPIP5K1, MARK2 and FNIP1), implying that vertebrates require alternative splicing to fully implement the instructions of transcriptional control networks.