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1.
Glia ; 68(12): 2613-2630, 2020 12.
Artículo en Inglés | MEDLINE | ID: mdl-32573857

RESUMEN

Glioblastomas (GBMs) are incurable brain tumors with a high degree of cellular heterogeneity and genetic mutations. Transcription factors that normally regulate neural progenitors and glial development are aberrantly coexpressed in GBM, conferring cancer stem-like properties to drive tumor progression and therapeutic resistance. However, the functional role of individual transcription factors in GBMs in vivo remains elusive. Here, we demonstrate that the basic-helix-loop-helix transcription factor ASCL1 regulates transcriptional targets that are central to GBM development, including neural stem cell and glial transcription factors, oncogenic signaling molecules, chromatin modifying genes, and cell cycle and mitotic genes. We also show that the loss of ASCL1 significantly reduces the proliferation of GBMs induced in the brain of a genetically relevant glioma mouse model, resulting in extended survival times. RNA-seq analysis of mouse GBM tumors reveal that the loss of ASCL1 is associated with downregulation of cell cycle genes, illustrating an important role for ASCL1 in controlling the proliferation of GBM.


Asunto(s)
Neoplasias Encefálicas , Glioblastoma , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neoplasias Encefálicas/genética , Regulación Neoplásica de la Expresión Génica , Genes cdc , Ratones , Factores de Transcripción/metabolismo
2.
Development ; 142(19): 3416-28, 2015 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-26443638

RESUMEN

V1 interneurons are inhibitory neurons that play an essential role in vertebrate locomotion. The molecular mechanisms underlying their genesis remain, however, largely undefined. Here, we show that the transcription factor Prdm12 is selectively expressed in p1 progenitors of the hindbrain and spinal cord in the frog embryo, and that a similar restricted expression profile is observed in the nerve cord of other vertebrates as well as of the cephalochordate amphioxus. Using frog, chick and mice, we analyzed the regulation of Prdm12 and found that its expression in the caudal neural tube is dependent on retinoic acid and Pax6, and that it is restricted to p1 progenitors, due to the repressive action of Dbx1 and Nkx6-1/2 expressed in the adjacent p0 and p2 domains. Functional studies in the frog, including genome-wide identification of its targets by RNA-seq and ChIP-Seq, reveal that vertebrate Prdm12 proteins act as a general determinant of V1 cell fate, at least in part, by directly repressing Dbx1 and Nkx6 genes. This probably occurs by recruiting the methyltransferase G9a, an activity that is not displayed by the amphioxus Prdm12 protein. Together, these findings indicate that Prdm12 promotes V1 interneurons through cross-repressive interactions with Dbx1 and Nkx6 genes, and suggest that this function might have only been acquired after the split of the vertebrate and cephalochordate lineages.


Asunto(s)
Proteínas Portadoras/metabolismo , Regulación del Desarrollo de la Expresión Génica/fisiología , Morfogénesis/fisiología , Proteínas del Tejido Nervioso/metabolismo , Células de Renshaw/fisiología , Xenopus/embriología , Animales , Secuencia de Bases , Embrión de Pollo , Inmunoprecipitación de Cromatina , Biología Computacional , Cartilla de ADN/genética , ADN Complementario/genética , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas de Homeodominio/metabolismo , Inmunohistoquímica , Hibridación in Situ , Ratones , Datos de Secuencia Molecular , Rombencéfalo/metabolismo , Análisis de Secuencia de ARN , Especificidad de la Especie , Médula Espinal/metabolismo
3.
Development ; 141(14): 2803-12, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24924197

