RESUMO
Astrocytes are known to regulate and support neuronal and synaptic functions. Changes in their size and morphology in mouse models result in mental retardation. However, the mechanism underlying these morphological changes remains unclear. In the present study, abnormal astrocyte morphology was found in the mouse brain following knockout of dendritic cell factor 1 (Dcf1). Immunoprecipitation-mass spectrometry (IP-Mass) identified that ATP1B1 is bound to DCF1, and co-immunoprecipitation and cell fluorescence further confirmed an interaction between these two proteins, with asparagine residue 266 of ATP1B1 being required for the interaction with DCF1. Moreover, Dcf1 knockout in mice resulted in upregulation of ATP1B1 expression in the hippocampus. Furthermore, DCF1 interaction with ATP1B1 in astrocytes impaired their structural plasticity. Ultimately, Dcf1 knockout increased glutamate release. Mechanism exploration proposed that Dcf1 knockout led to significantly perturbed expression of AMPA receptors (AMPARs) and induced morphological changes in astrocytes through the P38 signaling pathway. Our data shed light on the possible mechanisms underlying changes in astrocyte morphology and provide new avenues for the exploration of proteins involved in glutamate release.
Assuntos
Astrócitos/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Sistema de Sinalização das MAP Quinases/genética , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Transdução de Sinais/genética , ATPase Trocadora de Sódio-Potássio/metabolismo , Animais , Animais Recém-Nascidos , Asparagina/metabolismo , Astrócitos/metabolismo , Células Cultivadas , Proteína Glial Fibrilar Ácida/genética , Proteína Glial Fibrilar Ácida/metabolismo , Ácido Glutâmico/metabolismo , Hipocampo/citologia , Humanos , Imunoprecipitação , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Domínios e Motivos de Interação entre Proteínas , RNA Mensageiro/metabolismo , Receptores de AMPA/genética , Receptores de AMPA/metabolismo , ATPase Trocadora de Sódio-Potássio/genéticaRESUMO
The study of neural dendrite formation is of great significance both in theory and applications. However, the molecular mechanisms of regulation remain unclear. We previously described a novel EST, which has high homology with dentritic cell factors (DCF1), expressed differentially between undifferentiated and differentiated neural stem cells (NSCs). In this study, we cloned, expressed, and silenced the dcf1 gene and offered insight into its function in regulating dendrite formation during the differentiation of NSCs. The results indicated that dcf1 encoded a 42 kD protein and could be successfully expressed both in Escherichia coli and NSCs. In order to silence dcf1 gene, three different kinds of siRNA vectors were constructed and transformed into the NSC line C17.2 and primary NSCs, resulting in down regulation of the dcf1 mRNA. Analysis of immunofluorescence or GFP illuminated that with overexpression of the dcf1 gene, the NSCs were maintained in undifferentiated status. After the dcf1 gene was silenced, cells tended to differentiate into neurons and astrocytes.
