RESUMEN
Tourette disorder (TD) is poorly understood, despite affecting 1/160 children. A lack of animal models possessing construct, face, and predictive validity hinders progress in the field. We used CRISPR/Cas9 genome editing to generate mice with mutations orthologous to human de novo variants in two high-confidence Tourette genes, CELSR3 and WWC1. Mice with human mutations in Celsr3 and Wwc1 exhibit cognitive and/or sensorimotor behavioral phenotypes consistent with TD. Sensorimotor gating deficits, as measured by acoustic prepulse inhibition, occur in both male and female Celsr3 TD models. Wwc1 mice show reduced prepulse inhibition only in females. Repetitive motor behaviors, common to Celsr3 mice and more pronounced in females, include vertical rearing and grooming. Sensorimotor gating deficits and rearing are attenuated by aripiprazole, a partial agonist at dopamine type II receptors. Unsupervised machine learning reveals numerous changes to spontaneous motor behavior and less predictable patterns of movement. Continuous fixed-ratio reinforcement shows that Celsr3 TD mice have enhanced motor responding and reward learning. Electrically evoked striatal dopamine release, tested in one model, is greater. Brain development is otherwise grossly normal without signs of striatal interneuron loss. Altogether, mice expressing human mutations in high-confidence TD genes exhibit face and predictive validity. Reduced prepulse inhibition and repetitive motor behaviors are core behavioral phenotypes and are responsive to aripiprazole. Enhanced reward learning and motor responding occur alongside greater evoked dopamine release. Phenotypes can also vary by sex and show stronger affection in females, an unexpected finding considering males are more frequently affected in TD.
Asunto(s)
Dopamina , Mutación , Síndrome de Tourette , Animales , Síndrome de Tourette/genética , Síndrome de Tourette/fisiopatología , Síndrome de Tourette/metabolismo , Ratones , Femenino , Masculino , Humanos , Dopamina/metabolismo , Recompensa , Cuerpo Estriado/metabolismo , Modelos Animales de Enfermedad , Aprendizaje/fisiología , Conducta Animal , Inhibición Prepulso/genética , Filtrado Sensorial/genéticaRESUMEN
The ability of the skin to grow in response to stretching has been exploited in reconstructive surgery1. Although the response of epidermal cells to stretching has been studied in vitro2,3, it remains unclear how mechanical forces affect their behaviour in vivo. Here we develop a mouse model in which the consequences of stretching on skin epidermis can be studied at single-cell resolution. Using a multidisciplinary approach that combines clonal analysis with quantitative modelling and single-cell RNA sequencing, we show that stretching induces skin expansion by creating a transient bias in the renewal activity of epidermal stem cells, while a second subpopulation of basal progenitors remains committed to differentiation. Transcriptional and chromatin profiling identifies how cell states and gene-regulatory networks are modulated by stretching. Using pharmacological inhibitors and mouse mutants, we define the step-by-step mechanisms that control stretch-mediated tissue expansion at single-cell resolution in vivo.
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Mecanotransducción Celular/fisiología , Análisis de la Célula Individual , Piel/citología , Piel/crecimiento & desarrollo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Uniones Adherentes/metabolismo , Animales , Secuencia de Bases , Proteínas de Ciclo Celular/metabolismo , Diferenciación Celular/efectos de los fármacos , Autorrenovación de las Células/efectos de los fármacos , Cromatina/efectos de los fármacos , Cromatina/genética , Ensamble y Desensamble de Cromatina/efectos de los fármacos , Células Clonales/citología , Células Clonales/efectos de los fármacos , Células Clonales/metabolismo , Modelos Animales de Enfermedad , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Redes Reguladoras de Genes/efectos de los fármacos , Hidrogeles/administración & dosificación , Hidrogeles/farmacología , Mecanotransducción Celular/efectos de los fármacos , Mecanotransducción Celular/genética , Ratones , Ratones Transgénicos , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Mutación , ARN Mensajero/genética , RNA-Seq , Piel/efectos de los fármacos , Células Madre/citología , Células Madre/efectos de los fármacos , Células Madre/metabolismo , Transactivadores/antagonistas & inhibidores , Transactivadores/metabolismo , Factor de Transcripción AP-1/metabolismo , Transcripción Genética/efectos de los fármacos , Proteínas Señalizadoras YAPRESUMEN
KIF2A is an atypical kinesin that has the capacity to depolymerize microtubules. Patients carrying mutations in KIF2A suffer from progressive microcephaly and mental disabilities. While the role of this protein is well documented in neuronal migration, the relationship between its dysfunction and the pathobiology of brain disorders is unclear. Here, we report that KIF2A is dispensable for embryogenic neurogenesis but critical in postnatal stages for maturation, connectivity, and maintenance of neurons. We used a conditional approach to inactivate KIF2A in cortical progenitors, nascent postmitotic neurons, and mature neurons in mice. We show that the lack of KIF2A alters microtubule dynamics and disrupts several microtubule-dependent processes, including neuronal polarity, neuritogenesis, synaptogenesis, and axonal transport. KIF2A-deficient neurons exhibit aberrant electrophysiological characteristics, neuronal connectivity, and function, leading to their loss. The role of KIF2A is not limited to development, as fully mature neurons require KIF2A for survival. Our results emphasize an additional function of KIF2A and help explain how its mutations lead to brain disorders.
