NanoDam identifies novel temporal transcription factors conserved between the Drosophilacentral brain and visual system

Temporal patterning of neural progenitors is an evolutionarily conserved strategy for generating neuronal diversity. Type II neural stem cells in the Drosophilacentral brain produce transit-amplifying intermediate neural progenitors (INPs) that exhibit temporal patterning. However, the known temporal factors cannotaccount for the neuronal diversity in the adult brain. To search for new temporal factors, we developed NanoDam, which enablesrapid genome-wide profiling of endogenously-tagged proteins in vivowith a singlegenetic cross.Mapping the targets of known temporal transcription factorswith NanoDamidentified Homeobrain and Scarecrow (ARX and NKX2.1 orthologues) as novel temporal factors. We show that Homeobrainand Scarecrow define middle-aged and late INP temporal windows and play a role in cellular longevity. Strikingly, Homeobrain and Scarecrow have conserved functions as temporal factors in the developing visual system.NanoDam enables rapid cell type-specific genome-wideprofiling with temporal resolution and can be easily adapted for use in higher organisms. To establish and assess the efficiency and reproducibility of NanoDam, Chronophage-NanoDam was compared to Chronophage-Targeted DamID in Neural Stem Cells of Drosophila embryos. NanoDam was performed for Dichaete, Grainyhead and Eyeless in TypeII Neural Stem Cell-derived Intermediate Progenitors of the Drosophila L3 larval brain to identify novel temporal factors. scRNAseq was conducted to assess expression of the respective temporal factor candidates in Intermediate Progenitors and their offspring. In parallel, NanoDam for Dichaete and Eyeless in the Optic lobe medulla Neural Stem Cells allowed a comparative analysis between the two temporal cascades.