RESUMO
Stress granules are non-membranous structures that transiently form in the cytoplasm during cellular stress, where they promote translational repression of non-essential RNAs and modulate cell signaling by sequestering key signal transduction proteins. These and other functions of stress granules facilitate an adaptive cellular response to environmental adversity. A key component of stress granules is the prion-related RNA-binding protein, T cell intracellular antigen-1 (TIA-1). Here, we report that recombinant TIA-1 undergoes rapid multimerization and phase separation in the presence of divalent zinc, which can be reversed by the zinc chelator, TPEN. Similarly, the formation and maintenance of TIA-1-positive stress granules in arsenite-treated cells are inhibited by TPEN. In addition, Zn2+ is released in cells treated with arsenite, before stress granule formation. These findings suggest that Zn2+ is a physiological ligand of TIA-1, acting as a stress-inducible second messenger to promote multimerization of TIA-1 and subsequent localization into stress granules.
Assuntos
Arsenitos/farmacologia , Grânulos Citoplasmáticos , Multimerização Proteica/efeitos dos fármacos , Sistemas do Segundo Mensageiro/efeitos dos fármacos , Antígeno-1 Intracelular de Células T , Zinco , Linhagem Celular , Grânulos Citoplasmáticos/química , Grânulos Citoplasmáticos/genética , Grânulos Citoplasmáticos/metabolismo , Humanos , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Antígeno-1 Intracelular de Células T/química , Antígeno-1 Intracelular de Células T/genética , Antígeno-1 Intracelular de Células T/metabolismo , Zinco/química , Zinco/metabolismoRESUMO
Neurotrophins control the development and adult plasticity of the vertebrate nervous system. Failure to identify invertebrate neurotrophin orthologs, however, has precluded studies in invertebrate models, limiting our understanding of fundamental aspects of neurotrophin biology and function. We identified a neurotrophin (ApNT) and Trk receptor (ApTrk) in the mollusk Aplysia and found that they play a central role in learning-related synaptic plasticity. Blocking ApTrk signaling impairs long-term facilitation, whereas augmenting ApNT expression enhances it and induces the growth of new synaptic varicosities at the monosynaptic connection between sensory and motor neurons of the gill-withdrawal reflex. Unlike vertebrate neurotrophins, ApNT has multiple coding exons and exerts distinct synaptic effects through differentially processed and secreted splice isoforms. Our findings demonstrate the existence of bona fide neurotrophin signaling in invertebrates and reveal a posttranscriptional mechanism that regulates neurotrophin processing and the release of proneurotrophins and mature neurotrophins that differentially modulate synaptic plasticity.
Assuntos
Fatores de Crescimento Neural/metabolismo , Sinapses/metabolismo , Processamento Alternativo , Sequência de Aminoácidos , Animais , Aplysia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Células Cultivadas , Técnicas de Cocultura , Células HEK293 , Humanos , Dados de Sequência Molecular , Neurônios Motores/citologia , Neurônios Motores/metabolismo , Fatores de Crescimento Neural/química , Fatores de Crescimento Neural/genética , Plasticidade Neuronal , Células PC12 , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Ratos , Receptor trkA/química , Receptor trkA/genética , Receptor trkA/metabolismo , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/metabolismo , Serotonina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacosRESUMO
Here we describe a strategy designed to identify RNAs that are actively transported to synapses during learning. Our approach is based on the characterization of RNA transport complexes carried by molecular motor kinesin. Using this strategy in Aplysia, we have identified 5,657 unique sequences consisting of both coding and noncoding RNAs from the CNS. Several of these RNAs have key roles in the maintenance of synaptic function and growth. One of these RNAs, myosin heavy chain, is critical in presynaptic sensory neurons for the establishment of long-term facilitation, but not for its persistence.
Assuntos
Aplysia/genética , Perfilação da Expressão Gênica/métodos , Sinapses/genética , Transcriptoma/genética , Animais , Sistema Nervoso Central/metabolismo , Genoma/genética , Sequenciamento de Nucleotídeos em Larga Escala , Hibridização In Situ , Cinesinas/metabolismo , Potenciação de Longa Duração/genética , Cadeias Pesadas de Miosina/metabolismo , Neurônios/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Transporte Proteico/genética , RNA/genética , RNA/metabolismo , Transporte de RNA/genética , Análise de Sequência de RNARESUMO
Neurexin and neuroligin, which undergo heterophilic interactions with each other at the synapse, are mutated in some patients with autism spectrum disorder, a set of disorders characterized by deficits in social and emotional learning. We have explored the role of neurexin and neuroligin at sensory-to-motor neuron synapses of the gill-withdrawal reflex in Aplysia, which undergoes sensitization, a simple form of learned fear. We find that depleting neurexin in the presynaptic sensory neuron or neuroligin in the postsynaptic motor neuron abolishes both long-term facilitation and the associated presynaptic growth induced by repeated pulses of serotonin. Moreover, introduction into the motor neuron of the R451C mutation of neuroligin-3 linked to autism spectrum disorder blocks both intermediate-term and long-term facilitation. Our results suggest that activity-dependent regulation of the neurexin-neuroligin interaction may govern transsynaptic signaling required for the storage of long-term memory, including emotional memory that may be impaired in autism spectrum disorder.
Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Potenciação de Longa Duração/fisiologia , Proteínas de Membrana/metabolismo , Neurônios Motores/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Receptores de Superfície Celular/metabolismo , Células Receptoras Sensoriais/fisiologia , Análise de Variância , Animais , Aplysia , Arginina/genética , Moléculas de Adesão Celular Neuronais/genética , Células Cultivadas , Sistema Nervoso Central/citologia , Clonagem Molecular/métodos , Cisteína/genética , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Proteínas de Fluorescência Verde/genética , Humanos , Potenciação de Longa Duração/efeitos dos fármacos , Proteínas de Membrana/genética , Microinjeções/métodos , Dados de Sequência Molecular , Neurônios Motores/efeitos dos fármacos , Mutação/genética , Proteínas do Tecido Nervoso/genética , Oligodesoxirribonucleotídeos Antissenso/farmacologia , Ligação Proteica/fisiologia , Receptores de Superfície Celular/genética , Células Receptoras Sensoriais/efeitos dos fármacos , Serotonina/farmacologia , Sinapses/metabolismo , Sinapses/fisiologiaRESUMO
To explore how gene products, required for the initiation of synaptic growth, move from the cell body of the sensory neuron to its presynaptic terminals, and from the cell body of the motor neuron to its postsynaptic dendritic spines, we have investigated the anterograde transport machinery in both the sensory and motor neurons of the gill-withdrawal reflex of Aplysia. We found that the induction of long-term facilitation (LTF) by repeated applications of serotonin, a modulatory transmitter released during learning in Aplysia, requires upregulation of kinesin heavy chain (KHC) in both pre- and postsynaptic neurons. Indeed, upregulation of KHC in the presynaptic neurons alone is sufficient for the induction of LTF. However, KHC is not required for the persistence of LTF. Thus, in addition to transcriptional activation in the nucleus and local protein synthesis at the synapse, our studies have identified a third component critical for long-term learning-related plasticity: the coordinated upregulation of kinesin-mediated transport.