Optogenetic cleavage of the Miro GTPase reveals the direct consequences of real-time loss of function in Drosophila
Fig 7
Optogenetic control of Drosophila locomotor behaviour in Split-Miro flies.
(A) Adult miroSd32/B682 flies eclosed after Appl-Gal4-driven expression of UAS-wt-Miro, UAS-Split-Miro or UAS-Split-Miro + UAS-EGFP-SNPH (UAS-SNPH) were counted from 6, 5, and 3 independent crosses, respectively. Data are reported as percentage of flies expected from mendelian ratios. Data are mean ± SEM, one-way ANOVA with Tukey’s post hoc test showed no statistical difference. (B) Schematic of the “Opto-DART” behavioural setup consisting of a custom-made optogenetic enclosure equipped with LEDs for blue light stimulation and a camera to record fly activity. Flies are transferred to a 1D platform for automated recording of activity using the DART system [45,46]. DAC: Digital-to-Analog Converter for multiplatform integration. (C-H) Overall activity, action initiation, and average speed of 2-day-old flies (C-E) and 8-day-old flies (F-H) expressing UAS-wt-Miro or UAS-Split-Miro, before, during, and after blue light exposure (shaded blue rectangles). In (C) and (F), the average activity of flies expressing UAS-wt-Miro vs. UAS-Split-Miro is: before light exposure, 3.78% ± 0.94 vs. 2.34% ± 0.72 (p = 0.11) (C) and 5.62% ± 1.02 vs. 8.7% ±1.81 (p = 0.14) (F); under blue light, 9.52% ± 0.96 vs. 7.32% ± 0.81 (p = 0.13) (C) and 8.45% ± 0.82 vs. 21.87% ± 2.07**** (F); after blue light exposure, 3.36% ± 1.18 vs. 7.85% ± 1.96 (p = 0.13) (C) and 3.23% ± 1.1 vs. 13.19% ± 2.39** (F). (I, K) Overall activity, action initiation, and average speed of 8-day-old flies expressing UAS-Split-Miro or UAS-Split-Miro + UAS-SNPH, before, during, and after blue light exposure (shaded blue rectangles). In (I), the overall activity of flies expressing UAS-Split-Miro vs. UAS-Split-Miro + UAS-SNPH is: 6.09% ± 0.89 vs. 0.95% ± 0.35**** before light exposure; 18.62% ± 1.4 vs. 3.55% ± 0.75**** under blue light; 9.9% ± 1.4 vs. 3.91% ± 1.25* after blue light exposure. In (C-K), n = number of flies. Values are means ± SEM, Mann–Whitney test (C, F, I) and multiple unpaired t test (D, E, G, H, J, K) with Holm–Sidak correction for multiple comparisons. * p < 0.05, ** p < 0.01, **** p < 0.0001. The data underlying the graphs shown in the figures can be found in S1 Data.