Figure 2.
DCX-EMAP is required for fly mechanotransduction. (A) Schematic diagrams of DCX-EMAPKI, DCX-EMAPKO, and the domain organization of DCX-EMAP. The blue bar indicates the promoter sequence (1.5 kb) used to generate the DCX-EMAP-gal4 strain. The red bar marks the fragment of DCX-EMAP (amino acids 258–461, 204 amino acids) used as the antigen to generate the DCX-EMAP antibody (Liang et al., 2014). (B)DCX-EMAP expressed in fly mechanosensory organs. Left: Haltere campaniform mechanoreceptors (red arrowhead). Middle: Leg campaniform and bristle mechanoreceptors (red arrowheads). Right: The Johnston’s organ (blue arrowhead) and olfactory receptor cells (green arrowhead) in fly antenna (also see Fig. S2). Genotype: DCX-EMAP-gal4; uas-cd4-tdgfp. Scale bars, 50 µm. (C) Flight tests of wild type (n = 73 flies), DCX-EMAPKO (n = 151 flies), nompC3 (n = 18 flies), two rescue strains (rescue-1: DCX-EMAP-gal4/uas-gfp-DCX-EMAP; DCX-EMAPKO, n = 71 flies. rescue-2: nompC-gal4/uas-gfp-DCX-EMAP; DCX-EMAPKO, n = 42 flies), DCX-EMAPKI (n = 66 flies), and DCX-EMAPOE (DCX-EMAP-gal4/uas-gfp-DCX-EMAP, n = 54 flies). The schematic for the assay is shown in the left panel. (D) Crawling tests of wild type (n = 18 flies), DCX-EMAPKO (n = 11 flies), nompC3 (n = 13 flies), two rescue strains (rescue-1: n = 10 flies; rescue-2: n = 7 flies), DCX-EMAPKI (n = 15 flies), and DCX-EMAPOE (n = 12 flies). The schematics for the assay are shown in the left panels. Scale bar, 2 mm. Also see Video 3. (E) Representative images of the in vivo calcium recording in wild type (DCX-EMAP-gal4, uas-cd4-tdtom/uas-GCamp6s), DCX-EMAPKO (DCX-EMAP-gal4, uas-cd4-tdtom/uas-GCamp6s; DCX-EMAPKO), and rescue (DCX-EMAP-gal4, uas-cd4-tdtom, uas-DCX-EMAP/uas-GCamp6s; DCX-EMAPKO). Scale bar, 10 µm. (F) Statistical quantification of the calcium responses. Wild type (n = 8 cells), DCX-EMAPKO (n = 6 cells), and rescue-1 (n = 8 cells). In D and F, data are presented as mean ± SD with scattered data points. Two-sided unpaired Student’s test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; n.s., no significance.

DCX-EMAP is required for fly mechanotransduction. (A) Schematic diagrams of DCX-EMAPKI, DCX-EMAPKO, and the domain organization of DCX-EMAP. The blue bar indicates the promoter sequence (1.5 kb) used to generate the DCX-EMAP-gal4 strain. The red bar marks the fragment of DCX-EMAP (amino acids 258–461, 204 amino acids) used as the antigen to generate the DCX-EMAP antibody (Liang et al., 2014). (B)DCX-EMAP expressed in fly mechanosensory organs. Left: Haltere campaniform mechanoreceptors (red arrowhead). Middle: Leg campaniform and bristle mechanoreceptors (red arrowheads). Right: The Johnston’s organ (blue arrowhead) and olfactory receptor cells (green arrowhead) in fly antenna (also see Fig. S2). Genotype: DCX-EMAP-gal4; uas-cd4-tdgfp. Scale bars, 50 µm. (C) Flight tests of wild type (n = 73 flies), DCX-EMAPKO (n = 151 flies), nompC3 (n = 18 flies), two rescue strains (rescue-1: DCX-EMAP-gal4/uas-gfp-DCX-EMAP; DCX-EMAPKO, n = 71 flies. rescue-2: nompC-gal4/uas-gfp-DCX-EMAP; DCX-EMAPKO, n = 42 flies), DCX-EMAPKI (n = 66 flies), and DCX-EMAPOE (DCX-EMAP-gal4/uas-gfp-DCX-EMAP, n = 54 flies). The schematic for the assay is shown in the left panel. (D) Crawling tests of wild type (n = 18 flies), DCX-EMAPKO (n = 11 flies), nompC3 (n = 13 flies), two rescue strains (rescue-1: n = 10 flies; rescue-2: n = 7 flies), DCX-EMAPKI (n = 15 flies), and DCX-EMAPOE (n = 12 flies). The schematics for the assay are shown in the left panels. Scale bar, 2 mm. Also see Video 3. (E) Representative images of the in vivo calcium recording in wild type (DCX-EMAP-gal4, uas-cd4-tdtom/uas-GCamp6s), DCX-EMAPKO (DCX-EMAP-gal4, uas-cd4-tdtom/uas-GCamp6s; DCX-EMAPKO), and rescue (DCX-EMAP-gal4, uas-cd4-tdtom, uas-DCX-EMAP/uas-GCamp6s; DCX-EMAPKO). Scale bar, 10 µm. (F) Statistical quantification of the calcium responses. Wild type (n = 8 cells), DCX-EMAPKO (n = 6 cells), and rescue-1 (n = 8 cells). In D and F, data are presented as mean ± SD with scattered data points. Two-sided unpaired Student’s test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; n.s., no significance.

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