Virally encoded connectivity transgenic overlay RNA sequencing (VECTORseq) defines projection neurons involved in sensorimotor integration.
Virally encoded connectivity transgenic overlay RNA sequencing (VECTORseq) defines projection neurons involved in sensorimotor integration.
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DOI：
10.1016/j.celrep.2021.110131
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发表时间：
2021-12-21
期刊：
影响因子：
8.8
通讯作者：
Feinberg EH
中科院分区：
文献类型：
--
作者：
Cheung V;Chung P;Bjorni M;Shvareva VA;Lopez YC;Feinberg EH
行为产生于整个大脑的协同活动。因此，现代神经科学的一个主要焦点是定义连接不同大脑区域的投射神经元的生理和行为角色。单细胞RNA测序通过揭示细胞生理的分子决定因素和能够进行基因靶向干扰的标记，如光遗传学，促进了这些努力，但现有的对确定的投影群体进行测序的方法吞吐量低、费力且成本高昂。我们开发了一种简单、多重的方法，病毒编码的连通性转基因覆盖RNA测序(VECTORseq)。VECTORseq通过将病毒转基因mRNA视为单细胞数据集中的条形码，改变了商业逆行感染病毒的用途，这些病毒通常用于表达功能性转基因(例如重组酶和荧光蛋白)。VECTORseq与不同的病毒家族兼容，在一次测序运行中解析具有不同投影目标的多个种群，并识别皮质和皮质下的兴奋性和抑制性投影种群。我们的研究为基于连接性的高通量识别神经元亚型提供了路线图。具有不同轴突投射目标的神经元以及不同的分子和功能特性混杂在整个大脑中。Cheung等人。建立一种高通量的方法，VECTORseq，以转录方式定义投射神经元。该方法将广泛使用的逆行感染病毒表达的转基因重新想象为在单细胞测序中检测到的多重RNA条形码。
Behavior arises from concerted activity throughout the brain. Consequently, a major focus of modern neuroscience is defining the physiology and behavioral roles of projection neurons linking different brain areas. Single-cell RNA sequencing has facilitated these efforts by revealing molecular determinants of cellular physiology and markers that enable genetically targeted perturbations such as optogenetics, but existing methods for sequencing defined projection populations are low throughput, painstaking, and costly. We developed a straightforward, multiplexed approach, virally encoded connectivity transgenic overlay RNA sequencing (VECTORseq). VECTORseq repurposes commercial retrogradely infecting viruses typically used to express functional transgenes (e.g., recombinases and fluorescent proteins) by treating viral transgene mRNA as barcodes within single-cell datasets. VECTORseq is compatible with different viral families, resolves multiple populations with different projection targets in one sequencing run, and identifies cortical and subcortical excitatory and inhibitory projection populations. Our study provides a roadmap for high-throughput identification of neuronal subtypes based on connectivity. Neurons with different axonal projection targets and distinct molecular and functional properties are intermingled throughout the brain. Cheung et al. establish a high-throughput method, VECTORseq, to transcriptionally define projection neurons. The method reimagines transgenes expressed by widely used retrogradely infecting viruses as multiplexed RNA barcodes that are detected in single-cell sequencing.
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