RESUMEN
The development of the cerebellum depends on intricate processes of neurogenesis, migration, and differentiation of neural stem cells (NSCs) and progenitor cells. Defective cerebellar development often results in motor dysfunctions and psychiatric disorders. Understanding the molecular mechanisms that underlie the complex development of the cerebellum will facilitate the development of novel treatment options. Here, we report that the receptor for activated C kinase (Rack1), a multifaceted signaling adaptor protein, regulates mammalian cerebellar development in a cell type-specific manner. Selective deletion of Rack1 in mouse NSCs or granule neuron progenitors (GNPs), but not Bergmann glial cells (BGs), causes severe defects in cerebellar morphogenesis, including impaired folia and fissure formation. NSCs and GNPs lacking Rack1 exhibit enhanced Wnt/ß-catenin signaling but reduced Sonic hedgehog (Shh) signaling. Simultaneous deletion of ß-catenin in NSCs, but not GNPs, significantly rescues the Rack1 mutant phenotype. Interestingly, Rack1 controls the activation of Shh signaling by regulating the ubiquitylation and stability of histone deacetylase 1 (HDAC1)/HDAC2. Suppression of HDAC1/HDAC2 activity in the developing cerebellum phenocopies the Rack1 mutant. Together, these results reveal a previously unknown role of Rack1 in controlling mammalian cerebellar development by opposite regulation of Wnt/ß-catenin and Shh signaling pathways.
Asunto(s)
Cerebelo/crecimiento & desarrollo , Cerebelo/metabolismo , Proteínas Hedgehog/metabolismo , Receptores de Cinasa C Activada/metabolismo , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animales , Regulación del Desarrollo de la Expresión Génica , Proteínas Hedgehog/genética , Histona Desacetilasa 1/genética , Histona Desacetilasa 1/metabolismo , Histona Desacetilasa 2/genética , Histona Desacetilasa 2/metabolismo , Ratones , Ratones Noqueados , Células-Madre Neurales/metabolismo , Neuronas/metabolismo , Receptores de Cinasa C Activada/genética , Transducción de Señal , Proteínas Wnt/genética , beta Catenina/genéticaRESUMEN
The proper development of the cerebral cortex is essential for brain formation and functioning. O-GlcNAcylation, an important posttranslational modification, regulates the pathways critical for neuronal health and the survival of the cerebral cortex in neurodegenerative diseases. However, the role of O-GlcNAcylation in regulating cerebral cortical development at the embryonic and early postnatal (0-21 days) stages is still largely unknown. Here we report that the selective deletion of O-GlcNAc transferase (OGT) in neural stem cells (NSCs) in mice led to a series of severe brain developmental deficits, including dramatic shrinkage of cortical and hippocampal histoarchitecture, widespread neuronal apoptosis, decrease in cell proliferation, induction of endoplasmic reticulum (ER) stress, and inhibition of neuronal dendritic and axonal differentiation. The pathology of corticogenesis deficits caused by OGT deletion may largely rely on complicated biological processes, such as proliferation, apoptosis and differentiation. Our results suggest that dysfunctional O-GlcNAcylation in NSCs may be an important contributor to neurodevelopmental diseases.
