RESUMEN
A fundamental challenge in developing treatments for autism spectrum disorders is the heterogeneity of the condition. More than one hundred genetic mutations confer high risk for autism, with each individual mutation accounting for only a small fraction of cases1-3. Subsets of risk genes can be grouped into functionally related pathways, most prominently those involving synaptic proteins, translational regulation, and chromatin modifications. To attempt to minimize this genetic complexity, recent therapeutic strategies have focused on the neuropeptides oxytocin and vasopressin4-6, which regulate aspects of social behaviour in mammals7. However, it is unclear whether genetic risk factors predispose individuals to autism as a result of modifications to oxytocinergic signalling. Here we report that an autism-associated mutation in the synaptic adhesion molecule Nlgn3 results in impaired oxytocin signalling in dopaminergic neurons and in altered behavioural responses to social novelty tests in mice. Notably, loss of Nlgn3 is accompanied by a disruption of translation homeostasis in the ventral tegmental area. Treatment of Nlgn3-knockout mice with a new, highly specific, brain-penetrant inhibitor of MAP kinase-interacting kinases resets the translation of mRNA and restores oxytocin signalling and social novelty responses. Thus, this work identifies a convergence between the genetic autism risk factor Nlgn3, regulation of translation, and oxytocinergic signalling. Focusing on such common core plasticity elements might provide a pragmatic approach to overcoming the heterogeneity of autism. Ultimately, this would enable mechanism-based stratification of patient populations to increase the success of therapeutic interventions.
Asunto(s)
Trastorno Autístico/metabolismo , Trastorno Autístico/psicología , Modelos Animales de Enfermedad , Oxitocina/metabolismo , Conducta Social , Animales , Moléculas de Adhesión Celular Neuronal/deficiencia , Moléculas de Adhesión Celular Neuronal/genética , Factor 4E Eucariótico de Iniciación/metabolismo , Masculino , Proteínas de la Membrana/deficiencia , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Fosforilación/efectos de los fármacos , Biosíntesis de Proteínas/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de Señal/efectos de los fármacos , Área Tegmental Ventral/citología , Área Tegmental Ventral/efectos de los fármacosRESUMEN
OBJECTIVE: Temporal lobe epilepsy (TLE) is characterized by recurrent seizures generated in the limbic system, particularly in the hippocampus. In TLE, recurrent mossy fiber sprouting from dentate gyrus granule cells (DGCs) crea an aberrant epileptogenic network between DGCs which operates via ectopically expressed GluK2/GluK5-containing kainate receptors (KARs). TLE patients are often resistant to anti-seizure medications and suffer significant comorbidities; hence, there is an urgent need for novel therapies. Previously, we have shown that GluK2 knockout mice are protected from seizures. This study aims at providing evidence that downregulating KARs in the hippocampus using gene therapy reduces chronic epileptic discharges in TLE. METHODS: We combined molecular biology and electrophysiology in rodent models of TLE and in hippocampal slices surgically resected from patients with drug-resistant TLE. RESULTS: Here, we confirmed the translational potential of KAR suppression using a non-selective KAR antagonist that markedly attenuated interictal-like epileptiform discharges (IEDs) in TLE patient-derived hippocampal slices. An adeno-associated virus (AAV) serotype-9 vector expressing anti-grik2 miRNA was engineered to specifically downregulate GluK2 expression. Direct delivery of AAV9-anti grik2 miRNA into the hippocampus of TLE mice led to a marked reduction in seizure activity. Transduction of TLE patient hippocampal slices reduced levels of GluK2 protein and, most importantly, significantly reduced IEDs. INTERPRETATION: Our gene silencing strategy to knock down aberrant GluK2 expression demonstrates inhibition of chronic seizure in a mouse TLE model and IEDs in cultured slices derived from TLE patients. These results provide proof-of-concept for a gene therapy approach targeting GluK2 KARs for drug-resistant TLE patients. ANN NEUROL 2023;94:745-761.