Asunto(s)
Encefalopatías , Proteínas Represoras , Animales , Ratones , Proteínas Represoras/metabolismo , Cinesinas/genética , Microtúbulos/metabolismo , Neuronas/metabolismo , Encefalopatías/metabolismoRESUMEN
Neural repair is highly influenced by reactive astrocytes. Atypical cadherin Celsr2 regulates neuron development and axon regeneration, while its role in glial cells remains unexplored. In this study, we show that Celsr2 is highly expressed in spinal astrocytes of adult mice, and knockout of Celsr2 results in reactive astrocytes with longer protrusions preferentially orientated towards lesion borders in culture scratch assay and injured spinal cord, and elevation of total and active Cdc42 and Rac1 protein in western blots. Inactivation of Celsr2 enhances calcium influx in reactive astrocytes in time-lapse imaging. Morphological phenotypes of cultured Celsr2-/- astrocytes are rescued by Cdc42 or Rac1 inhibitors. Following spinal cord injury (SCI), Celsr2-/- mice exhibit smaller lesion cavity and glial scar, enhanced fiber regeneration, weaker microglial response, and improved functional recovery than control animals. Similar phenotypes are found in mice with conditional knockout of Celsr2 in astrocytes. In Celsr2-/- mice, astrocyte phenotype is changed and neuroinflammation is alleviated after injury. Inhibiting Cdc42/Rac1 activities compromises astrocyte polarization and the improvement of neural repair and functional recovery in Celsr2-/- mice with SCI. In conclusion, Celsr2 regulates morphological polarization and functional phenotype of reactive astrocytes and inactivating Celsr2 is a potential therapeutic strategy for neural repair.
Asunto(s)
Astrocitos , Traumatismos de la Médula Espinal , Ratones , Animales , Astrocitos/metabolismo , Axones/metabolismo , Regeneración Nerviosa/fisiología , Traumatismos de la Médula Espinal/metabolismo , Fenotipo , Cadherinas/metabolismoRESUMEN
The molecular mechanisms by which cilia orientation is coordinated within and between multi-ciliated cells (MCCs) are not fully understood. In the mouse oviduct, MCCs exhibit a characteristic basal body (BB) orientation and microtubule gradient along the tissue axis. The intracellular polarities were moderately maintained in cells lacking CELSR1 (cadherin EGF LAG seven-pass G-type receptor 1), a planar cell polarity (PCP) factor involved in tissue polarity regulation, although the intercellular coordination of the polarities was disrupted. However, CAMSAP3 (calmodulin-regulated spectrin-associated protein 3), a microtubule minus-end regulator, was found to be critical for determining the intracellular BB orientation. CAMSAP3 localized to the base of cilia in a polarized manner, and its mutation led to the disruption of intracellular coordination of BB orientation, as well as the assembly of microtubules interconnecting BBs, without affecting PCP factor localization. Thus, both CELSR1 and CAMSAP3 are responsible for BB orientation but in distinct ways; their cooperation should therefore be critical for generating functional multi-ciliated tissues.
