Enhancer viruses and a transgenic platform for combinatorial cell subclass-specific labeling

Abstract

SummaryThe rapid pace of cell type identification by new single-cell analysis methods has not been met with efficient experimental access to the newly discovered types. To enable flexible and efficient access to specific neural populations in the mouse cortex, we collected chromatin accessibility data from individual cells and clustered the single-cell data to identify enhancers specific for cell classes and subclasses. When cloned into adeno-associated viruses (AAVs) and delivered to the brain by retro-orbital injections, these enhancers drive transgene expression in specific cell subclasses in the cortex. We characterize several enhancer viruses in detail to show that they result in labeling of different projection neuron subclasses in mouse cortex, and that one of them can be used to label the homologous projection neuron subclass in human cortical slices. To enable the combinatorial labeling of more than one cell type by enhancer viruses, we developed a three-color Cre-, Flp- and Nigri-recombinase dependent reporter mouse line, Ai213. The delivery of three enhancer viruses driving these recombinases via a single retroorbital injection into a single Ai213 transgenic mouse results in labeling of three different neuronal classes/subclasses in the same brain tissue. This approach combines unprecedented flexibility with specificity for investigation of cell types in the mouse brain and beyond.

Single cell epigenomic atlas of the developing human brain and organoids

“…In order to determine the fidelity of organoids as a model for the epigenomic landscape of the developing brain, functional characterization of conserved and non-conserved chromatin features in vivo and in vitro will be required. In addition to serving as a reference for evaluating in vitro models of human brain development, our data will enable scalable prediction of candidate cell type specific regulatory elements that could facilitate the development of enhancer-based tools for accessing molecularly-defined cell types [57][58][59] .…”

Transcriptomic evidence that von Economo neurons are regionally specialized extratelencephalic-projecting excitatory neurons

“…Furthermore, novel viral tools provide cell type-specific genetic targeting in this system 61 , and application of enhancers for ET cells, such as the Fam84b enhancer that labels mouse ET cells with >90% specificity 62 , could be used to label VENs in ex vivo human brain tissue. Such studies can help to further refine our understanding of the defining characteristics of VENs, potentially providing information about their local connectivity and teasing out subtle gene expression differences between ET cells with spindle, fork, and pyramidal shapes.…”

Broad gene expression throughout the mouse and marmoset brain after intravenous delivery of engineered AAV capsids

“…Previously, we harnessed the power and specificity of Cre transgenic mice to apply increased selective pressure to viral engineering, leading to the variants AAV-PHP.B and AAV-PHP.eB, which cross the blood-brain barrier following intravenous (IV) administration and broadly transduce cells throughout the CNS 17,18 . These variants transformed the way we study the CNS in rodents, opening up completely novel approaches for measuring and affecting brain activity, both in exploratory and translational contexts [19][20][21][22] . However, these variants also transduce cells in the liver and other offtarget organs.…”