Figure 1
From: Nervous system-wide profiling of presynaptic mRNAs reveals regulators of associative memory
Isolation of presynaptic compartments for RNA-seq. (A) Neuronal cell isolation results in fragmentation of cells, where cell bodies (left) can be detected separately from neurites (right). (B) Schematic of the dual-fluorescent protein strategy for isolating synaptic and somatic compartments in C. elegans. (C) Representative confocal images of rab-3p::mCherry; rab-3p::RAB-3::GFP transgenic worms. Neurons in the head (outlined in white) and tail (outlined in gray) show distinct expression of fluorescent proteins. Cell bodies (outlined in white, arrows) express somatic mCherry (red), while nerve ring synapses exclusively express GFP (green, arrows. Colocalization of the two fluorescent proteins (yellow) is also evident. (D) FACS plot displaying ability to isolate synaptic (GFP+, green), somatic (mCherry+, red), and synaptic + somatic (GFP+/mCherry+, blue) compartments using adult neuron cell isolation method18. (E,F) Principle components analysis of six mCherry+ (red) somatic and GFP+ (green) synaptic RNA-seq samples performed as part of DESeq2 analysis. (E) Principle component analysis of four synaptic (green) and six somatic (red) RNA-seq samples after removal of outliers. Remaining samples cluster by subcompartment.
