RbFox1-Nestin hippocampus

Dysregulation of the brain-enriched RNA binding protein Rbfox1 has been linked to neurologic diseases such as epilepsy and autism spectrum disorders. However, it remains unexplored how distinct neuronal populations might contribute to neurologic dysfunction resulting from Rbfox1 loss. To examine these issues we profiled gene expression specifically in the hippocampus of wildtype and Rbfox1-/- mice. We identified transcripts whose expression was strongly Rbfox1-dependent and exhibited significant Rbfox1 binding in their 3’UTRs. One prominent target, Vamp1, was found to be specifically expressed in GABAergic interneurons. Both Vamp1 knockdown and Rbfox1 loss led to decreased synaptic transmission, and altered E/I balance in the Rbfox1-/- hippocampus, indicating that Vamp1 loss is a major component of the Rbfox1-/- physiological phenotype. The cytoplasmic isoform of Rbfox1 was sufficient to rescue Vamp1 expression in Rbfox1-/- neurons. We show that Rbfox1 binding in the Vamp1 3'UTR promotes its expression in part by antagonizing the brain-enriched microRNA-9. These results demonstrate that inhibitory neurons maintain specialized synaptic vesicle release machinery containing Vamp1 that is critically regulated by Rbfox1. We performed RNA-seq to profile gene expression and splicing changes. Adult (P60-P70) hippocampal tissue was dissected from 3 pairs of male wildtype (Rbfox1 fl/fl; +/+) and Nes-cKO (Rbfox1 fl/fl; Nes/+) mice. Gene expression and splicing profiles were compared between wildtype and Nes-cKO. 6 total samples were analyzed.脑富集RNA结合蛋白（RNA binding protein）Rbfox1的失调与癫痫、自闭症谱系障碍等神经系统疾病密切相关。然而，Rbfox1缺失所引发的神经系统功能障碍具体由哪些不同的神经元群体介导，目前仍未得到阐明。 为探究上述问题，我们对野生型与Rbfox1基因敲除（Rbfox1-/-）小鼠的海马体组织开展了特异性基因表达谱分析。我们鉴定出一批表达显著依赖Rbfox1，且其3'非翻译区（3' untranslated region, 3'UTR）存在显著Rbfox1结合位点的转录本。其中一个核心靶标Vamp1，被证实特异性表达于γ-氨基丁酸能中间神经元（GABAergic interneurons）中。 Vamp1敲低与Rbfox1缺失均可导致突触传递功能减弱，并改变Rbfox1-/-小鼠海马体的兴奋-抑制平衡，提示Vamp1缺失是Rbfox1-/-小鼠生理表型的关键介导组分。Rbfox1的细胞质亚型足以挽救Rbfox1-/-神经元中Vamp1的表达水平。 我们证实，Rbfox1结合于Vamp1的3'UTR，可通过拮抗脑富集微小RNA-9（microRNA-9, miR-9）的作用，部分促进Vamp1的表达。上述结果表明，抑制性神经元维持着一套包含Vamp1的特异性突触囊泡释放机制，而该机制受到Rbfox1的关键调控。 我们通过RNA测序（RNA-seq）对基因表达及剪接变化进行了谱分析。实验所用组织为成年（P60-P70）小鼠的海马体，分别从3对雄性野生型（Rbfox1 fl/fl; +/+）与Nes条件性敲除（Nes-cKO, Rbfox1 fl/fl; Nes/+）小鼠中分离获取。随后我们对野生型与Nes条件性敲除小鼠的基因表达及剪接谱进行了比较分析，共对6个样本进行了分析。