Figure 5. Presynaptic reduction of dTip60 in the nervous system leads to suppression of active zone marker bruchpilot at larval NMJs.

Confocal imaging analysis of larval boutons on muscles 6/7 at abdominal segment 4 immunohistochemical double labeled with HRP antibody (green) that labels the entire presynaptic membrane and nc82 antibody (red) that recognizes bruchpilot, a protein associated with functional active zones and neurotransmission. (A,B) is w¹¹¹⁸ (C,D) is dTip60^E431Q (E,F) is dTip60^RNAi. In the analyses, w¹¹¹⁸ genotype is represented by 20 larval preparations (n = 20), dTip60^RNAi (n = 18) and dTip60^E431Q (n = 21). Scale bar is 5 um. Quantification of the fluorescence intensity as a measure of nc82 abundance at the NMJ muscle 6/7 of segment A4. The relative fluorescence intensities were measured as number of pixels per measured area and represented as % maximum intensities. Fluorescence intensity was determined by first quantitatively measuring the intensity profiles of all the nc82 punctae in a given marked area of the boutons and then calculating the average intensity of the brightest punctae in all of the samples as given by their intensity values. The nc82 intensities were normalized against HRP fluorescence intensities and corrected for background before all intensity measurements. Histogram depicts quantitative analysis of antibody fluorescence intensity where (G) is percentage maximum intensity of antibody stain in all boutons (H) is percentage maximum intensity of antibody stain in Is boutons. Asterisks (*) indicates statistically significant difference in relation to control where double asterisks indicate p≤0.001 and three asterisks indicate p≤0.0001. All error bars depict standard error of the mean.