RESUMO
Synaptotagmin-7 (Syt7) plays direct or redundant Ca2+ sensor roles in multiple forms of vesicle exocytosis in synapses. Here, we show that Syt7 is a redundant Ca2+ sensor with Syt1/Doc2 to drive spontaneous glutamate release, which functions uniquely to activate the postsynaptic GluN2B-containing NMDARs that significantly contribute to mental illness. In mouse hippocampal neurons lacking Syt1/Doc2, Syt7 inactivation largely diminishes spontaneous release. Using 2 approaches, including measuring Ca2+ dose response and substituting extracellular Ca2+ with Sr2+, we detect that Syt7 directly triggers spontaneous release via its Ca2+ binding motif to activate GluN2B-NMDARs. Furthermore, modifying the localization of Syt7 in the active zone still allows Syt7 to drive spontaneous release, but the GluN2B-NMDAR activity is abolished. Finally, Syt7 SNPs identified in bipolar disorder patients destroy the function of Syt7 in spontaneous release in patient iPSC-derived and mouse hippocampal neurons. Therefore, Syt7 could contribute to neuropsychiatric disorders through driving spontaneous glutamate release.
Assuntos
Transtorno Bipolar/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Sinaptotagminas/fisiologia , Animais , Transtorno Bipolar/genética , Cálcio/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Modelos Animais de Doenças , Predisposição Genética para Doença , Ácido Glutâmico/metabolismo , Células HEK293 , Hipocampo/citologia , Hipocampo/metabolismo , Humanos , Camundongos , Camundongos Knockout , Neurônios/metabolismo , Sinaptotagminas/genéticaRESUMO
microRNA-218 (miR-218) has been linked to several cognition related neurodegenerative and neuropsychiatric disorders. However, whether miR-218 plays a direct role in cognitive functions remains unknown. Here, using the miR-218 knockout (KO) mouse model and the sponge/overexpression approaches, we showed that miR-218-2 but not miR-218-1 could bidirectionally regulate the contextual and spatial memory in the mice. Furthermore, miR-218-2 deficiency induced deficits in the morphology and presynaptic neurotransmitter release in the hippocampus to impair the long term potentiation. Combining the RNA sequencing analysis and luciferase reporter assay, we identified complement component 3 (C3) as a main target gene of miR-218 in the hippocampus to regulate the presynaptic functions. Finally, we showed that restoring the C3 activity in the miR-218-2 KO mice could rescue the synaptic and learning deficits. Therefore, miR-218-2 played an important role in the cognitive functions of mice through C3, which can be a mechanism for the defective cognition of miR-218 related neuronal disorders.
Assuntos
Complemento C3/genética , Hipocampo/metabolismo , Potenciação de Longa Duração , MicroRNAs/metabolismo , Vesículas Sinápticas/metabolismo , Regiões 3' não Traduzidas , Animais , Células Cultivadas , Complemento C3/metabolismo , Exocitose , Hipocampo/citologia , Hipocampo/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Neurônios/metabolismo , Neurônios/fisiologiaRESUMO
Synaptotagmin-7 (Syt7) probably plays an important role in bipolar-like behavioral abnormalities in mice; however, the underlying mechanisms for this have remained elusive. Unlike antidepressants that cause mood overcorrection in bipolar depression, N-methyl-d-aspartate receptor (NMDAR)-targeted drugs show moderate clinical efficacy, for unexplained reasons. Here we identified Syt7 single nucleotide polymorphisms (SNPs) in patients with bipolar disorder and demonstrated that mice lacking Syt7 or expressing the SNPs showed GluN2B-NMDAR dysfunction, leading to antidepressant behavioral consequences and avoidance of overcorrection by NMDAR antagonists. In human induced pluripotent stem cell (iPSC)-derived and mouse hippocampal neurons, Syt7 and GluN2B-NMDARs were localized to the peripheral synaptic region, and Syt7 triggered multiple forms of glutamate release to efficiently activate the juxtaposed GluN2B-NMDARs. Thus, while Syt7 deficiency and SNPs induced GluN2B-NMDAR dysfunction in mice, patient iPSC-derived neurons showed Syt7 deficit-induced GluN2B-NMDAR hypoactivity that was rescued by Syt7 overexpression. Therefore, Syt7 deficits induced mania-like behaviors in mice by attenuating GluN2B activity, which enabled NMDAR antagonists to avoid mood overcorrection.