RESUMEN

The proper balance of excitatory and inhibitory neurons is crucial for normal processing of somatosensory information in the dorsal spinal cord. Two neural basic helix-loop-helix transcription factors (TFs), Ascl1 and Ptf1a, have contrasting functions in specifying these neurons. To understand how Ascl1 and Ptf1a function in this process, we identified their direct transcriptional targets genome-wide in the embryonic mouse neural tube using ChIP-Seq and RNA-Seq. We show that Ascl1 and Ptf1a directly regulate distinct homeodomain TFs that specify excitatory or inhibitory neuronal fates. In addition, Ascl1 directly regulates genes with roles in several steps of the neurogenic program, including Notch signaling, neuronal differentiation, axon guidance and synapse formation. By contrast, Ptf1a directly regulates genes encoding components of the neurotransmitter machinery in inhibitory neurons, and other later aspects of neural development distinct from those regulated by Ascl1. Moreover, Ptf1a represses the excitatory neuronal fate by directly repressing several targets of Ascl1. Ascl1 and Ptf1a bind sequences primarily enriched for a specific E-Box motif (CAGCTG) and for secondary motifs used by Sox, Rfx, Pou and homeodomain factors. Ptf1a also binds sequences uniquely enriched in the CAGATG E-box and in the binding motif for its co-factor Rbpj, providing two factors that influence the specificity of Ptf1a binding. The direct transcriptional targets identified for Ascl1 and Ptf1a provide a molecular understanding of how these DNA-binding proteins function in neuronal development, particularly as key regulators of homeodomain TFs required for neuronal subtype specification.


Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Redes Reguladoras de Genes , Inhibición Neural , Neuronas/metabolismo , Médula Espinal/citología , Factores de Transcripción/metabolismo , Animales , Secuencia de Bases , Sitios de Unión , Tipificación del Cuerpo/genética , Pollos , Cromatina/metabolismo , Elementos E-Box/genética , Neuronas GABAérgicas/metabolismo , Regulación del Desarrollo de la Expresión Génica , Genoma/genética , Glutamatos/metabolismo , Ratones , Datos de Secuencia Molecular , Tubo Neural/citología , Tubo Neural/embriología , Tubo Neural/metabolismo , Neurogénesis/genética , Neuronas/citología , Motivos de Nucleótidos/genética , Unión Proteica , Médula Espinal/embriología
4.
J Neurosci ; 35(15): 6028-37, 2015 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-25878276

RESUMEN

The intracellular transcriptional milieu wields considerable influence over the induction of neuronal identity. The transcription factor Ptf1a has been proposed to act as an identity "switch" between developmentally related precursors in the spinal cord (Glasgow et al., 2005; Huang et al., 2008), retina (Fujitani et al., 2006; Dullin et al., 2007; Nakhai et al., 2007; Lelièvre et al., 2011), and cerebellum (Hoshino et al., 2005; Pascual et al., 2007; Yamada et al., 2014), where it promotes an inhibitory over an excitatory neuronal identity. In this study, we investigate the potency of Ptf1a to cell autonomously confer a specific neuronal identity outside of its endogenous environment, using mouse in utero electroporation and a conditional genetic strategy to misexpress Ptf1a exclusively in developing cortical pyramidal cells. Transcriptome profiling of Ptf1a-misexpressing cells using RNA-seq reveals that Ptf1a significantly alters pyramidal cell gene expression, upregulating numerous Ptf1a-dependent inhibitory interneuron markers and ultimately generating a gene expression profile that resembles the transcriptomes of both Ptf1a-expressing spinal interneurons and endogenous cortical interneurons. Using RNA-seq and in situ hybridization analyses, we also show that Ptf1a induces expression of the peptidergic neurotransmitter nociceptin, while minimally affecting the expression of genes linked to other neurotransmitter systems. Moreover, Ptf1a alters neuronal morphology, inducing the radial redistribution and branching of neurites in cortical pyramidal cells. Thus Ptf1a is sufficient, even in a dramatically different neuronal precursor, to cell autonomously promote characteristics of an inhibitory peptidergic identity, providing the first example of a single transcription factor that can direct an inhibitory peptidergic fate.