Asunto(s)
Corteza Cerebral/crecimiento & desarrollo , N-Acetilglucosaminiltransferasas/fisiología , Células-Madre Neurales/enzimología , Animales , Apoptosis , Axones/ultraestructura , Corteza Cerebral/anatomía & histología , Corteza Cerebral/embriología , Corteza Cerebral/enzimología , Dendritas/ultraestructura , Homólogo 4 de la Proteína Discs Large/metabolismo , Estrés del Retículo Endoplásmico , Masculino , Ratones Noqueados , Morfogénesis , Células Madre Multipotentes/enzimología , N-Acetilglucosaminiltransferasas/genética , Neuronas/citología , Neuronas/metabolismoRESUMEN
We collected 61 craniopharyngioma (CP) specimens to investigate the expression of TrkA, ß-catenin, BRAF gene mutation, and NTRK1 fusion in CP. There were 37 male and 24 female individuals with a median age of 34 years (range, 4-75 years). Histologically, there were 46 cases of adamantinomatous craniopharyngioma (ACP), 14 cases of papillary craniopharyngioma (PCP), and 1 case with a mixed adamantinomatous and papillary pattern. By immunohistochemistry, we found that moderate/high TrkA expression was detected in 47% (28/60) CP and was significantly higher in adult patients (p = 0.018). Interestingly, TrkA is more expressed in "whorled epithelium" cells in ACP, similar to the localization of abnormal ß-catenin. The abnormal expression rate of ß-catenin was 70% (43/61), and the medium/high cyclin D1 expression rate was 73% (44/60), both of which were significantly higher in ACP than in PCP. Of the CP, 41% (21/51) had a moderate/strong P16-positive signal; 58% (34/59) showed a high Ki-67 expression, and there was a significant correlation between high Ki-67 L.I. and high tumor recurrence (p = 0.021). NTRK1 fusion was not found in CP by fluorescence in situ hybridization (FISH). By PCR, 26% (15/58) CP showed BRAF V600E gene mutation, which mainly occurred in PCP (100%, 14/14) except one case of mixed CP. Moreover, TrkA expression was negatively correlated with Ki-67 index and positively correlated with P16 expression. There was a significantly negative correlation between BRAF V600E mutation and abnormal ß-catenin expression. Our results demonstrate for the first time that TrkA expression might occur in CP, especially in adult CP patients, and suggest that cyclin D1 could be used for ACP histological classification in addition to ß-catenin and BRAF V600E mutation, while Ki-67 could be used as a marker to predict CP recurrence.
Asunto(s)
Craneofaringioma , Neoplasias Hipofisarias , Proteínas Proto-Oncogénicas B-raf , Receptor trkA , beta Catenina , Adolescente , Adulto , Anciano , Ciclo Celular , Niño , Preescolar , Craneofaringioma/genética , Ciclina D1/genética , Femenino , Humanos , Hibridación Fluorescente in Situ , Antígeno Ki-67/genética , Masculino , Persona de Mediana Edad , Mutación , Neoplasias Hipofisarias/diagnóstico , Neoplasias Hipofisarias/genética , Neoplasias Hipofisarias/metabolismo , Pronóstico , Proteínas Proto-Oncogénicas B-raf/genética , Receptor trkA/genética , Adulto Joven , beta Catenina/genéticaRESUMEN
Mutations of the X-linked methyl-CpG-binding protein 2 (MECP2) gene in humans are responsible for most cases of Rett syndrome (RTT), an X-linked progressive neurological disorder. While genome-wide screens in clinical trials have revealed several putative RTT-associated mutations in MECP2, their causal relevance regarding the functional regulation of MeCP2 at the etiologic sites at the protein level requires more evidence. In this study, we demonstrated that MeCP2 was dynamically modified by O-linked-ß-N-acetylglucosamine (O-GlcNAc) at threonine 203 (T203), an etiologic site in RTT patients. Disruption of the O-GlcNAcylation of MeCP2 specifically at T203 impaired dendrite development and spine maturation in cultured hippocampal neurons, and disrupted neuronal migration, dendritic spine morphogenesis, and caused dysfunction of synaptic transmission in the developing and juvenile mouse cerebral cortex. Mechanistically, genetic disruption of O-GlcNAcylation at T203 on MeCP2 decreased the neuronal activity-induced induction of Bdnf transcription. Our study highlights the critical role of MeCP2 T203 O-GlcNAcylation in neural development and synaptic transmission potentially via brain-derived neurotrophic factor.
Asunto(s)
Trastornos del Neurodesarrollo , Síndrome de Rett , Animales , Humanos , Proteína 2 de Unión a Metil-CpG/genética , Proteína 2 de Unión a Metil-CpG/metabolismo , Ratones , Trastornos del Neurodesarrollo/genética , Síndrome de Rett/genética , Transmisión Sináptica , TreoninaRESUMEN
Previous studies demonstrate an accumulation of transferrin and transferrin receptor 1 (TfR1) in regenerating peripheral nerves. However, the expression and function of transferrin and TfR1 in the denervated skeletal muscle remain poorly understood. In this study, a mouse model of denervation was produced by complete tear of the left brachial plexus nerve. RNA-sequencing revealed that transferrin expression in the denervated skeletal muscle was upregulated, while TfR1 expression was downregulated. We also investigated the function of TfR1 during development and in adult skeletal muscles in mice with inducible deletion or loss of TfR1. The ablation of TfR1 in skeletal muscle in early development caused severe muscular atrophy and early death. In comparison, deletion of TfR1 in adult skeletal muscles did not affect survival or glucose metabolism, but caused skeletal muscle atrophy and motor functional impairment, similar to the muscular atrophy phenotype observed after denervation. These findings suggest that TfR1 plays an important role in muscle development and denervation-induced muscular atrophy. This study was approved by the Institutional Animal Care and Use Committee of Beijing Institute of Basic Medical Sciences, China (approval No. SYXK 2017-C023) on June 1, 2018.