Assuntos
Comportamento Animal , Mania/patologia , Receptores de N-Metil-D-Aspartato/metabolismo , Sinaptotagminas/deficiência , Adulto , Idoso , Animais , Transtorno Bipolar/genética , Transtorno Bipolar/patologia , Exocitose , Feminino , Ácido Glutâmico/metabolismo , Hipocampo/patologia , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Masculino , Mania/fisiopatologia , Camundongos Knockout , Pessoa de Meia-Idade , Neurônios/metabolismo , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Vesículas Sinápticas/metabolismo , Sinaptotagminas/genética , Sinaptotagminas/metabolismo , Adulto JovemRESUMO
The pathogenesis of bipolar disorder (BD) has remained enigmatic, largely because genetic animal models based on identified susceptible genes have often failed to show core symptoms of spontaneous mood cycling. However, pedigree and induced pluripotent stem cell (iPSC)-based analyses have implicated that dysfunction in some key signaling cascades might be crucial for the disease pathogenesis in a subpopulation of BD patients. We hypothesized that the behavioral abnormalities of patients and the comorbid metabolic abnormalities might share some identical molecular mechanism. Hence, we investigated the expression of insulin/synapse dually functioning genes in neurons derived from the iPSCs of BD patients and the behavioral phenotype of mice with these genes silenced in the hippocampus. By these means, we identified synaptotagmin-7 (Syt7) as a candidate risk factor for behavioral abnormalities. We then investigated Syt7 knockout (KO) mice and observed nocturnal manic-like and diurnal depressive-like behavioral fluctuations in a majority of these animals, analogous to the mood cycling symptoms of BD. We treated the Syt7 KO mice with clinical BD drugs including olanzapine and lithium, and found that the drug treatments could efficiently regulate the behavioral abnormalities of the Syt7 KO mice. To further verify whether Syt7 deficits existed in BD patients, we investigated the plasma samples of 20 BD patients and found that the Syt7 mRNA level was significantly attenuated in the patient plasma compared to the healthy controls. We therefore concluded that Syt7 is likely a key factor for the bipolar-like behavioral abnormalities.
Assuntos
Transtorno Bipolar/metabolismo , Transtorno Bipolar/psicologia , Sinaptotagminas/metabolismo , Adulto , Animais , Comportamento , Transtorno Bipolar/sangue , Transtorno Bipolar/genética , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Masculino , Camundongos , Camundongos Knockout , Neurônios/metabolismo , Sinaptotagminas/genética , Adulto JovemRESUMO
Bipolar disorder is a complex neuropsychiatric disorder that is characterized by intermittent episodes of mania and depression; without treatment, 15% of patients commit suicide. Hence, it has been ranked by the World Health Organization as a top disorder of morbidity and lost productivity. Previous neuropathological studies have revealed a series of alterations in the brains of patients with bipolar disorder or animal models, such as reduced glial cell number in the prefrontal cortex of patients, upregulated activities of the protein kinase A and C pathways and changes in neurotransmission. However, the roles and causation of these changes in bipolar disorder have been too complex to exactly determine the pathology of the disease. Furthermore, although some patients show remarkable improvement with lithium treatment for yet unknown reasons, others are refractory to lithium treatment. Therefore, developing an accurate and powerful biological model for bipolar disorder has been a challenge. The introduction of induced pluripotent stem-cell (iPSC) technology has provided a new approach. Here we have developed an iPSC model for human bipolar disorder and investigated the cellular phenotypes of hippocampal dentate gyrus-like neurons derived from iPSCs of patients with bipolar disorder. Guided by RNA sequencing expression profiling, we have detected mitochondrial abnormalities in young neurons from patients with bipolar disorder by using mitochondrial assays; in addition, using both patch-clamp recording and somatic Ca(2+) imaging, we have observed hyperactive action-potential firing. This hyperexcitability phenotype of young neurons in bipolar disorder was selectively reversed by lithium treatment only in neurons derived from patients who also responded to lithium treatment. Therefore, hyperexcitability is one early endophenotype of bipolar disorder, and our model of iPSCs in this disease might be useful in developing new therapies and drugs aimed at its clinical treatment.
Assuntos
Potenciais de Ação/efeitos dos fármacos , Antipsicóticos/farmacologia , Transtorno Bipolar/patologia , Compostos de Lítio/farmacologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Sinalização do Cálcio/efeitos dos fármacos , Giro Denteado/efeitos dos fármacos , Giro Denteado/patologia , Endofenótipos , Humanos , Células-Tronco Pluripotentes Induzidas/patologia , Masculino , Mitocôndrias/patologia , Técnicas de Patch-ClampRESUMO
Copy-number variations (CNVs) of the human 16p11.2 genetic locus are associated with neurodevelopmental disorders, including autism spectrum disorders (ASDs) and schizophrenia. However, it remains largely unclear how this locus is involved in the disease pathogenesis. Doc2α is localized within this locus. Here, using in vivo and ex vivo electrophysiological and morphological approaches, we show that Doc2α-deficient mice have neuronal morphological abnormalities and defects in neural activity. Moreover, the Doc2α-deficient mice exhibit social and repetitive behavioral deficits. Furthermore, we demonstrate that Doc2α functions in behavioral and neural phenotypes through interaction with Secretagogin (SCGN). Finally, we demonstrate that SCGN functions in social/repetitive behaviors, glutamate release, and neuronal morphology of the mice through its Doc2α-interacting activity. Therefore, Doc2α likely contributes to neurodevelopmental disorders through its interaction with SCGN.
