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A light- and calcium-gated transcription factor for imaging and manipulating activated neurons
Nature Biotechnology ( IF 46.9 ) Pub Date : 2017-06-26 00:00:00 , DOI: 10.1038/nbt.3909
Wenjing Wang , Craig P Wildes , Tanyaporn Pattarabanjird , Mateo I Sanchez , Gordon F Glober , Gillian A Matthews , Kay M Tye , Alice Y Ting

Activity remodels neurons, altering their molecular, structural, and electrical characteristics. To enable the selective characterization and manipulation of these neurons, we present FLARE, an engineered transcription factor that drives expression of fluorescent proteins, opsins, and other genetically encoded tools only in the subset of neurons that experienced activity during a user-defined time window. FLARE senses the coincidence of elevated cytosolic calcium and externally applied blue light, which together produce translocation of a membrane-anchored transcription factor to the nucleus to drive expression of any transgene. In cultured rat neurons, FLARE gives a light-to-dark signal ratio of 120 and a high- to low-calcium signal ratio of 10 after 10 min of stimulation. Opsin expression permitted functional manipulation of FLARE-marked neurons. In adult mice, FLARE also gave light- and motor-activity-dependent transcription in the cortex. Due to its modular design, minute-scale temporal resolution, and minimal dark-state leak, FLARE should be useful for the study of activity-dependent processes in neurons and other cells that signal with calcium.

中文翻译:

光和钙门控转录因子,用于成像和操纵活化的神经元

活动会重塑神经元,从而改变其分子,结构和电学特征。为了能够选择性地表征和操纵这些神经元,我们提出了FLARE,这是一种工程化的转录因子,仅在用户定义的时间窗口内经历活动的神经元子集中驱动荧光蛋白,视蛋白和其他遗传编码工具的表达。FLARE感觉到细胞质钙的增加和外部施加的蓝光的巧合,它们共同产生膜锚定转录因子向核的转运,从而驱动任何转基因的表达。在培养的大鼠神经元中,刺激10分钟后,FLARE产生的明暗信号比为120,高钙离子比低钙信号比为10。视蛋白表达允许FLARE标记的神经元的功能操纵。在成年小鼠中,FLARE在皮质中也提供了光和运动活性相关的转录。由于其模块化的设计,微小的时间分辨率和最小的暗态泄漏,FLARE对于研究神经元和其他以钙发出信号的细胞中与活动有关的过程很有用。
更新日期:2017-09-12
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