Asunto(s)
Corteza Cerebral/citología , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas del Tejido Nervioso/metabolismo , Células Piramidales/metabolismo , Factores de Transcripción/metabolismo , Transcriptoma/fisiología , Animales , Animales Recién Nacidos , Corteza Cerebral/embriología , Corteza Cerebral/crecimiento & desarrollo , Biología Computacional , Electroporación , Embrión de Mamíferos , Proteínas del Ojo/metabolismo , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Antígeno Ki-67/metabolismo , Ratones , Ratones Transgénicos , Proteínas del Tejido Nervioso/genética , Factor de Transcripción PAX6 , Factores de Transcripción Paired Box/metabolismo , Péptidos/genética , Péptidos/metabolismo , Proteínas Represoras/metabolismo , Estadísticas no Paramétricas , Factores de Transcripción/genética , Tubulina (Proteína)/metabolismo
6.
bioRxiv ; 2023 Oct 02.
Artículo en Inglés | MEDLINE | ID: mdl-37873200

RESUMEN

Glioblastomas (GBMs) are highly aggressive, infiltrative, and heterogeneous brain tumors driven by complex driver mutations and glioma stem cells (GSCs). The neurodevelopmental transcription factors ASCL1 and OLIG2 are co-expressed in GBMs, but their role in regulating the heterogeneity and hierarchy of GBM tumor cells is unclear. Here, we show that oncogenic driver mutations lead to dysregulation of ASCL1 and OLIG2, which function redundantly to initiate brain tumor formation in a mouse model of GBM. Subsequently, the dynamic levels and reciprocal binding of ASCL1 and OLIG2 to each other and to downstream target genes then determine the cell types and degree of migration of tumor cells. Single-cell RNA sequencing (scRNA-seq) reveals that a high level of ASCL1 is key in defining GSCs by upregulating a collection of ribosomal protein, mitochondrial, neural stem cell (NSC), and cancer metastasis genes - all essential for sustaining the high proliferation, migration, and therapeutic resistance of GSCs.

7.
Dev Biol ; 328(2): 529-40, 2009 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-19389376

RESUMEN

Delta-like 3 (Dll3) is a Delta family member expressed broadly in the developing nervous system as neural progenitor cells initiate differentiation. A proximal promoter sequence for Dll3 is conserved across multiple species and is sufficient to direct GFP expression in a Dll3-like pattern in the neural tube of transgenic mice. This promoter contains multiple E-boxes, the consensus binding site for bHLH factors. Dll3 expression and the activity of the Dll3-promoter in the dorsal neural tube depends on the basic helix-loop-helix (bHLH) transcription factors Ascl1 (Mash1) and Neurog2 (Ngn2). Mutations in each E-box identified in the Dll3-promoter allowed distinct enhancer or repressor properties to be assigned to each site individually or in combination. In addition, each E-box has distinct characteristics relative to binding of bHLH factors Ascl1, Neurog1, and Neurog2. Surprisingly, novel Ascl1 containing DNA binding complexes are identified that interact with specific E-box sites within the Dll3-promoter in vitro. These complexes include Ascl1/Ascl1 homodimers and Ascl1/Neurog2 heterodimers, complexes that in some cases require additional undefined factors for efficient DNA binding. Thus, a complex interplay of E-box binding proteins spatially and temporally regulate Dll3 levels during neural tube development.


Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/fisiología , Péptidos y Proteínas de Señalización Intracelular/fisiología , Proteínas de la Membrana/fisiología , Proteínas del Tejido Nervioso/fisiología , Tubo Neural/fisiología , Animales , Secuencia de Bases , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Sitios de Unión , Dimerización , Elementos E-Box , Regulación del Desarrollo de la Expresión Génica , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas de la Membrana/genética , Ratones , Ratones Transgénicos , Datos de Secuencia Molecular , Mutación , Proteínas del Tejido Nervioso/genética , Tubo Neural/embriología , Neuronas/fisiología , Regiones Promotoras Genéticas , Células Madre/fisiología
8.
Transl Oncol ; 11(2): 292-299, 2018 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-29413762

