Figure S6.
Validation of UAS-RNAi lines targeting NMIIHC/Zip and NMIILC/Sqh. (A) Distribution of Sqh::GFP (cyan) in axons (labeled with HRP in magenta) proximal to muscle 4 in Sqh::GFP/+ heterozygous larvae, and larvae expressing in neurons two independent UAS-RNAi lines (Zip65947 and Zip36727) targeting NMIIHC/Zip, as well as the NMIILC/Sqh targeting lines Sqh32439 and Sqh33892. (B) Quantification of the Sqh::GFP fluorescence in axons of larvae with the different genotypes shown in A. (C) Example of image analysis in Fiji, where the Sqh::GFP fluorescence was measured within the HRP-delineated NMJ ROI, in a 1-µm rim surrounding the bouton (postsynaptic ROI), and in the muscle ROI (rectangle with yellow dashed lines) (excluding the HRP-delineated area and the surrounding 1-µm rim). (D) Quantifications of the Sqh::GFP fluorescence at NMJs from larvae carrying Zip65947 in the absence and presence of the neuronal C155-Gal driver, in an experiment validating the lack of nonspecific expression of Zip65947 in neurons and muscles in the absence of the Gal4 driver. (E) Immunostaining of endogenous Zip (cyan) at NMJs (HRP magenta) and muscles of larvae expressing C57-Gal4–driven control and RNAi lines targeting NMIIHC/Zip and NMIILC/Sqh, respectively. (F and G) Quantifications of muscle Zip fluorescence, and (G) Zip particle abundance, size, and brightness in larvae expressing C57-Gal4–driven CTRL35785, Zip65947, and Sqh32439. Box-and-whisker graphs were used to represent the results of the area and fluorescence intensity of the individual Zip particles. Whiskers represent 10th to 90th percentile, while the rest of the data points are shown as individual values. The y axis in these graphs represents log10, to capture the broad distribution of the individual values. In bar graphs with linear y axis, error bars represent the SEM. N—axons in B, NMJs in D and G, muscle ROIs in F. N in box and whiskers—individual Zip particles. **P < 0.01, ***P < 0.001 after one-way ANOVA with the Kruskal–Wallis multiple comparisons test or upon the unpaired, nonparametric Mann–Whitney test. Scale bar in A—5 µm; all the rest—2 µm. See Table S1 for detailed genotypes and N.

Validation of UAS-RNAi lines targeting NMII HC /Zip and NMII LC /Sqh. (A) Distribution of Sqh::GFP (cyan) in axons (labeled with HRP in magenta) proximal to muscle 4 in Sqh::GFP/+ heterozygous larvae, and larvae expressing in neurons two independent UAS-RNAi lines (Zip65947 and Zip36727) targeting NMIIHC/Zip, as well as the NMIILC/Sqh targeting lines Sqh32439 and Sqh33892. (B) Quantification of the Sqh::GFP fluorescence in axons of larvae with the different genotypes shown in A. (C) Example of image analysis in Fiji, where the Sqh::GFP fluorescence was measured within the HRP-delineated NMJ ROI, in a 1-µm rim surrounding the bouton (postsynaptic ROI), and in the muscle ROI (rectangle with yellow dashed lines) (excluding the HRP-delineated area and the surrounding 1-µm rim). (D) Quantifications of the Sqh::GFP fluorescence at NMJs from larvae carrying Zip65947 in the absence and presence of the neuronal C155-Gal driver, in an experiment validating the lack of nonspecific expression of Zip65947 in neurons and muscles in the absence of the Gal4 driver. (E) Immunostaining of endogenous Zip (cyan) at NMJs (HRP magenta) and muscles of larvae expressing C57-Gal4–driven control and RNAi lines targeting NMIIHC/Zip and NMIILC/Sqh, respectively. (F and G) Quantifications of muscle Zip fluorescence, and (G) Zip particle abundance, size, and brightness in larvae expressing C57-Gal4–driven CTRL35785, Zip65947, and Sqh32439. Box-and-whisker graphs were used to represent the results of the area and fluorescence intensity of the individual Zip particles. Whiskers represent 10th to 90th percentile, while the rest of the data points are shown as individual values. The y axis in these graphs represents log10, to capture the broad distribution of the individual values. In bar graphs with linear y axis, error bars represent the SEM. N—axons in B, NMJs in D and G, muscle ROIs in F. N in box and whiskers—individual Zip particles. **P < 0.01, ***P < 0.001 after one-way ANOVA with the Kruskal–Wallis multiple comparisons test or upon the unpaired, nonparametric Mann–Whitney test. Scale bar in A—5 µm; all the rest—2 µm. See Table S1 for detailed genotypes and N.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal