Fig. 1: Unc13A and Un13B localization at active zones of ePN and iPN presynapses.
From: Unc13A and Unc13B contribute to the decoding of distinct sensory information in Drosophila
a Expression of myr-RFP in iPNs using MZ699-Gal4 (UAS/Gal4 system) and mCD8-GFP in ePNs using GH146-QF (QUAS/QF system). Image shows maximal projection fluorescence intensity across a confocal stack of a whole brain, immunostained against GFP (green), RFP (red), and Unc13B (blue). Lateral horn, MB calyx, iPN, and axonal tracks are indicated (n = 2, experiment was performed once). b Expression of UAS: brp-short GFP under control of GH146-Gal4 (n = 5, experiment was performed two times). Images represent single focal planes from calyx and LH regions immunostained against Unc13A (red), Unc13B (green), and GFP (blue). c, d Expression of UAS: brp-short GFP using GH146-Gal4 (c) and MZ699-Gal4 (d). Images represent single focal planes from calyx and LH regions immunostained against Unc13A or Unc13B (red), Drep2 (magenta), and GFP (green) (n = 5, experiment was performed two times). e–i Time-gated STED (gSTED) analysis of Unc13A and B nano-distribution at LH and MB calyx synapses. e Confocal (left) and gSTED (right) images of BRP Nc82 (green) and the GFP-labeled Ca2+ channel subunit sfCac (red) at LH AZs. Insets show a planar AZ with sfCac localizing at the center of BRP rings (upper) and a vertical AZ (lower) (n = 3, experiment was performed once). f gSTED images of Unc13A or Unc13B (green) and sfCac (red) at PN:KC synapses in the MB calyx (n = 3, experiment was performed once). g, h Confocal (left) and gSTED (right) images of Unc13A (g) or Unc13B (h) (green) and sfCac (red) in the LH. Insets show a Drep2-positive excitatory synapse (upper) and a Drep2-negative inhibitory synapse (lower) from triple-channel gSTED images. i Mean Unc13A- and Unc13B-sfCac distances at Drep2-positive and Drep2-negative LH synapses (n = 7 brains/group). Drep2-positive synapses: Unc13A-sfCac (61 ± 1 nm) versus Unc13B-sfCac (101 ± 4 nm). Drep2-negative synapses: Unc13A-sfCac (60 ± 2 nm) versus Unc13B-sfCac (102 ± 4 nm). Values represent mean ± SEM. Statistics: **p < 0.01, Kruskal–Wallis test followed by two-tailed Dunn´s multiple comparison test. White squares: magnified regions. White arrowheads: region of interest. Scale bars: a 50 μm, b 10 μm, c–d 2 μm, e, g, h 1 μm, insets: 200 nm, f 200 nm. Source data including the exact sample sizes and the p values are provided as a Source Data file. See also Supplementary Fig. 1 and 2.