Asunto(s)
Cadherinas , Cilios , Células Epiteliales , Proteínas Asociadas a Microtúbulos , Animales , Polaridad Celular , Femenino , Ratones , Oviductos , Receptores Acoplados a Proteínas GRESUMEN
Understanding new modulators of axon regeneration is central to neural repair. Our previous work demonstrated critical roles of atypical cadherin Celsr2 during neural development, including cilia organization, neuron migration and axon navigation. Here, we address its role in axon regeneration. We show that Celsr2 is highly expressed in both mouse and human spinal motor neurons. Celsr2 knockout promotes axon regeneration and fasciculation in mouse cultured spinal explants. Similarly, cultured Celsr2 mutant motor neurons extend longer neurites and larger growth cones, with increased expression of end-binding protein 3 and higher potassium-induced calcium influx. Mice with Celsr2 conditional knockout in spinal motor neurons do not exhibit any behavioural deficits; however, after branchial plexus injury, axon regeneration and functional forelimb locomotor recovery are significantly improved. Similarly, knockdown of CELSR2 using shRNA interference in cultured human spinal motor explants and motor neurons increases axonal fasciculation and growth. In mouse adult spinal cord after root avulsion, in mouse embryonic spinal cords, and in cultured human motor neurons, Celsr2 downregulation is accompanied by increased levels of GTP-bound Rac1 and Cdc42, and of JNK and c-Jun. In conclusion, Celsr2 negatively regulates motor axon regeneration and is a potential target to improve neural repair.
Asunto(s)
Fasciculación Axonal , Traumatismos de la Médula Espinal , Animales , Axones/metabolismo , Cadherinas , Humanos , Ratones , Neuronas Motoras/metabolismo , Regeneración Nerviosa , Médula Espinal , Traumatismos de la Médula Espinal/metabolismoRESUMEN
BACKG ROUND: Glioblastoma is an aggressive tumor that has a dismal prognosis even with multimodal treatment. However, some patients survive longer than expected. The objective of this study was to revisit patients diagnosed with glioblastoma according to the 2021 WHO classification and analyze clinical and molecular characteristics associated with long-term survival (LTS). METHODS: We retrospectively analyzed 120 IDH-wildtype glioblastomas operated on at our institution between 2013 and 2018. We divided them into LTS patients, surviving more than 3 years, and non-LTS patients, and then compared their features. Additionally, we performed DNA methylation-based brain tumor classification in LTS patients. RESULTS: Sixteen patients were long-term survivors. Age < 70 years, MGMT promoter methylation, extent of resection ≥ 95%, and administration of radiochemotherapy were associated with LTS (P = 0.005, P < 0.001, P = 0.048, and P = 0.008, respectively). In addition, when these factors were combined, the probability of LTS was 74% (95% CI: 62--84). The methylome analysis confirmed the diagnosis of glioblastoma in the majority of the tested LTS patients. Regarding subtypes, 29% of cases were mesenchymal (MES), 43% were RTK1, and 29% were RTK2. Interestingly, RTK1 and RTK2 cases tended to have longer overall survival than MES cases (P = 0.057). Moreover, the only tested LTS patient with an unmethylated MGMT promoter had an "adult-type diffuse high-grade glioma, IDH-wildtype, subtype E" rather than a glioblastoma. This tumor was characterized by multinucleated giant cells and a somatic mutation in POLE. CONCLUSIONS: We suggest that glioblastoma patients with a combination of favorable prognostic factors can achieve LTS in 74% of cases. In addition, methylome analysis is important to ascertain the type of glioma in LTS patients, especially when the MGMT promoter is unmethylated.
Asunto(s)
Neoplasias Encefálicas , Glioblastoma , Glioma , Adulto , Humanos , Anciano , Glioblastoma/terapia , Glioblastoma/tratamiento farmacológico , Estudios Retrospectivos , Glioma/genética , Pronóstico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/terapia , Neoplasias Encefálicas/patología , Metilación de ADN/genética , Isocitrato Deshidrogenasa/genética , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genéticaRESUMEN
Paralog factors are usually described as consolidating biological systems by displaying redundant functionality in the same cells. Here, we report that paralogs can also cooperate in distinct cell populations at successive stages of differentiation. In mouse embryonic spinal cord, motor neurons and V2 interneurons differentiate from adjacent progenitor domains that share identical developmental determinants. Therefore, additional strategies secure respective cell fate. In particular, Hb9 promotes motor neuron identity while inhibiting V2 differentiation, whereas Chx10 stimulates V2a differentiation while repressing motor neuron fate. However, Chx10 is not present at the onset of V2 differentiation and in other V2 populations. In the present study, we show that Vsx1, the single paralog of Chx10, which is produced earlier than Chx10 in V2 precursors, can inhibit motor neuron differentiation and promote V2 interneuron production. However, the single absence of Vsx1 does not impact on V2 fate consolidation, suggesting that lack of Vsx1 may be compensated by other factors. Nevertheless, Vsx1 cooperates with Chx10 to prevent motor neuron differentiation in early V2 precursors although these two paralog factors are not produced in the same cells. Hence, this study uncovers an original situation, namely labor division, wherein paralog genes cooperate at successive steps of neuronal development.
