Iterative assay for transposase-accessible chromatin by sequencing to home in on functionally relevant neuronal subtypes
The Drosophila brain contains tens of thousands of distinct cell types. Numerous different transgenic lines reproducibly target specific neuron subsets, yet most still express in several distinct cell types. Furthermore, most lines were developed without a priori knowledge of where the transgenes would be expressed. To aid in the development of cell type-specific tools for neuronal identification and manipulation, we used an iterative assay for transposase-accessible chromatin by sequencing (ATAC-seq) approach. Open chromatin regions (OCRs) that were enriched in neurons compared to whole bodies drove transgene expression preferentially in subsets of neurons. A second round of ATAC-seq from these specific neuron subsets revealed additional enriched OCRs that further restricted transgene expression within the chosen neuron subset. This approach allows for continued refinement of transgene expression and can be used to identify neurons relevant for sleep behavior. Furthermore, this approach is widely applicable to other cell types and to other organisms. Overall design: GFP-positive nuclei from whole flies, neurons in the head, and neuronal subtypes were isolated by fluorescence-activated nuclei sorting and were used to generate ATAC-seq libraries.