Asunto(s)
Epilepsia Refractaria , Epilepsia del Lóbulo Temporal , MicroARNs , Humanos , Ratones , Animales , Epilepsia del Lóbulo Temporal/terapia , Lóbulo Temporal , Hipocampo , Epilepsia Refractaria/genética , Epilepsia Refractaria/terapia , ConvulsionesRESUMEN
Despite the pivotal functions of the NMDA receptor (NMDAR) for neural circuit development and synaptic plasticity, the molecular mechanisms underlying the dynamics of NMDAR trafficking are poorly understood. The cell adhesion molecule neuroligin-1 (NL1) modifies NMDAR-dependent synaptic transmission and synaptic plasticity, but it is unclear whether NL1 controls synaptic accumulation or function of the receptors. Here, we provide evidence that NL1 regulates the abundance of NMDARs at postsynaptic sites. This function relies on extracellular, NL1 isoform-specific sequences that facilitate biochemical interactions between NL1 and the NMDAR GluN1 subunit. Our work uncovers NL1 isoform-specific cis-interactions with ionotropic glutamate receptors as a key mechanism for controlling synaptic properties.
Asunto(s)
Moléculas de Adhesión Celular Neuronal/metabolismo , Plasticidad Neuronal/fisiología , Receptores Ionotrópicos de Glutamato/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Sinapsis/fisiología , Transmisión Sináptica/fisiología , Análisis de Varianza , Animales , Western Blotting , Maleato de Dizocilpina , Inmunoprecipitación , Microscopía Confocal , Microscopía Inmunoelectrónica , Ratas , Estadísticas no ParamétricasRESUMEN
The immunoreceptor-associated protein CD3ζ is known for its role in immunity and has also been implicated in neuronal development and synaptic plasticity. However, the mechanism by which CD3ζ regulates synaptic transmission remains unclear. In this study, we showed that mice lacking CD3ζ exhibited defects in spatial learning and memory as examined by the Barnes maze and object location memory tasks. Given that peripheral T cells have been shown to support cognitive functions and neural plasticity, we generated CD3ζ(-/-) mice in which the peripheral T cells were repopulated to a normal level by syngeneic bone marrow transplantation. Using this approach, we showed that T-cell replenishment in CD3ζ(-/-) mice did not restore spatial memory defects, suggesting that the cognitive deficits in CD3ζ(-/-) mice were most likely mediated through a T-cell-independent mechanism. In support of this idea, we showed that CD3ζ proteins were localized to glutamatergic postsynaptic sites, where they interacted with the NMDAR subunit GluN2A. Loss of CD3ζ in brain decreased GluN2A-PSD95 association and GluN2A synaptic localization. This effect was accompanied by a reduced interaction of GluN2A with the key NMDAR downstream signaling protein calcium/calmodulin-dependent protein kinase II (CaMKII). Using the glycine-induced, NMDA-dependent form of chemical long-term potentiation (LTP) in cultured cortical neurons, we showed that CD3ζ was required for activity-dependent CaMKII autophosphorylation and for the synaptic recruitment of the AMPAR subunit GluA1. Together, these results support the model that the procognitive function of CD3ζ may be mediated through its involvement in the NMDAR downstream signaling pathway leading to CaMKII-dependent LTP induction.
Asunto(s)
Complejo CD3/metabolismo , Trastornos de la Memoria/genética , Receptores AMPA/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Linfocitos T/patología , Animales , Trasplante de Médula Ósea , Complejo CD3/genética , Células Cultivadas , Corteza Cerebral/citología , Modelos Animales de Enfermedad , Embrión de Mamíferos , Regulación de la Expresión Génica/genética , Glicina/farmacología , Antígenos Comunes de Leucocito/genética , Aprendizaje por Laberinto , Trastornos de la Memoria/fisiopatología , Trastornos de la Memoria/cirugía , Memoria a Corto Plazo/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/citología , Neuronas/efectos de los fármacos , Reconocimiento en Psicología/fisiologíaRESUMEN
Autism is a developmental disorder characterised by a high heterogeneity of clinical diagnoses and genetic associations. This heterogeneity is a challenge for the identification of the pathophysiology of the disease and for the development of new therapeutic strategies. New conceptual approaches are being used to try to challenge this complexity and gene cluster analysis studies suggest that the pathophysiology of autism is associated with a dysregulation of specific cellular mechanisms. This review will present the experimental evidence for a convergence of synaptic pathophysiology between syndromic and non-syndromic forms of autism, grouped under the generic term of autism spectrum disorders. In particular I will highlight the results from genetic mouse models identifying a convergence of dysregulation of the synaptic type I metabotropic glutamate receptor pathway in mouse models for autism spectrum disorders. These results help to build a new conceptual framework for the study of the synaptic phenotype of autism, which is important for the identification of new therapeutic strategies.