Asunto(s)
Proteínas del Ojo/genética , Proteínas de Homeodominio/genética , Interneuronas/fisiología , Neuronas Motoras/fisiología , Médula Espinal/fisiología , Factores de Transcripción/genética , Animales , Diferenciación Celular/genética , Línea Celular , Regulación del Desarrollo de la Expresión Génica/genética , Células HEK293 , Humanos , RatonesRESUMEN
Understanding the developmental steps that shape formation of the neuromuscular junction (NMJ) connecting motoneurons to skeletal muscle fibers is crucial. Wnt morphogens are key players in the formation of this specialized peripheral synapse, but their individual and collaborative functions and downstream pathways remain poorly understood at the NMJ. Here, we demonstrate through Wnt4 and Wnt11 gain-of-function studies in cell culture or in mice that Wnts enhance acetylcholine receptor (AChR) clustering and motor axon outgrowth. By contrast, loss of Wnt11 or Wnt-dependent signaling in vivo decreases AChR clustering and motor nerve terminal branching. Both Wnt4 and Wnt11 stimulate AChR mRNA levels and AChR clustering downstream of activation of the ß-catenin pathway. Strikingly, Wnt4 and Wnt11 co-immunoprecipitate with Vangl2, a core component of the planar cell polarity (PCP) pathway, which accumulates at embryonic NMJs. Moreover, mice bearing a Vangl2 loss-of-function mutation (loop-tail) exhibit fewer AChR clusters and overgrowth of motor axons bypassing AChR clusters. Together, our results provide genetic and biochemical evidence that Wnt4 and Wnt11 cooperatively contribute to mammalian NMJ formation through activation of both the canonical and Vangl2-dependent core PCP pathways.
Asunto(s)
Unión Neuromuscular/metabolismo , Transducción de Señal , Proteínas Wnt/metabolismo , Proteína Wnt4/metabolismo , Animales , Axones/metabolismo , Polaridad Celular , Embrión de Mamíferos/metabolismo , Espacio Extracelular/metabolismo , Ratones Endogámicos C57BL , Mutación/genética , Proteínas del Tejido Nervioso/metabolismo , Fenotipo , Receptores Colinérgicos/metabolismo , Sinapsis/metabolismoRESUMEN
Group I metabotropic glutamate receptors (mGluR) are involved in various forms of synaptic plasticity that are believed to underlie declarative memory. We previously showed that mGluR5 specifically activates channels containing TRPC1, an isoform of the canonical family of Transient Receptor Potential channels highly expressed in the CA1-3 regions of the hippocampus. Using a tamoxifen-inducible conditional knockout model, we show here that the acute deletion of the Trpc1 gene alters the extinction of spatial reference memory. mGluR-induced long-term depression, which is partially responsible for memory extinction, was impaired in these mice. Similar results were obtained in vitro and in vivo by inhibiting the channel by its most specific inhibitor, Pico145. Among the numerous known postsynaptic pathways activated by type I mGluR, we observed that the deletion of Trpc1 impaired the activation of ERK1/2 and the subsequent expression of Arc, an immediate early gene that plays a key role in AMPA receptors endocytosis and subsequent long-term depression.
