Neuronal diversity and convergence in a visual system developmental atlas

Abstract

Summary Deciphering how neuronal diversity is established and maintained requires a detailed knowledge of neuronal gene expression throughout development. In contrast to mammalian brains 1 , 2 , the large neuronal diversity of the Drosophila optic lobes 3 and its connectome 4 – 6 are almost completely characterized. However, a molecular characterization of this diversity, particularly during development, has been lacking. We present novel insights into brain development through a nearly exhaustive description of the transcriptomic diversity of the optic lobes. We acquired the transcriptome of 275,000 single-cells at adult and five pupal stages, and developed a machine learning framework to assign them to almost 200 cell-types at all timepoints. We discovered two large neuronal populations that wrap neuropils during development but die just before adulthood, as well as neuronal subtypes that partition dorsal and ventral visual circuits by differential Wnt signaling throughout development. Moreover, we showed that neurons of the same type but produced days apart synchronize their transcriptomes shortly after being produced. We also resolved during synaptogenesis neuronal subtypes that converge to indistinguishable transcriptomic profiles in adults while greatly differing in morphology and connectivity. Our datasets almost completely account for the known neuronal diversity of the optic lobes and serve as a paradigm to understand brain development across species.

Temporal evolution of single-cell transcriptomes of Drosophila olfactory projection neurons

“…Furthermore, whereas TFs are generally expressed in a binary fashion throughout development ( Figure 9C and Figure 9figure supplement 1), many CSMs exhibit graded expression with complex temporal dynamics ( Figure 9D and Figure 9-figure supplement 2). This is consistent with observations made from single-cell transcriptome studies in the developing Drosophila optic lobe (Kurmangaliyev et al, 2020;Ozel et al, 2020). Among the CSMs that are differentially expressed in any of the 4 stages, we observed many molecules in protein families that have been implicated in wiring, including Beaten Path (Beat), Dpr, DIP, Dscam, Fasciclin (Fas), and Robo (Figure 9-figure supplement 2) (Kolodkin & Tessier-Lavigne, 2011;Sanes & Zipursky, 2020).…”

“…As a consequence, it may be challenging to identify genes carrying type-specific information at certain timepoints even when sophisticated algorithms are applied, which can lead to underestimation of cell type diversity. Our observation of peaked transcriptome diversity in developing projection neurons has also been observed in the Drosophila optic lobe recently (Ozel et al, 2020). Thus, in order to accurately classify single-cell transcriptomes, especially for connectivity-defined neuronal types such as fly olfactory PNs, it may be a general strategy to first obtain their single-cell transcriptomes during circuit assembly and then use this information to supervise cell-type classification in other developmental stages, including adults.…”

“…By applying both methods, we can confidently track the transcriptomes of the same cell type throughout development and study the unique molecular features of each stage. We note that two other methods for tracing transcriptomes of the same neuronal types across development-batch-correction to cluster same cell types across different stages, and training an artificial neural network to classify cell type-have been applied successfully in recent single-cell transcriptome studies of cells in the developing Drosophila optic lobe (Kurmangaliyev et al, 2020;Ozel et al, 2020).…”

“…In addition, driver lines for specific types of PNs can be made using genes that show consistent expression pattern across different stages (Figure 7-figure supplement 2) to label and genetically manipulate specific PN types. Together with several recent in depth scRNAseq studies of cells in the visual and olfactory system across multiple stages (Jain et al, 2020;Kurmangaliyev et al, 2020;McLaughlin et al, 2020;Ozel et al, 2020), these studies have established foundations of gene expression for Drosophila olfactory and visual systems and should catalyze new biological discoveries.…”

R7 photoreceptor axon targeting depends on the relative levels oflost and foundexpression in R7 and its synaptic partners

“…(1:500, Sigma F3165), mouse anti-Dac (1:40; DSHB mAbdac2-3), rabbit anti-Bsh (1:1800) (Ozel et al, 2020), and guinea pig anti-Runt (1:600; GenScript). Rhodamine-phalloidin (Invitrogen R415) was used at 1:20.…”