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. 2021 Dec 17;15:784649. doi: 10.3389/fnins.2021.784649

FIGURE 1.

FIGURE 1

Expression of VAChT/ChAT cholinergic locus in the AMG5 neuron. (A) Cartoon schematic of the VAChT/ChAT locus encoding VAChT and ChAT from two mutually exclusive, alternatively spliced transcript variants sharing the same non-coding exon 1 (purple bar). Dotted line indicates additional exons encoding ChAT not shown. Colored bars underneath locus indicate cis-regulatory sequences tested, numbers reflect distance from the start codon (+ 1) of VAChT. Expression in “core” motor ganglion (MG) and Ascending Motor Ganglion neuron 5 (AMG5) was assessed qualitatively. An Ebf binding site previously reported (Kratsios et al., 2012) is situated -4111 bp upstream of VAChT. (B) Co-electroporation of VAChT/ChAT -4315/ + 15 > Unc-76:mCherry and Msx > H2B:GFP revealed co-expression in AMG5 at 19 h post-fertilization (hpf) at 20°C, a cholinergic interneuron of the dorsal MG, derived from the dorsal cells of the neural tube. (C) A minimal fragment surrounding the Ebf binding site is sufficient to drive AMG5-specific reporter expression at 18 hpf, 20°C. Image is a confocal Z stack projection. (D) A smaller proximal fragment from VAChT/ChAT (-2083/ + 15) lacking the Ebf binding site is sufficient to drive expression in other cholinergic neurons including those of the core MG, but not in AMG5, at 19 hpf, 18°C.