Asunto(s)
Hipocampo/metabolismo , Canales Catiónicos TRPC/metabolismo , Animales , Depresión/genética , Depresión/metabolismo , Depresión/fisiopatología , Hipocampo/fisiología , Sistema de Señalización de MAP Quinasas/genética , Sistema de Señalización de MAP Quinasas/fisiología , Masculino , Ratones , Ratones Noqueados , Plasticidad Neuronal/genética , Plasticidad Neuronal/fisiología , Receptores AMPA/genética , Receptores AMPA/metabolismo , Receptores de Glutamato Metabotrópico/genética , Receptores de Glutamato Metabotrópico/metabolismo , Transducción de Señal/genética , Transducción de Señal/fisiología , Memoria Espacial/fisiología , Canales Catiónicos TRPC/genéticaRESUMEN
Cadherin EGF LAG seven-pass G-type receptors 1, 2 and 3 (CELSR1-3) form a family of three atypical cadherins with multiple functions in epithelia and in the nervous system. During the past decade, evidence has accumulated for a key role of CELSR1 in epithelial planar cell polarity (PCP), and for CELSR2 and CELSR3 in ciliogenesis and neural development, especially neuron migration and axon guidance in the central, peripheral and enteric nervous systems. Phenotypes in mutant mice indicate that CELSR proteins work in concert with FZD3 and FZD6, but several questions remain. Apart from PCP signaling pathways implicating CELSR1 that begin to be unraveled, little is known about other signals generated by CELSR2 and CELSR3. A crucial question concerns the putative ligands that trigger signaling, in particular what is the role of WNT factors. Another critical issue is the identification of novel intracellular pathways and effectors that relay and transmit signals in receptive cells? Answers to those questions should further our understanding of the role of those important molecules not only in development but also in regeneration and disease.
Asunto(s)
Cadherinas/química , Cadherinas/metabolismo , Animales , Orientación del Axón , Tipificación del Cuerpo , Encéfalo/fisiología , Cadherinas/genética , Humanos , Dominios ProteicosRESUMEN
Cajal-Retzius neurons (CRN) are the main source of Reelin in the marginal zone of the developing neocortex and hippocampus (HC). They also express the transcription factor p73 and are complemented by later-appearing GABAergic Reelin+ interneurons. The human dorsal HC forms at gestational week 10 (GW10), when it develops a rudimentary Ammonic plate and incipient dentate migration, although the dorsal hippocampal fissure (HF) remains shallow and contains few CRN. The dorsal HC transforms into the indusium griseum (IG), concurrently with the rostro-caudal appearance of the corpus callosum, by GW14-17. Dorsal and ventral HC merge at the site of the former caudal hem, which is located at the level of the future atrium of the lateral ventricle and closely connected with the choroid plexus. The ventral HC forms at GW11 in the temporal lobe. The ventral HF is wide open at GW14-16 and densely populated by large numbers of CRNs. These are in intimate contact with the meninges and meningeal blood vessels, suggesting signalling through diverse pathways. At GW17, the fissure deepens and begins to fuse, although it is still marked by p73/Reelin+ CRNs. The p73KO mouse illustrates the importance of p73 in CRN for HF formation. In the mutant, Tbr1/Reelin+ CRNs are born in the hem but do not leave it and subsequently disappear, so that the mutant cortex and HC lack CRN from the onset of corticogenesis. The HF is absent, which leads to profound architectonic alterations of the HC. To determine which p73 isoform is important for HF formation, isoform-specific TAp73- and DeltaNp73-deficient embryonic and early postnatal mice were examined. In both mutants, the number of CRNs was reduced, but each of their phenotypes was much milder than in the global p73KO mutant missing both isoforms. In the TAp73KO mice, the HF of the dorsal HC failed to form, but was present in the ventral HC. In the DeltaNp73KO mice, the HC had a mild patterning defect along with a shorter HF. Complex interactions between both isoforms in CRNs may contribute to their crucial activity in the developing brain.
Asunto(s)
Hipocampo/embriología , Proteína Tumoral p73/fisiología , Animales , Hipocampo/citología , Humanos , Lóbulo Límbico/embriología , Ratones Noqueados , Neuronas/fisiología , Proteína ReelinaRESUMEN
Vestibular hair cells of the inner ear are specialized receptors that detect mechanical stimuli from gravity and motion via the deflection of a polarized bundle of stereocilia located on their apical cell surfaces. The orientation of stereociliary bundles is coordinated between neighboring cells by core PCP proteins including the large adhesive G-protein coupled receptor Celsr1. We show that mice lacking Celsr1 have vestibular behavioral phenotypes including circling. In addition, we show that Celsr1 is asymmetrically distributed at cell boundaries between hair cells and neighboring supporting cells in the developing vestibular and auditory sensory epithelia. In the absence of Celsr1 the stereociliary bundles of vestibular hair cells are misoriented relative to their neighbors, a phenotype that is greatest in the cristae of the semicircular canals. Since horizontal semi-circular canal defects lead to circling in other mutant mouse lines, we propose that this PCP phenotype is the cellular basis of the circling behavior in Celsr1 mutants.
