RESUMEN
Although ubiquitin ligases have been implicated in autism, their roles and mechanisms in brain development remain incompletely understood. Here, we report that in vivo knockdown or conditional knockout of the autism-linked ubiquitin ligase RNF8 or associated ubiquitin-conjugating enzyme UBC13 in rodent cerebellar granule neurons robustly increases the number of parallel fiber presynaptic boutons and functional parallel fiber/Purkinje cell synapses. In contrast to the role of nuclear RNF8 in proliferating cells, RNF8 operates in the cytoplasm in neurons to suppress synapse differentiation in vivo. Proteomics analyses reveal that neuronal RNF8 interacts with the HECT domain protein HERC2 and scaffold protein NEURL4, and knockdown of HERC2 or NEURL4 phenocopies the inhibition of RNF8/UBC13 signaling on synapse differentiation. In behavior analyses, granule neuron-specific knockout of RNF8 or UBC13 impairs cerebellar-dependent learning. Our study defines RNF8 and UBC13 as components of a novel cytoplasmic ubiquitin-signaling network that suppresses synapse formation in the brain.
Asunto(s)
Encéfalo/crecimiento & desarrollo , Neuronas/metabolismo , Sinapsis/ultraestructura , Enzimas Ubiquitina-Conjugadoras/genética , Ubiquitina-Proteína Ligasas/genética , Animales , Cerebelo/citología , Técnicas de Silenciamiento del Gen , Técnicas de Inactivación de Genes , Factores de Intercambio de Guanina Nucleótido/metabolismo , Ratones , Microscopía Electrónica , Técnicas de Placa-Clamp , Terminales Presinápticos/metabolismo , Terminales Presinápticos/ultraestructura , Proteómica , Células de Purkinje/metabolismo , Células de Purkinje/ultraestructura , Ratas , Transducción de Señal , Sinapsis/metabolismoRESUMEN
Activity-dependent transcription influences neuronal connectivity, but the roles and mechanisms of inactivation of activity-dependent genes have remained poorly understood. Genome-wide analyses in the mouse cerebellum revealed that the nucleosome remodeling and deacetylase (NuRD) complex deposits the histone variant H2A.z at promoters of activity-dependent genes, thereby triggering their inactivation. Purification of translating messenger RNAs from synchronously developing granule neurons (Sync-TRAP) showed that conditional knockout of the core NuRD subunit Chd4 impairs inactivation of activity-dependent genes when neurons undergo dendrite pruning. Chd4 knockout or expression of NuRD-regulated activity genes impairs dendrite pruning. Imaging of behaving mice revealed hyperresponsivity of granule neurons to sensorimotor stimuli upon Chd4 knockout. Our findings define an epigenetic mechanism that inactivates activity-dependent transcription and regulates dendrite patterning and sensorimotor encoding in the brain.