Asunto(s)
Trastornos Generalizados del Desarrollo Infantil/fisiopatología , Sinapsis/metabolismo , Animales , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Moléculas de Adhesión Celular Neuronal/genética , Moléculas de Adhesión Celular Neuronal/metabolismo , Trastornos Generalizados del Desarrollo Infantil/genética , Trastornos Generalizados del Desarrollo Infantil/metabolismo , Humanos , Receptores de Glutamato Metabotrópico/genética , Receptores de Glutamato Metabotrópico/metabolismo , Sinapsis/fisiologíaRESUMEN
Recent studies have highlighted the key role of the immune protein CD3ζ in the maturation of neuronal circuits in the CNS. Yet, the upstream signals that might recruit and activate CD3ζ in neurons are still unknown. In this study, we show that CD3ζ functions early in neuronal development and we identify ephrinA1-dependent EphA4 receptor activation as an upstream regulator of CD3ζ. When newly born neurons are still spherical, before neurite extension, we found a transient CD3ζ aggregation at the cell periphery matching the initiation site of the future neurite. This accumulation of CD3ζ correlated with a stimulatory effect on filopodia extension via a Rho-GEF Vav2 pathway and a repression of neurite outgrowth. Conversely, cultured neurons lacking CD3ζ isolated from CD3ζ(-/-) mice showed a decreased number of filopodia and an enhanced neurite number. Stimulation with ephrinA1 induces the translocation of both CD3ζ and its activated effector molecules, ZAP-70/Syk tyrosine kinases, to EphA4 receptor clusters. EphrinA1-induced growth cone collapse was abrogated in CD3ζ(-/-) neurons and was markedly reduced by ZAP-70/Syk inhibition. Moreover, ephrinA1-induced ZAP-70/Syk activation was inhibited in CD3ζ(-/-) neurons. Altogether, our data suggest that CD3ζ mediates the ZAP-70/Syk kinase activation triggered by ephrinA-activated pathway to regulate early neuronal morphogenesis.
Asunto(s)
Complejo CD3/metabolismo , Efrinas/metabolismo , Neuritas/fisiología , Neuronas/fisiología , Transducción de Señal/fisiología , Proteína Tirosina Quinasa ZAP-70/metabolismo , Animales , Animales Recién Nacidos , Encéfalo/citología , Complejo CD3/genética , Células COS , Células Cultivadas , Chlorocebus aethiops , Embrión de Mamíferos , Efrinas/genética , Efrinas/farmacología , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Inmunoprecipitación/métodos , Ratones , Ratones Noqueados , Células-Madre Neurales , Neuronas/citología , Neuronas/efectos de los fármacos , Seudópodos/efectos de los fármacos , Seudópodos/fisiología , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Factores de Tiempo , Transfección/métodos , Tubulina (Proteína)/metabolismo , Proteína Tirosina Quinasa ZAP-70/genéticaRESUMEN
The regulation of neuronal soma size is essential for appropriate brain circuit function and its dysregulation is associated with several neurodevelopmental disorders. A defect in the dendritic growth and elaboration of motor neocortical pyramidal neurons in neonates lacking neuregulin-4 (NRG4) has previously been reported. In this study, we investigated whether the loss of NRG4 causes further morphologic defects that are specific to these neurons. We analyzed the soma size of pyramidal neurons of layer (L)2/3 and L5 of the motor cortex and a subpopulation of multipolar interneurons in this neocortical region in Nrg4+/+ and Nrg4-/- mice. There were significant decreases in pyramidal neuron soma size in Nrg4-/- mice compared with Nrg4+/+ littermates at all stages studied [postnatal day (P)10, P30, and P60]. The reduction was especially marked at P10 and in L5 pyramidal neurons. Soma size was not significantly different for multipolar interneurons at any age. This in vivo phenotype was replicated in pyramidal neurons cultured from Nrg4-/- mice and was rescued by NRG treatment. Analysis of a public single-cell RNA sequencing repository revealed discrete Nrg4 and Erbb4 expression in subpopulations of L5 pyramidal neurons, suggesting that the observed defects were due in part to loss of autocrine Nrg4/ErbB4 signaling. The pyramidal phenotype in the motor cortex of Nrg4-/- mice was associated with a lack of Rotarod test improvement in P60 mice, suggesting that absence of NRG4 causes alterations in motor performance.
