RESUMEN
Excitotoxicity induced by NMDA receptors (NMDARs) is thought to be intimately linked to high intracellular calcium load. Unexpectedly, NMDAR-mediated toxicity can be eliminated without affecting NMDAR-induced calcium signals. Instead, excitotoxicity requires physical coupling of NMDARs to TRPM4. This interaction is mediated by intracellular domains located in the near-membrane portions of the receptors. Structure-based computational drug screening using the interaction interface of TRPM4 in complex with NMDARs identified small molecules that spare NMDAR-induced calcium signaling but disrupt the NMDAR/TRPM4 complex. These interaction interface inhibitors strongly reduce NMDA-triggered toxicity and mitochondrial dysfunction, abolish cyclic adenosine monophosphate-responsive element-binding protein (CREB) shutoff, boost gene induction, and reduce neuronal loss in mouse models of stroke and retinal degeneration. Recombinant or small-molecule NMDAR/TRPM4 interface inhibitors may mitigate currently untreatable human neurodegenerative diseases.
Asunto(s)
Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Descubrimiento de Drogas , Enfermedades Neurodegenerativas/metabolismo , Fármacos Neuroprotectores/química , Receptores de N-Metil-D-Aspartato/metabolismo , Proteínas Recombinantes/metabolismo , Canales Catiónicos TRPM/metabolismo , Animales , Señalización del Calcio , AMP Cíclico/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/genética , Técnicas de Silenciamiento del Gen , Humanos , Ratones , Enfermedades Neurodegenerativas/tratamiento farmacológico , Dominios Proteicos , Receptores de N-Metil-D-Aspartato/química , Receptores de N-Metil-D-Aspartato/uso terapéutico , Proteínas Recombinantes/química , Proteínas Recombinantes/uso terapéutico , Canales Catiónicos TRPM/genética , Activación TranscripcionalRESUMEN
Synapse-to-nucleus signaling triggered by synaptic NMDA receptors can lead to the buildup of a neuroprotective shield. Nuclear calcium activating the cAMP response element binding protein (CREB) plays a key role in neuroprotection acquired by synaptic activity. Here we show that in mouse hippocampal neurons, the transcription factor Atf3 (activating transcription factor 3) is a direct target of CREB. Induction of ATF3 expression by CREB in hippocampal neurons was initiated by calcium entry through synaptic NMDA receptors and required nuclear calcium transients and calcium/calmodulin-dependent protein kinase IV activity. Acting as a transcriptional repressor, ATF3 protects cultured hippocampal neurons from apoptosis and extrasynaptic NMDA receptor-induced cell death triggered by bath application of NMDA or oxygen-glucose deprivation. Expression of ATF3 in vivo using stereotaxic delivery of recombinant adeno-associated virus reduces brain damage following a cerebral ischemic insult in mice. Conversion of ATF3 to a transcriptional activator transforms ATF3 into a potent prodeath protein that kills neurons in cell culture and, when expressed in vivo in the hippocampus, ablates the neuronal cell layer. These results link nuclear calcium-CREB signaling to an ATF3-mediated neuroprotective gene repression program, indicating that activity-dependent shutoff of genes is an important process for survival. ATF3 supplementation may counteract age- and disease-related neuronal cell loss caused by a reduction in synaptic activity, malfunctioning of calcium signaling toward and within the nucleus ("nuclear calciopathy"), or increases in death signaling by extrasynaptic NMDA receptors.