Asunto(s)
Polaridad Celular , Oído Interno/citología , Oído Interno/embriología , Células Ciliadas Vestibulares/citología , Receptores Acoplados a Proteínas G/metabolismo , Animales , Conducta Animal , Oído Interno/metabolismo , Epitelio/metabolismo , Eliminación de Gen , Ratones Noqueados , Órgano Espiral/citología , Órgano Espiral/embriología , Órgano Espiral/metabolismo , Fenotipo , Transducción de Señal , Estereocilios/metabolismoRESUMEN
Planar cell polarity (PCP) is complementary to the intrinsic polarization of single cells and refers to the global coordination of cell behaviour in the plane of a tissue and, by extension, to the signalling pathways that control it. PCP is most evident in cell sheets, and research into PCP was for years confined to studies in Drosophila melanogaster. However, PCP has more recently emerged as an important phenomenon in vertebrates, in which it regulates various developmental processes and is associated with multiple disorders. In particular, core PCP genes are crucial for the development and function of the nervous system. They are involved in neural tube closure, ependymal polarity, neuronal migration, dendritic growth and axon guidance.
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Polaridad Celular/genética , Proteínas de la Membrana/genética , Sistema Nervioso/citología , Sistema Nervioso/metabolismo , Animales , Tipificación del Cuerpo/genética , Movimiento Celular/genética , Proteínas de la Membrana/metabolismo , Modelos Biológicos , Sistema Nervioso/embriología , Sistema Nervioso/crecimiento & desarrollo , Neuronas/citología , Neuronas/fisiología , Transducción de Señal/genéticaRESUMEN
A unique population of cells, called "lot cells," circumscribes the path of the lateral olfactory tract (LOT) in the rodent brain and acts to restrict its position at the lateral margin of the telencephalon. Lot cells were believed to originate in the dorsal pallium (DP). We show that Lhx2 null mice that lack a DP show a significant increase in the number of mGluR1/lot cells in the piriform cortex, indicating a non-DP origin of these cells. Since lot cells present common developmental features with Cajal-Retzius (CR) cells, we analyzed Wnt3a- and Dbx1-reporter mouse lines and found that mGluR1/lot cells are not generated in the cortical hem, ventral pallium, or septum, the best characterized sources of CR cells. Finally, we identified a novel origin for the lot cells by combining in utero electroporation assays and histochemical characterization. We show that mGluR1/lot cells are specifically generated in the lateral thalamic eminence and that they express mitral cell markers, although a minority of them express ΔNp73 instead. We conclude that most mGluR1/lot cells are prospective mitral cells migrating to the accessory olfactory bulb (OB), whereas mGluR1+, ΔNp73+ cells are CR cells that migrate through the LOT to the piriform cortex and the OB.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica/fisiología , Receptores de Glutamato Metabotrópico/metabolismo , Células Madre/fisiología , Tálamo/citología , Tálamo/metabolismo , Animales , Movimiento Celular , Células Cultivadas , Embrión de Mamíferos , Femenino , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Proteínas con Homeodominio LIM/genética , Proteínas con Homeodominio LIM/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Ratones Transgénicos , Proteínas del Tejido Nervioso/metabolismo , Neurogénesis/fisiología , Embarazo , Receptores de Glutamato Metabotrópico/genética , Proteínas de Dominio T Box/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteína Tumoral p73/genética , Proteína Tumoral p73/metabolismoRESUMEN
The caudal migration of facial branchiomotor (FBM) neurons from rhombomere (r) 4 to r6 in the hindbrain is an excellent model to study neuronal migration mechanisms. Although several Wnt/Planar Cell Polarity (PCP) components are required for FBM neuron migration, only Celsr1, an atypical cadherin, regulates the direction of migration in mice. In Celsr1 mutants, a subset of FBM neurons migrates rostrally instead of caudally. Interestingly, Celsr1 is not expressed in the migrating FBM neurons, but rather in the adjacent floor plate and adjoining ventricular zone. To evaluate the contribution of different expression domains to neuronal migration, we conditionally inactivated Celsr1 in specific cell types. Intriguingly, inactivation of Celsr1 in the ventricular zone of r3-r5, but not in the floor plate, leads to rostral migration of FBM neurons, greatly resembling the migration defect of Celsr1 mutants. Dye fill experiments indicate that the rostrally-migrated FBM neurons in Celsr1 mutants originate from the anterior margin of r4. These data suggest strongly that Celsr1 ensures that FBM neurons migrate caudally by suppressing molecular cues in the rostral hindbrain that can attract FBM neurons.