Asunto(s)
Corteza Motora , Neurregulinas/genética , Neuronas/citología , Células Piramidales/citología , Animales , Ratones , Ratones Noqueados , Corteza Motora/metabolismoRESUMEN
Autism Spectrum Disorders (ASD) are characterized by heterogeneity both in their presentation and their genetic aetiology. In order to discover points of convergence common to different cases of ASD, attempts were made to identify the biological pathways genes associated with ASD contribute to. Many of these genes were found to play a role in neuronal and synaptic development and function. Among these genes are FMR1, CYFIP1 and NLGN3, all present at the synapse and reliably linked to ASD. In this review, we evaluate the evidence for the contribution of these genes to the same biological pathway responsible for the regulation of structural and physiological plasticity. Alterations in dendritic spine density and turnover, as well as long-term depression (LTD), were found in mouse models of mutations of all three genes. This overlap in the phenotypes associated with these mouse models likely arises from the molecular interaction between the protein products of FMR1, CYFIP1, and NLG3. A number of other proteins linked to ASD are also likely to participate in these pathways, resulting in further downstream effects. Overall, a synaptic pathway centered around FMR1, CYFIP1, and NLG3 is likely to contribute to the phenotypes associated with structural and physiological plasticity characteristic of ASD.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Trastorno del Espectro Autista , Moléculas de Adhesión Celular Neuronal/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Animales , Trastorno del Espectro Autista/genética , Proteínas Portadoras , Modelos Animales de Enfermedad , Humanos , Ratones , Plasticidad Neuronal , FenotipoRESUMEN
The etiology of autism spectrum disorders (ASDs) is a complex combination of genetic and environmental factors. Neuroligin3, a synaptic adhesion protein, and cytoplasmic FMR1 interacting protein 1 (CYFIP1), a regulator of protein translation and actin polymerization, are two proteins associated with ASDs that interact in neurons in vivo Here, we investigated the role of the Neuroligin3/CYFIP1 pathway in behavioral functioning and synapse formation in mice and found that it contributes to motor learning and synapse formation in males. Similar investigation in female mice revealed an absence of such phenotypes, suggesting that females are protected against mutations affecting this pathway. Previously, we showed that the social environment influences the behavior of male mice. We extended this finding and found that the transcriptome of wild-type mice housed with their mutant littermates, lacking Neuroligin3, differed from the transcriptome of wild-type mice housed together. Altogether, these results identify the role of the Neuroligin3/CYFIP1 pathway in male mouse behavior and highlight its sensitivity to social environment.
Asunto(s)
Trastorno del Espectro Autista , Animales , Trastorno del Espectro Autista/genética , Modelos Animales de Enfermedad , Femenino , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas , Fenotipo , Conducta Social , Medio SocialRESUMEN
Deletions in the 15q11.2 region of the human genome are associated with neurobehavioral deficits, and motor development delay, as well as in some cases, symptoms of autism or schizophrenia. The cytoplasmic FMRP-interacting protein 1 (CYFIP1) is one of the four genes contained within this locus and has been associated with other genetic forms of autism spectrum disorders (ASD). In mice, Cyfip1 haploinsufficiency leads to alteration of dendritic spine morphology and defects in synaptic plasticity, two pathophysiological hallmarks of mouse models of ASD. At the behavioral level, however, Cyfip1 haploinsufficiency leads to minor phenotypes, not directly relevant for 15q11.2 deletion syndrome or ASD. A fundamental question is whether neuronal phenotypes caused by the mutation of Cyfip1 are relevant for the human condition. Here, we describe a synaptic cluster of ASD-associated proteins centered on CYFIP1 and the adhesion protein Neuroligin-3. Cyfip1 haploinsufficiency in mice led to decreased dendritic spine density and stability associated with social behavior and motor learning phenotypes. Behavioral training early in development resulted in alleviating the motor learning deficits caused by Cyfip1 haploinsufficiency. Altogether, these data provide new insight into the neuronal and behavioral phenotypes caused by Cyfip1 mutation and proof-of-concept for the development of a behavioral therapy to treat phenotypes associated with 15q11.2 syndromes and ASD.