RESUMEN
Normal neurodevelopment relies on intricate signaling pathways that balance neural stem cell (NSC) self-renewal, maturation, and survival. Disruptions lead to neurodevelopmental disorders, including microcephaly. Here, we implicate the inhibition of NSC senescence as a mechanism underlying neurogenesis and corticogenesis. We report that the receptor for activated C kinase (Rack1), a family member of WD40-repeat (WDR) proteins, is highly enriched in NSCs. Deletion of Rack1 in developing cortical progenitors leads to a microcephaly phenotype. Strikingly, the absence of Rack1 decreases neurogenesis and promotes a cellular senescence phenotype in NSCs. Mechanistically, the senescence-related p21 signaling pathway is dramatically activated in Rack1 null NSCs, and removal of p21 significantly rescues the Rack1-knockout phenotype in vivo. Finally, Rack1 directly interacts with Smad3 to suppress the activation of transforming growth factor (TGF)-ß/Smad signaling pathway, which plays a critical role in p21-mediated senescence. Our data implicate Rack1-driven inhibition of p21-induced NSC senescence as a critical mechanism behind normal cortical development.
Asunto(s)
Senescencia Celular , Corteza Cerebral/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Microcefalia/metabolismo , Células-Madre Neurales/metabolismo , Neurogénesis , Receptores de Cinasa C Activada/metabolismo , Animales , Proliferación Celular , Senescencia Celular/efectos de los fármacos , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/crecimiento & desarrollo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Regulación del Desarrollo de la Expresión Génica , Predisposición Genética a la Enfermedad , Células HEK293 , Humanos , Ratones Endogámicos C57BL , Ratones Noqueados , Microcefalia/genética , Microcefalia/patología , Microcefalia/fisiopatología , Células-Madre Neurales/efectos de los fármacos , Células-Madre Neurales/patología , Neurogénesis/efectos de los fármacos , Fenotipo , Receptores de Cinasa C Activada/genética , Transducción de Señal , Proteína smad3/genética , Proteína smad3/metabolismo , Factor de Crecimiento Transformador beta1/farmacologíaRESUMEN
Purkinje cells (PCs) in the cerebellum receive two excitatory afferents including granule cells-derived parallel fiber (PF) and the climbing fiber. Scaffolding protein Rack1 is highly expressed in the cerebellar PCs. Here, we found delayed formation of specific cerebellar vermis lobule and impaired motor coordination in PC-specific Rack1 conditional knockout mice. Our studies further revealed that Rack1 is essential for PF-PC synapse formation. In addition, Rack1 plays a critical role in regulating synaptic plasticity and long-term depression (LTD) induction of PF-PC synapses without changing the expression of postsynaptic proteins. Together, we have discovered Rack1 as the crucial molecule that controls PF-PC synaptogenesis and synaptic plasticity. Our studies provide a novel molecular insight into the mechanisms underlying the neural development and neuroplasticity in the cerebellum.
RESUMEN
Chromosome alignment is required for accurate chromosome segregation. Chromosome misalignment can result in genomic instability and tumorigenesis. Here, we show that NF-κB activating protein (NKAP) is critical for chromosome alignment through anchoring CENP-E to kinetochores. NKAP knockdown causes chromosome misalignment and prometaphase arrest in human cells. NKAP dynamically localizes to kinetochores, and is required for CENP-E kinetochore localization. NKAP is SUMOylated predominantly in mitosis and the SUMOylation is needed for NKAP to bind CENP-E. A SUMOylation-deficient mutant of NKAP cannot support the localization of CENP-E on kinetochores or proper chromosome alignment. Moreover, Bub3 recruits NKAP to stabilize the binding of CENP-E to BubR1 at kinetochores. Importantly, loss of NKAP expression causes aneuploidy in cultured cells, and is observed in human soft tissue sarcomas. These findings indicate that NKAP is a novel and key regulator of mitosis, and its dysregulation might contribute to tumorigenesis by causing chromosomal instability.