Assuntos
Transtorno do Espectro Autista , Esquizofrenia , Animais , Humanos , Camundongos , Transtorno do Espectro Autista/genética , Deleção Cromossômica , Cromossomos Humanos Par 16/genética , Variações do Número de Cópias de DNA/genética , Esquizofrenia/genética , Secretagoginas/genética , Comportamento SocialRESUMO
Synaptic vesicle (SV) docking is a dynamic multi-stage process that is required for efficient neurotransmitter release in response to nerve impulses. Although the steady-state SV docking likely involves the cooperation of Synaptotagmin-1 (Syt1) and soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), where and how the docking process initiates remains unknown. Phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) can interact with Syt1 and SNAREs to contribute to vesicle exocytosis. In the present study, using the CRISPRi-mediated multiplex gene knockdown and 3D electron tomography approaches, we show that in mouse hippocampal synapses, SV docking initiates at â¼12 nm to the active zone (AZ) by Syt1. Furthermore, we demonstrate that PI(4,5)P2 is the membrane partner of Syt1 to initiate SV docking, and disrupting their interaction could abolish the docking initiation. In contrast, the SNARE complex contributes only to the tight SV docking within 0-2 nm. Therefore, Syt1 interacts with PI(4,5)P2 to loosely dock SVs within 2-12 nm to the AZ in hippocampal neurons.
Assuntos
Hipocampo/citologia , Neurônios/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Vesículas Sinápticas/metabolismo , Sinaptotagmina I/metabolismo , Animais , Células HEK293 , Humanos , Camundongos Endogâmicos C57BL , Neurônios/ultraestrutura , Ligação Proteica , Proteínas SNARE/metabolismo , Vesículas Sinápticas/ultraestrutura , Proteína 25 Associada a Sinaptossoma/metabolismo , Sintaxina 1/metabolismo , Proteína 2 Associada à Membrana da Vesícula/metabolismoRESUMO
NSF (N-ethylmaleimide-sensitive factor) and α-SNAP (α-soluble NSF attachment protein) bind to the SNARE (soluble NSF attachment protein receptor) complex, the minimum machinery to mediate membrane fusion, to form a 20S complex, which disassembles the SNARE complex for reuse. We report the cryo-EM structures of the α-SNAP-SNARE subcomplex and the NSF-D1D2 domain in the 20S complex at 3.9- and 3.7-Å resolutions, respectively. Combined with the biochemical and electrophysiological analyses, we find that α-SNAPs use R116 through electrostatic interactions and L197 through hydrophobic interactions to apply force mainly on two positions of the VAMP protein to execute disassembly process. Furthermore, we define the interaction between the amino terminus of the SNARE helical bundle and the pore loop of the NSF-D1 domain and demonstrate its essential role as a potential anchor for SNARE complex disassembly. Our studies provide a rotation model of α-SNAP-mediated disassembly of the SNARE complex.
Assuntos
Fenômenos Mecânicos , Complexos Multiproteicos/química , Proteínas SNARE/química , Aminoácidos , Sítios de Ligação , Modelos Moleculares , Complexos Multiproteicos/metabolismo , Complexos Multiproteicos/ultraestrutura , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Proteínas SNARE/metabolismo , Proteínas de Ligação a Fator Solúvel Sensível a N-Etilmaleimida/química , Proteínas de Ligação a Fator Solúvel Sensível a N-Etilmaleimida/metabolismo , Relação Estrutura-AtividadeRESUMO
Recently, a novel bunyavirus, severe fever with thrombocytopenia syndrome virus (SFTSV), was isolated in central China. The virus can cause multi-clinical symptoms: severe fever, thrombocytopenia, leukocytopenia, with a mortality rate of ~10%. Several studies show that SFTSV could undergo rapid evolution via gene mutation and homologous recombination. However, as an important evolutionary force for segmented-genome viruses, reassortment has not been reported in SFTSV. In this study, we identified two SFTSV strains of which the S segment has different origin from M and L, suggesting that reassortment might be potential force driving rapid change of SFTSV. This result might shed new light on the evolutionary behavior of the novel virus.