Asunto(s)
Trastorno del Espectro Autista/genética , Haploinsuficiencia/genética , Discapacidad Intelectual/genética , Proteínas del Tejido Nervioso/genética , Conducta Social , Proteínas Adaptadoras Transductoras de Señales , Animales , Moléculas de Adhesión Celular Neuronal/metabolismo , Aberraciones Cromosómicas , Cromosomas Humanos Par 15/genética , Espinas Dendríticas/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/metabolismo , Fenotipo , Distribución AleatoriaRESUMEN
In most mammals, including humans, the postnatal acquisition of normal social and nonsocial behavior critically depends on interactions with peers. Here we explore the possibility that mixed-group housing of mice carrying a deletion of Nlgn3, a gene associated with autism spectrum disorders, and their wild-type littermates induces changes in each other's behavior. We have found that, when raised together, male Nlgn3 knockout mice and their wild-type littermates displayed deficits in sociability. Moreover, social submission in adult male Nlgn3 knockout mice correlated with an increase in their anxiety. Re-expression of Nlgn3 in parvalbumin-expressing cells in transgenic animals rescued their social behavior and alleviated the phenotype of their wild-type littermates, further indicating that the social behavior of Nlgn3 knockout mice has a direct and measurable impact on wild-type animals' behavior. Finally, we showed that, unlike male mice, female mice lacking Nlgn3 were insensitive to their peers' behavior but modified the social behavior of their littermates. Altogether, our findings show that the environment is a critical factor in the development of behavioral phenotypes in transgenic and wild-type mice. In addition, these results reveal that the social environment has a sexually dimorphic effect on the behavior of mice lacking Nlgn3, being more influential in males than females.
Asunto(s)
Moléculas de Adhesión Celular Neuronal/deficiencia , Proteínas de la Membrana/deficiencia , Proteínas del Tejido Nervioso/deficiencia , Caracteres Sexuales , Conducta Social , Vocalización Animal/fisiología , Animales , Animales Recién Nacidos , Moléculas de Adhesión Celular Neuronal/genética , Familia 2 del Citocromo P450/metabolismo , Conducta Exploratoria/fisiología , Femenino , Regulación de la Expresión Génica/genética , Regulación del Desarrollo de la Expresión Génica/genética , Masculino , Aprendizaje por Laberinto , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mutación/genética , Proteínas del Tejido Nervioso/genética , Parvalbúminas/genética , Parvalbúminas/metabolismo , ARN Mensajero/metabolismo , Testosterona/orinaRESUMEN
Immune signaling and neuroinflammatory mediators have recently emerged as influential variables that regulate neural precursor/stem cell (NPC) behavior and function. In this study, we investigated whether the signaling adaptor protein CD3ζ, a transmembrane protein involved in T cell differentiation and function and recently shown to regulate neuronal development in the central nervous system (CNS), may have a role in NPC differentiation. We analyzed the expression profile of CD3ζ in embryonic rat brain during neurogenic periods and in neurosphere-derived neural cells, and we investigated the action of CD3ζ on cell differentiation. We found that CD3ζ expression coincided with neuronal commitment, but its forced expression in NPCs prevented the production of neurons and oligodendrocytes, but not astroglial cells. This blockade of neuronal differentiation was operated through an ITAM-independent mechanism, but required the Asp36 of the CD3ζ transmembrane domain involved in membrane receptor interaction. Together, our findings show that ectopic CD3ζ expression in NPCs impaired their normal cell-fate specification and suggest that variations of CD3ζ expression in the developing CNS might result in neurodevelopmental anomalies.
Asunto(s)
Complejo CD3/fisiología , Linaje de la Célula , Regulación del Desarrollo de la Expresión Génica , Proteínas del Tejido Nervioso/fisiología , Células-Madre Neurales/enzimología , Neurogénesis/fisiología , Sustitución de Aminoácidos , Animales , Complejo CD3/biosíntesis , Complejo CD3/química , Complejo CD3/genética , Supervivencia Celular , Células Cultivadas/enzimología , Inducción Enzimática , Femenino , Edad Gestacional , Hipocampo/citología , Hipocampo/embriología , Microscopía Fluorescente , Mutagénesis Sitio-Dirigida , Proteínas del Tejido Nervioso/biosíntesis , Proteínas del Tejido Nervioso/química , Proteínas del Tejido Nervioso/genética , Células-Madre Neurales/patología , Neuronas/citología , Neuronas/enzimología , Bulbo Olfatorio/embriología , Bulbo Olfatorio/enzimología , Embarazo , Ratas , Ratas Sprague-Dawley , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/fisiología , TransfecciónRESUMEN
The genetic heterogeneity of autism poses a major challenge for identifying mechanism-based treatments. A number of rare mutations are associated with autism, and it is unclear whether these result in common neuronal alterations. Monogenic syndromes, such as fragile X, include autism as one of their multifaceted symptoms and have revealed specific defects in synaptic plasticity. We discovered an unexpected convergence of synaptic pathophysiology in a nonsyndromic form of autism with those in fragile X syndrome. Neuroligin-3 knockout mice (a model for nonsyndromic autism) exhibited disrupted heterosynaptic competition and perturbed metabotropic glutamate receptor-dependent synaptic plasticity, a hallmark of fragile X. These phenotypes could be rescued by reexpression of neuroligin-3 in juvenile mice, highlighting the possibility of reverting neuronal circuit alterations in autism after the completion of development.
