Single cell RNA-seq analysis reveals temporally-regulated and quiescence-regulated gene expression in Drosophila larval neuroblasts

Abstract

The mechanisms that generate neural diversity during development remains largely unknown. Here, we use scRNA-seq methodology to discover new features of the Drosophila larval CNS across several key developmental timepoints. We identify multiple progenitor subtypes – both stem cell-like neuroblasts and intermediate progenitors – that change gene expression across larval development, and report on new candidate markers for each class of progenitors. We identify a pool of quiescent neuroblasts in newly hatched larvae and show that they are transcriptionally primed to respond to the insulin signaling pathway to exit from quiescence, including relevant pathway components in the adjacent glial signaling cell type. We identify candidate “temporal transcription factors” (TTFs) that are expressed at different times in progenitor lineages. Our work identifies many cell type specific genes that are candidates for functional roles, and generates new insight into the differentiation trajectory of larval neurons.

Eight principal chromatin states functionally segregate the fly genome into developmental and housekeeping roles

“…g. Pal2 42 , GABA-B-R2 43 ) . Reanalysis of recent scRNA-seq data for the Drosophila larval brain 39 confirmed that these MRG15-enriched genes were silent in NSCs and broadly expressed across mature neuron subtypes (Fig. 5d).…”

“…These compartments can be further divided into sub-compartments, partially separated by histone modifications 46 , and we wondered whether TADs of similar chromatin state would also exhibit longrange associations within the 3D genome. Reprocessing existing HiC 47 and Lamin B binding Although not significantly enriched for specific ontologies, investigation of the highest-ranking genes transitioning from Yellow-R in NSCs to Yellow states in neurons revealed a number of genes involved in synaptic structure and signalling, with scRNA-seq data (reanalysed from 39 ) confirming the change in expression status between cell types (d).…”

“…(a) Assigning all genes in NSCs and neurons a chromatin state based on modal state occupancy reveals extensive chromatin remodelling during differentiation, including (b) genes transitioning between active Swi/Snf and repressive states (i) and between active and repressive forms of Yellow chromatin (ii). Functional roles of different transitions are illustrated via GO term enrichments (c).Although not significantly enriched for specific ontologies, investigation of the highest-ranking genes transitioning from Yellow-R in NSCs to Yellow states in neurons revealed a number of genes involved in synaptic structure and signalling, with scRNA-seq data (reanalysed from39 ) confirming the change in expression status between cell types (d).…”

“…Functional roles of different transitions are illustrated via GO term enrichments (c). Although not significantly enriched for specific ontologies, investigation of the highest-ranking genes transitioning from Yellow-R in NSCs to Yellow states in neurons revealed a number of genes involved in synaptic structure and signalling, with scRNA-seq data (reanalysed from 39 ) confirming the change in expression status between cell types (d).…”

“…Seurat v4 69 was used to manually annotate the Seurat object from a published scRNA-seq dataset of the larval brain 39 , and identify NSCs, cholinergic, glutamatergic, and GABAergic neuronal clusters using the established markers indicated in Fig. 5d.…”