RESUMEN
Neuronal polarity in the developing cortex begins during the early stages of neural progenitor migration toward the cortical plate and culminates with the specification of the axon and dendrites. Here, we demonstrate that the Ran-dependent nucleocytoplasmic transport machinery is essential for the establishment of cortical neuron polarity. We found that Ran-binding protein 1 (RanBP1) regulates axon specification and dendritic arborization in cultured neurons in vitro and radial neural migration in vivo. During axonogenesis, RanBP1 regulates the cytoplasmic levels of the polarity protein LKB1/Par4, and this is dependent on the nuclear export machinery. Our results show that downstream of RanBP1, LKB1 function is mediated by the STK25-GM130 pathway, which promotes axonogenesis through Golgi regulation. Our results indicate that the nucleocytoplasmic transport machinery is a main regulator of neuron polarity, including radial migration, and that the regulated export of LKB1 through RanBP1 is a limiting step of axonogenesis.
Asunto(s)
Proteínas de Drosophila/metabolismo , Aparato de Golgi/metabolismo , Neuronas/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Quinasas/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP , Animales , Axones/metabolismo , Western Blotting , Movimiento Celular/fisiología , Polaridad Celular/genética , Polaridad Celular/fisiología , Células Cultivadas , Citoplasma/metabolismo , Proteínas de Unión al ADN/metabolismo , Drosophila , Proteínas de Drosophila/genética , Femenino , Ratones , Neuronas/citología , Proteínas Nucleares/genética , Células PC12 , Embarazo , Proteínas Quinasas/genética , Ratas , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteína de Unión al GTP ran/metabolismoRESUMEN
Spatiotemporal regulation of gene expression depends on the cooperation of multiple mechanisms, including the functional interaction of promoters with distally located enhancers. Here, we show that, in cortical neurons, a subset of short interspersed nuclear elements (SINEs) located in the proximity of activity-regulated genes bears features of enhancers. Enhancer SINEs (eSINEs) recruit the Pol III cofactor complex TFIIIC in a stimulus-dependent manner and are transcribed by Pol III in response to neuronal depolarization. Characterization of an eSINE located in proximity to the Fos gene (FosRSINE1) indicated that the FosRSINE1-encoded transcript interacts with Pol II at the Fos promoter and mediates Fos relocation to Pol II factories, providing an unprecedented molecular link between Pol III and Pol II transcription. Strikingly, knockdown of the FosRSINE1 transcript induces defects of both cortical radial migration in vivo and activity-dependent dendritogenesis in vitro, demonstrating that FosRSINE1 acts as a strong enhancer of Fos expression in diverse physiological contexts.
Asunto(s)
ARN Polimerasa III/metabolismo , ARN Polimerasa II/metabolismo , Animales , Ratones , Neuronas/metabolismo , Regiones Promotoras Genéticas/genética , ARN Polimerasa II/genética , ARN Polimerasa III/genética , Secuencias Reguladoras de Ácidos Nucleicos/genética , Elementos de Nucleótido Esparcido Corto/genética , Factores de Transcripción TFIII/genética , Factores de Transcripción TFIII/metabolismo , Transcripción Genética/genéticaRESUMEN
Gain-of-function mutations in histone 3 (H3) variants are found in a substantial proportion of pediatric high-grade gliomas (pHGG), often in association with TP53 loss and platelet-derived growth factor receptor alpha (PDGFRA) amplification. Here, we describe a somatic mouse model wherein H3.3K27M and Trp53 loss alone are sufficient for neoplastic transformation if introduced in utero. H3.3K27M-driven lesions are clonal, H3K27me3 depleted, Olig2 positive, highly proliferative, and diffusely spreading, thus recapitulating hallmark molecular and histopathological features of pHGG. Addition of wild-type PDGFRA decreases latency and increases tumor invasion, while ATRX knockdown is associated with more circumscribed tumors. H3.3K27M-tumor cells serially engraft in recipient mice, and preliminary drug screening reveals mutation-specific vulnerabilities. Overall, we provide a faithful H3.3K27M-pHGG model which enables insights into oncohistone pathogenesis and investigation of future therapies.
Asunto(s)
Células Madre Embrionarias/metabolismo , Glioma/genética , Histonas/genética , Células-Madre Neurales/metabolismo , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/genética , Proteína p53 Supresora de Tumor/genética , Animales , Encéfalo/metabolismo , Encéfalo/patología , Transformación Celular Neoplásica/genética , Regulación Neoplásica de la Expresión Génica , Glioma/metabolismo , Glioma/patología , Humanos , Ratones , Mutación , Clasificación del Tumor , Invasividad Neoplásica , Interferencia de ARN , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Proteína Nuclear Ligada al Cromosoma X/genética , Proteína Nuclear Ligada al Cromosoma X/metabolismoRESUMEN
Histone modifications and chromatin remodeling represent universal mechanisms by which cells adapt their transcriptional response to rapidly changing environmental conditions. Extensive chromatin remodeling takes place during neuronal development, allowing the transition of pluripotent cells into differentiated neurons. Here, we report that the NuRD complex, which couples ATP-dependent chromatin remodeling with histone deacetylase activity, regulates mouse brain development. Subunit exchange of CHDs, the core ATPase subunits of the NuRD complex, is required for distinct aspects of cortical development. Whereas CHD4 promotes the early proliferation of progenitors, CHD5 facilitates neuronal migration and CHD3 ensures proper layer specification. Inhibition of each CHD leads to defects of neuronal differentiation and migration, which cannot be rescued by expressing heterologous CHDs. Finally, we demonstrate that NuRD complexes containing specific CHDs are recruited to regulatory elements and modulate the expression of genes essential for brain development.