RESUMEN

Small cell lung cancer (SCLC) is an aggressive neuroendocrine carcinoma, designated as a recalcitrant cancer by the National Cancer Institute, in urgent need of new rational therapeutic targets. Previous studies have determined that the basic helix-loop-helix transcription factor achaete-scute homolog 1 (ASCL1) is essential for the survival and progression of a fraction of pulmonary neuroendocrine cancer cells, which include both SCLC and a subset of non-SCLC. Previously, to understand how ASCL1 initiates tumorigenesis in pulmonary neuroendocrine cancer and identify the transcriptional targets of ASCL1, whole-genome RNA-sequencing analysis combined with chromatin immunoprecipitation-sequencing was performed with a series of lung cancer cell lines. From this analysis, we discovered that the gene SCNN1A, which encodes the alpha subunit of the epithelial sodium channel (αENaC), is highly correlated with ASCL1 expression in SCLC. The product of the SCNN1A gene ENaC can be pharmacologically inhibited with amiloride, a drug that has been used clinically for close to 50 years. Amiloride inhibited growth of ASCL1-dependent SCLC more strongly than ASCL1-independent SCLC in vitro and slowed growth of ASCL1-driven SCLC in xenografts. We conclude that SCNN1A/αENaC is a direct transcriptional target of the neuroendocrine lung cancer lineage oncogene ASCL1 that can be pharmacologically targeted with antitumor effects.

9.
Elife ; 62017 08 29.
Artículo en Inglés | MEDLINE | ID: mdl-28850031

RESUMEN

The mechanisms that activate some genes while silencing others are critical to ensure precision in lineage specification as multipotent progenitors become restricted in cell fate. During neurodevelopment, these mechanisms are required to generate the diversity of neuronal subtypes found in the nervous system. Here we report interactions between basic helix-loop-helix (bHLH) transcriptional activators and the transcriptional repressor PRDM13 that are critical for specifying dorsal spinal cord neurons. PRDM13 inhibits gene expression programs for excitatory neuronal lineages in the dorsal neural tube. Strikingly, PRDM13 also ensures a battery of ventral neural tube specification genes such as Olig1, Olig2 and Prdm12 are excluded dorsally. PRDM13 does this via recruitment to chromatin by multiple neural bHLH factors to restrict gene expression in specific neuronal lineages. Together these findings highlight the function of PRDM13 in repressing the activity of bHLH transcriptional activators that together are required to achieve precise neuronal specification during mouse development.


Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Regulación del Desarrollo de la Expresión Génica , N-Metiltransferasa de Histona-Lisina/genética , Neuronas Motoras/metabolismo , Células-Madre Neurales/metabolismo , Neurogénesis/genética , Factores de Transcripción/genética , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Linaje de la Célula/genética , Embrión de Pollo , Embrión de Mamíferos , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , N-Metiltransferasa de Histona-Lisina/metabolismo , Interneuronas/citología , Interneuronas/metabolismo , Ratones , Ratones Transgénicos , Neuronas Motoras/citología , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Células-Madre Neurales/citología , Tubo Neural/citología , Tubo Neural/crecimiento & desarrollo , Tubo Neural/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos/genética , Factor de Transcripción 2 de los Oligodendrocitos/metabolismo , Transducción de Señal , Médula Espinal/citología , Médula Espinal/crecimiento & desarrollo , Médula Espinal/metabolismo , Factores de Transcripción/metabolismo
10.
Cell Rep ; 16(5): 1259-1272, 2016 08 02.
Artículo en Inglés | MEDLINE | ID: mdl-27452466

RESUMEN

Small cell lung carcinoma (SCLC) is a high-grade pulmonary neuroendocrine tumor. The transcription factors ASCL1 and NEUROD1 play crucial roles in promoting malignant behavior and survival of human SCLC cell lines. Here, we find that ASCL1 and NEUROD1 identify heterogeneity in SCLC, bind distinct genomic loci, and regulate mostly distinct genes. ASCL1, but not NEUROD1, is present in mouse pulmonary neuroendocrine cells, and only ASCL1 is required in vivo for tumor formation in mouse models of SCLC. ASCL1 targets oncogenic genes including MYCL1, RET, SOX2, and NFIB while NEUROD1 targets MYC. ASCL1 and NEUROD1 regulate different genes that commonly contribute to neuronal function. ASCL1 also regulates multiple genes in the NOTCH pathway including DLL3. Together, ASCL1 and NEUROD1 distinguish heterogeneity in SCLC with distinct genomic landscapes and distinct gene expression programs.


Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Tumores Neuroendocrinos/genética , Tumores Neuroendocrinos/metabolismo , Animales , Línea Celular Tumoral , Femenino , Expresión Génica/genética , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Masculino , Ratones , Células Neuroendocrinas/metabolismo , Oncogenes/genética , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , Carcinoma Pulmonar de Células Pequeñas/genética , Carcinoma Pulmonar de Células Pequeñas/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
11.
Dev Cell ; 25(2): 182-95, 2013 Apr 29.
Artículo en Inglés | MEDLINE | ID: mdl-23639443

RESUMEN

Generating a balanced network of inhibitory and excitatory neurons during development requires precise transcriptional control. In the dorsal spinal cord, Ptf1a, a basic helix-loop-helix (bHLH) transcription activator, maintains this delicate balance by inducing homeodomain (HD) transcription factors such as Pax2 to specify the inhibitory lineage while suppressing HD factors such as Tlx1/3 that specify the excitatory lineage. We uncover the mechanism by which Ptf1a represses excitatory cell fate in the inhibitory lineage. We identify Prdm13 as a direct target of Ptf1a and reveal that Prdm13 actively represses excitatory cell fate by binding to regulatory sequences near the Tlx1 and Tlx3 genes to silence their expression. Prdm13 acts through multiple mechanisms, including interactions with the bHLH factor Ascl1, to repress Ascl1 activation of Tlx3. Thus, Prdm13 is a key component of a highly coordinated transcriptional network that determines the balance of inhibitory versus excitatory neurons in the dorsal spinal cord.


Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Proteínas de Homeodominio/metabolismo , Neuronas/metabolismo , Médula Espinal/metabolismo , Factores de Transcripción/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Western Blotting , Diferenciación Celular , Células Cultivadas , Embrión de Pollo , Inmunoprecipitación de Cromatina , Cartilla de ADN/química , Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente , Proteínas de Homeodominio/genética , Hibridación in Situ , Ratones , Ratones Noqueados , Neuronas/citología , Factor de Transcripción PAX2/metabolismo , ARN Interferente Pequeño/genética , Médula Espinal/citología , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/genética , Factores de Transcripción/fisiología , Dedos de Zinc
12.
Mol Cell Biol ; 33(16): 3166-79, 2013 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-23754747

RESUMEN

The lineage-specific basic helix-loop-helix transcription factor Ptf1a is a critical driver for development of both the pancreas and nervous system. How one transcription factor controls diverse programs of gene expression is a fundamental question in developmental biology. To uncover molecular strategies for the program-specific functions of Ptf1a, we identified bound genomic regions in vivo during development of both tissues. Most regions bound by Ptf1a are specific to each tissue, lie near genes needed for proper formation of each tissue, and coincide with regions of open chromatin. The specificity of Ptf1a binding is encoded in the DNA surrounding the Ptf1a-bound sites, because these regions are sufficient to direct tissue-restricted reporter expression in transgenic mice. Fox and Sox factors were identified as potential lineage-specific modifiers of Ptf1a binding, since binding motifs for these factors are enriched in Ptf1a-bound regions in pancreas and neural tube, respectively. Of the Fox factors expressed during pancreatic development, Foxa2 plays a major role. Indeed, Ptf1a and Foxa2 colocalize in embryonic pancreatic chromatin and can act synergistically in cell transfection assays. Together, these findings indicate that lineage-specific chromatin landscapes likely constrain the DNA binding of Ptf1a, and they identify Fox and Sox gene families as part of this process.


Asunto(s)
Cromatina/metabolismo , Regulación del Desarrollo de la Expresión Génica , Tubo Neural/embriología , Páncreas/embriología , Factores de Transcripción/metabolismo , Animales , Secuencia de Bases , Línea Celular , Cromatina/genética , Secuencia de Consenso , ADN/genética , ADN/metabolismo , Factor Nuclear 3-beta del Hepatocito/metabolismo , Humanos , Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas/metabolismo , Ratones , Ratones Transgénicos , Tubo Neural/metabolismo , Páncreas/metabolismo , Unión Proteica , Factores de Transcripción SOXB1/metabolismo
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