Classifying Drosophila Olfactory Projection Neuron Subtypes by Single-Cell RNA Sequencing

Abstract

Summary The definition of neuronal type and how this relates to the transcriptome are open questions. Drosophila olfactory projection neurons (PNs) are among the best-characterized neuronal types: different PN classes target dendrites to distinct olfactory glomeruli, while PNs of the same class exhibit indistinguishable anatomical and physiological properties. Using single-cell RNA-sequencing, we comprehensively characterized the transcriptomes of most PN classes and unequivocally mapped transcriptomes to specific olfactory function for 6 classes. Transcriptomes of closely related PN classes exhibit the largest differences during circuit assembly but become indistinguishable in adults, suggesting that neuronal subtype diversity peaks during development. Transcription factors and cell-surface molecules are the most differentially expressed genes between classes and are highly informative in encoding cell identity, enabling us to identify a new lineage-specific transcription factor that instructs PN dendrite targeting. These findings establish that neuronal transcriptomic identity corresponds with anatomical and physiological identity defined by connectivity and function.

Building blocks of the human brain

“…Although future experiments are needed to compare cortical microcircuit architecture in specific cortical areas, our findings suggest that in the developing human cortex, topographically distinct minicolumns consist of different classes of excitatory neurons. Importantly, similar area-specific neuronal signatures have been reported in the adult mouse brain (20), indicating that the topographic variation that we described reflects differences in neuronal identities, rather than transient developmental variation (21).…”

Molecular, spatial and projection diversity of neurons in primary motor cortex revealed by in situ single-cell transcriptomics

“…We selected a panel of 258 genes for MERFISH imaging (Figure 1a) by combining three distinct approaches: (i) 62 canonical marker genes for major neuronal and non-neuronal cell types in the cortex were selected based on prior knowledge; (ii) 168 genes were selected based on pair-wise differential gene expression analysis on the neuronal clusters identified by a concurrent singlecell / single-nucleus RNA-sequencing (sc/snRNA-seq) study reported in a companion BICCN paper 38 ; (iii) a set of genes (50 for glutamatergic and 50 for GABAergic neuronal clusters) were selected which contained the highest mutual information 39,40 among the clusters identified by sc/snRNA-seq. The overlapping gene lists generated by these three approaches were combined to form a panel of 258 genes total, in which 242 were imaged using MERFISH, and the remaining 16 genes, which were either relatively short or highly expressed, were measured in eight sequential rounds of a two-color FISH following MERFISH.…”

“…To discriminate different neuronal types, we took two approaches to select genes. In the first approach, we selected a panel of most informative genes using mutual information analysis as reported previously 39 . Briefly, we used an information theory quantity known as mutual information 40 to determine the relative amount of information each gene carries in defining the sc/snRNA-seq clusters.…”

Recent advances in neuropeptide signaling in Drosophila, from genes to physiology and behavior

“…Recent progress in single-cell transcriptome sequencing has also facilitated the creation of databases that allow mining of gene expression in the brain (http://scope.aertslab.org) and gut (https://www.flyrnai.org/scRNA/) at the cellular level [117][118][119][120]. In some cases the name of the precursor is used (when the peptides derived have multiple names and/or when multiple peptides derived from the same precursor are listed).…”