Asunto(s)
Trastorno Autístico/fisiopatología , Síndrome del Cromosoma X Frágil/fisiopatología , Plasticidad Neuronal , Sinapsis/fisiología , Animales , Moléculas de Adhesión Celular Neuronal/genética , Moléculas de Adhesión Celular Neuronal/metabolismo , Modelos Animales de Enfermedad , Síndrome del Cromosoma X Frágil/genética , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Red Nerviosa/metabolismo , Red Nerviosa/fisiopatología , Red Nerviosa/ultraestructura , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Sinapsis/metabolismo , Sinapsis/ultraestructuraRESUMEN
A novel idea is emergxsing that a large molecular repertoire is common to the nervous and immune systems, which might reflect the existence of novel neuronal functions for immune molecules in the brain. Here, we show that the transmembrane adaptor signaling protein CD3zeta, first described in the immune system, has a previously uncharacterized role in regulating neuronal development. Biochemical and immunohistochemical analyses of the rat brain and cultured neurons showed that CD3zeta is mainly expressed in neurons. Distribution of CD3zeta in developing cultured hippocampal neurons, as determined by immunofluorescence, indicates that CD3zeta is preferentially associated with the somatodendritic compartment as soon as the dendrites initiate their differentiation. At this stage, CD3zeta was selectively concentrated at dendritic filopodia and growth cones, actin-rich structures involved in neurite growth and patterning. siRNA-mediated knockdown of CD3zeta in cultured neurons or overexpression of a loss-of-function CD3zeta mutant lacking the tyrosine phosphorylation sites in the immunoreceptor tyrosine-based activation motifs (ITAMs) increased dendritic arborization. Conversely, activation of endogenous CD3zeta by a CD3zeta antibody reduced the size of the dendritic arbor. Altogether, our findings reveal a novel role for CD3zeta in the nervous system, suggesting its contribution to dendrite development through ITAM-based mechanisms.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Complejo CD3/metabolismo , Dendritas/metabolismo , Actinas/metabolismo , Animales , Anticuerpos/farmacología , Encéfalo/citología , Encéfalo/metabolismo , Células COS , Calcio/metabolismo , Células Cultivadas , Chlorocebus aethiops , Dendritas/efectos de los fármacos , Dendritas/enzimología , Conos de Crecimiento/efectos de los fármacos , Conos de Crecimiento/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Masculino , Mutación/genética , Neuritas/efectos de los fármacos , Neuritas/enzimología , Fosforilación/efectos de los fármacos , Fosfotirosina/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Seudópodos/efectos de los fármacos , Seudópodos/metabolismo , ARN Interferente Pequeño/metabolismo , Ratas , Ratas Sprague-Dawley , Quinasa Syk , Proteína Tirosina Quinasa ZAP-70/metabolismoRESUMEN
The chemokine SDF-1 is a secreted protein that plays a critical role in several aspects of neuron development through interaction with its unique receptor CXCR4. A key mechanism that controls neuron responsiveness to extracellular signals during neuronal growth is receptor endocytosis. Since we previously reported that SDF-1 regulates axon development without affecting the other neurites, we asked whether this could correlate with a compartment-selective trafficking of CXCR4. We thus studied CXCR4 behavior upon SDF-1 exposure in rat hippocampus slices and in transfected neuron cultures. A massive agonist-induced redistribution of CXCR4 in endosomes was observed in dendrites whereas no modification was evidenced in axons. Our data suggest that CXCR4 trafficking may play a role in mediating selective effects of SDF-1 on distinct neuronal membrane subdomains.