Asunto(s)
Movimiento Celular/fisiología , Nervio Facial/embriología , Neurogénesis/fisiología , Receptores Acoplados a Proteínas G/metabolismo , Rombencéfalo/embriología , Animales , Nervio Facial/metabolismo , Regulación del Desarrollo de la Expresión Génica , Ratones , Ratones Noqueados , Neuronas Motoras/citología , Receptores Acoplados a Proteínas G/genéticaRESUMEN
The oviduct is an important organ in reproduction where fertilization occurs, and through which the fertilized eggs are carried to the uterus in mammals. This organ is highly polarized, where the epithelium forms longitudinal folds along the ovary-uterus axis, and the epithelial multicilia beat towards the uterus to transport the ovulated ova. Here, we analyzed the postnatal development of mouse oviduct and report that multilevel polarities of the oviduct are regulated by a planar cell polarity (PCP) gene, Celsr1. In the epithelium, Celsr1 is concentrated in the specific cellular boundaries perpendicular to the ovary-uterus axis from postnatal day 2. We found a new feature of cellular polarity in the oviduct - the apical surface of epithelial cells is elongated along the ovary-uterus axis. In Celsr1-deficient mice, the ciliary motion is not orchestrated along the ovary-uterus axis and the transport ability of beating cilia is impaired. Epithelial cells show less elongation and randomized orientation, and epithelial folds show randomized directionality and ectopic branches in the mutant. Our mosaic analysis suggests that the geometry of epithelial cells is primarily regulated by Celsr1 and as a consequence the epithelial folds are aligned. Taken together, we reveal the characteristics of the multilevel polarity formation processes in the mouse oviduct epithelium and suggest a novel function of the PCP pathway for proper tissue morphogenesis.
Asunto(s)
Polaridad Celular/fisiología , Células Epiteliales/fisiología , Organogénesis/fisiología , Oviductos/embriología , Receptores Acoplados a Proteínas G/metabolismo , Animales , Bromodesoxiuridina , Polaridad Celular/genética , Forma de la Célula/fisiología , Cilios/fisiología , Cartilla de ADN/genética , Femenino , Fluorescencia , Inmunohistoquímica , Ratones , Ratones Noqueados , Microscopía Electrónica de Transmisión , Microesferas , Oviductos/ultraestructura , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
In the nervous system, cilia dysfunction perturbs the circulation of the cerebrospinal fluid, thus affecting neurogenesis and brain homeostasis. A role for planar cell polarity (PCP) signaling in the orientation of cilia (rotational polarity) and ciliogenesis is established. However, whether and how PCP regulates cilia positioning in the apical domain (translational polarity) in radial progenitors and ependymal cells remain unclear. By analysis of a large panel of mutant mice, we show that two PCP signals are operating in ciliated cells. The first signal, controlled by cadherin, EGF-like, laminin G-like, seven-pass, G-type receptor (Celsr) 2, Celsr3, Frizzled3 (Fzd3) and Van Gogh like2 (Vangl2) organizes multicilia in individual cells (single-cell polarity), whereas the second signal, governed by Celsr1, Fzd3, and Vangl2, coordinates polarity between cells in both radial progenitors and ependymal cells (tissue polarity). Loss of either of these signals is associated with specific defects in the cytoskeleton. Our data reveal unreported functions of PCP and provide an integrated view of planar polarization of the brain ciliated cells.
