Identification of Neuronal Lineages in the Drosophila Peripheral Nervous System with a “Digital” Multi-spectral Lineage Tracing System

Abstract

SUMMARY Elucidating cell lineages provides crucial understanding of development. Recently developed sequencing-based techniques enhance the scale of lineage tracing but eliminate the spatial information offered by conventional approaches. Multi-spectral labeling techniques, such as Brainbow, have the potential to identify lineage-related cells in situ. Here, we report nuclear Bitbow (nBitbow), a “digital” version of Brainbow that greatly expands the color diversity for scoring cells, and a suite of statistical methods for quantifying the lineage relationship of any two cells. Applying these tools to the Drosophila peripheral nervous system (PNS), we determined lineage relationship between all neuronal pairs. This study demonstrates nBitbow as an efficient tool for in situ lineage mapping, and the complete lineage relationship among larval PNS neurons opens new possibilities for studying how neurons gain specific features and circuit connectivity.

Light microscopy based approach for mapping connectivity with molecular specificity

“…We can already observe tens of easily distinguishable color barcodes, which can be exponentially increased in the future by probing more FPs. For example, five FPs were used in the recently developed Bitbow, a "digital" format of Brainbow 40 . Expansion of the sample is important for increasing imaging resolution, but comes at the cost of diluting antibody signal.…”

Light microscopy based approach for mapping connectivity with molecular specificity

“…We can already observe tens of easily distinguishable color barcodes, which can be exponentially increased in the future by probing more FPs. For example, 5 FPs were used in the recently developed Bitbow, a "digital" format of Brainbow 39 . Expansion of the sample is important for increasing imaging resolution, but comes at the cost of diluting antibody signal.…”

The molecular landscape of neural differentiation in the developing Drosophila brain revealed by targeted scRNA-seq and multi-informatic analysis

“…Genetic constructs based around CRISPR-Cas9 (Raj et al, 2017;Spanjaard et al, 2018) and the Cre/Lox system (Kalhor et al, 2018;Pei et al, 2017;Weber et al, 2016) have been developed for this purpose, although which exact lineage was labeled by a particular barcode was still unknown. The introduction of a spectrally unique barcode for each neuroblast lineage, in a similar vein to the recently developed Bitbow lineage tracking strategy (Li et al, 2020;Veling et al, 2019), would be advantageous as they can provide direct in situ evidence for neuroblast lineage identity.…”