Asunto(s)
Corteza Cerebral/embriología , Corteza Cerebral/metabolismo , Ensamble y Desensamble de Cromatina , Complejo Desacetilasa y Remodelación del Nucleosoma Mi-2/metabolismo , Subunidades de Proteína/metabolismo , Animales , Ciclo Celular , Movimiento Celular , Eliminación de Gen , Regulación del Desarrollo de la Expresión Génica , Ratones Endogámicos C57BL , Microcefalia/patología , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Regiones Promotoras Genéticas/genéticaRESUMEN
Dynamic epigenetic modifications play a key role in mediating the expression of genes required for neuronal development. We previously identified nitric oxide (NO) as a signaling molecule that mediates S-nitrosylation of histone deacetylase 2 (HDAC2) and epigenetic changes in neurons. Here, we show that HDAC2 nitrosylation regulates neuronal radial migration during cortical development. Bead-array analysis performed in the developing cortex revealed that brahma (Brm), a subunit of the ATP-dependent chromatin-remodeling complex BRG/brahma-associated factor, is one of the genes regulated by S-nitrosylation of HDAC2. In the cortex, expression of a mutant form of HDAC2 that cannot be nitrosylated dramatically inhibits Brm expression. Our study identifies NO and HDAC2 nitrosylation as part of a signaling pathway that regulates cortical development and the expression of Brm in neurons.
Asunto(s)
Movimiento Celular , Ensamble y Desensamble de Cromatina , Histona Desacetilasa 2/metabolismo , Neuronas/citología , Óxido Nítrico/metabolismo , Factores de Transcripción/metabolismo , Animales , Separación Celular , Corteza Cerebral/citología , Corteza Cerebral/metabolismo , Electroporación , Femenino , Citometría de Flujo , Ratones , Embarazo , Transducción de SeñalRESUMEN
During development of the cerebral cortex, neural stem cells (NSCs) undergo a temporal switch from proliferative (symmetric) to neuron-generating (asymmetric) divisions. We investigated the role of Schwann cell factor 1 (SC1/PRDM4), a member of the PRDM family of transcription factors, in this critical transition. We discovered that SC1 recruits the chromatin modifier PRMT5, an arginine methyltransferase that catalyzes symmetric dimethylation of histone H4 arginine 3 (H4R3me2s) and that this modification is preferentially associated with undifferentiated cortical NSCs. Overexpressing SC1 in embryonic NSCs led to an increase in the number of Nestin-expressing precursors; mutational analysis of SC1 showed that this was dependent on recruitment of PRMT5. We found that SC1 protein levels are down-regulated at the onset of neurogenesis and that experimental knockdown of SC1 in primary NSCs triggers precocious neuronal differentiation. We propose that SC1 and PRMT5 are components of an epigenetic regulatory complex that maintains the "stem-like" cellular state of the NSC by preserving their proliferative capacity and modulating their cell cycle progression. Our findings provide evidence that histone arginine methylation regulates NSC differentiation.
Asunto(s)
Diferenciación Celular , Proteínas de Unión al ADN/metabolismo , Histonas/metabolismo , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Proteína Metiltransferasas/metabolismo , Factores de Transcripción/metabolismo , Animales , Arginina , Proliferación Celular , Corteza Cerebral/citología , Corteza Cerebral/embriología , Proteínas de Unión al ADN/química , Embrión de Mamíferos/citología , Técnicas de Silenciamiento del Gen , Células HEK293 , Humanos , Inmunoprecipitación , Metilación , Ratones , Células PC12 , Estructura Terciaria de Proteína , Proteína-Arginina N-Metiltransferasas , ARN Interferente Pequeño/metabolismo , Ratas , Factores de Tiempo , Factores de Transcripción/químicaRESUMEN
A fundamental feature of central nervous system development is that neurons are generated before glia. In the embryonic spinal cord, for example, a group of neuroepithelial stem cells (NSCs) generates motor neurons (MNs), before switching abruptly to oligodendrocyte precursors (OLPs). We asked how transcription factor OLIG2 participates in this MN-OLP fate switch. We found that Serine 147 in the helix-loop-helix domain of OLIG2 was phosphorylated during MN production and dephosphorylated at the onset of OLP genesis. Mutating Serine 147 to Alanine (S147A) abolished MN production without preventing OLP production in transgenic mice, chicks, or cultured P19 cells. We conclude that S147 phosphorylation, possibly by protein kinase A, is required for MN but not OLP genesis and propose that dephosphorylation triggers the MN-OLP switch. Wild-type OLIG2 forms stable homodimers, whereas mutant (unphosphorylated) OLIG2(S147A) prefers to form heterodimers with Neurogenin 2 or other bHLH partners, suggesting a molecular basis for the switch.