Asunto(s)
Polaridad Celular/fisiología , Citoesqueleto/metabolismo , Epéndimo/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neurogénesis/fisiología , Transducción de Señal/fisiología , Animales , Cilios/genética , Cilios/metabolismo , Citoesqueleto/genética , Epéndimo/citología , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/genéticaRESUMEN
Celsr3 and Fzd3, members of "core planar cell polarity" (PCP) genes, were shown previously to control forebrain axon guidance and wiring by acting in axons and/or guidepost cells. Here, we show that Celsr2 acts redundantly with Celsr3, and that their combined mutation mimics that of Fzd3. The phenotypes generated upon inactivation of Fzd3 in different forebrain compartments are similar to those in conditional Celsr2-3 mutants, indicating that Fzd3 and Celsr2-3 act in the same population of cells. Inactivation of Celsr2-3 or Fzd3 in thalamus does not affect forebrain wiring, and joint inactivation in cortex and thalamus adds little to cortical inactivation alone in terms of thalamocortical projections. On the other hand, joint inactivation perturbs strongly the formation of the barrel field, which is unaffected upon single cortical or thalamic inactivation, indicating a role for interactions between thalamic axons and cortical neurons in cortical arealization. Unexpectedly, forebrain wiring is normal in mice defective in Vangl1 and Vangl2, showing that, contrary to epithelial PCP, axon guidance can be Vangl independent in some contexts. Our results suggest that Celsr2-3 and Fzd3 regulate axonal navigation in the forebrain by using mechanisms different from classical epithelial PCP, and require interacting partners other than Vangl1-2 that remain to be identified.
Asunto(s)
Cadherinas/metabolismo , Proteínas Portadoras/metabolismo , Receptores Frizzled/metabolismo , Proteínas de la Membrana/metabolismo , Red Nerviosa/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Prosencéfalo/metabolismo , Receptores de Superficie Celular/metabolismo , Animales , Axones/metabolismo , Corteza Cerebral/metabolismo , Silenciador del Gen , Integrasas/metabolismo , Ratones , Mutación/genética , Fenotipo , Tálamo/metabolismoRESUMEN
The striatum is a large brain nucleus with an important role in the control of movement and emotions. Medium spiny neurons (MSNs) are striatal output neurons forming prominent descending axon tracts that target different brain nuclei. However, how MSN axon tracts in the forebrain develop remains poorly understood. Here, we implicate the Wnt binding receptor Frizzled3 in several uncharacterized aspects of MSN pathway formation [i.e., anterior-posterior guidance of MSN axons in the striatum and their subsequent growth into the globus pallidus (GP), an important (intermediate) target]. In Frizzled3 knock-out mice, MSN axons fail to extend along the anterior-posterior axis of the striatum, and many do not reach the GP. Wnt5a acts as an attractant for MSN axons in vitro, is expressed in a posterior high, anterior low gradient in the striatum, and Wnt5a knock-out mice phenocopy striatal anterior-posterior defects observed in Frizzled3 mutants. This suggests that Wnt5a controls anterior-posterior guidance of MSN axons through Frizzled3. Axons that reach the GP in Frizzled3 knock-out mice fail to enter this structure. Surprisingly, entry of MSN axons into the GP non-cell-autonomously requires Frizzled3, and our data suggest that GP entry may be contingent on the correct positioning of "corridor" guidepost cells for thalamocortical axons by Frizzled3. Together, these data dissect MSN pathway development and reveal (non)cell-autonomous roles for Frizzled3 in MSN axon guidance. Further, they are the first to identify a gene that provides anterior-posterior axon guidance in a large brain nucleus and link Frizzled3 to corridor cell development. SIGNIFICANCE STATEMENT: Striatal axon pathways mediate complex physiological functions and are an important therapeutic target, underscoring the need to define how these connections are established. Remarkably, the molecular programs regulating striatal pathway development remain poorly characterized. Here, we determine the embryonic ontogeny of the two main striatal pathways (striatonigral and striatopallidal) and identify novel (non)cell-autonomous roles for the axon guidance receptor Frizzled3 in uncharacterized aspects of striatal pathway formation (i.e., anterior-posterior axon guidance in the striatum and axon entry into the globus pallidus). Further, our results link Frizzled3 to corridor guidepost cell development and suggest that an abnormal distribution of these cells has unexpected, widespread effects on the development of different axon tracts (i.e., striatal and thalamocortical axons).