Figure 5.

Neuronal knockdown of candidate essential chaperones cause disruption of the presynaptic cytoskeleton. (A–J) Representative images of third instar larval NMJs from muscle 6/7 of A2 hemisegment in (A) control or elav-Gal4-driven (B) CG4183 RNAi, (C) CG15266 RNAi, (D) CG7770 RNAi, (E) CG7048 RNAi, (F) CG17187 RNAi, (G) CG5525 RNAi, (H) CG12101 RNAi, (I) CG8542 RNAi, and (J) CG1242 RNAi animals labeled with mAb22C10. Compared to the control NMJ, elav^C155-Gal4-driven knockdown of above essential chaperones shows a significant reduction in the number of Futsch-positive loops and, in some cases, synapses with broken Futsch loops were also seen (marked with arrows). The Roman numerals in the image correlate the NMJ morphological defect of each panel with the corresponding phenotypic class depicted in Figure 3. Bar in (J) represents 4 µm. (K) Histogram showing quantification of the percentage of boutons containing Futsch loops of control, elav-Gal4/+, CG4183 RNAi, elav-Gal4-driven CG4183 RNAi, CG15266 RNAi, elav-Gal4-driven CG15266 RNAi, CG7770 RNAi, elav-Gal4-driven CG7770 RNAi, CG7048 RNAi, elav-Gal4-driven CG7048 RNAi, CG17187 RNAi, elav-Gal4-driven CG17187 RNAi, CG5525 RNAi, elav-Gal4-driven CG5525 RNAi, CG12101 RNAi, elav-Gal4-driven CG12101 RNAi, CG8542 RNAi, elav-Gal4-driven CG8542 RNAi, CG1242 RNAi, and elav-Gal4-driven CG1242 RNAi. Histogram in black represents controls including w¹¹¹⁸, elav-Gal4/+, and parental lines. Percentage of boutons containing Futsch loops upon pan-neuronal knockdown of sHSP (green), Prefoldins (magenta), Hsp40 (violet), Hsp60 (orange), Hsp70 (dark blue), and Hsp90 (light blue) have been represented. At least eight NMJ synapses from four larvae of each genotype were used for Futsch loop quantification. *** P < 0.0001. Error bars represent SEM (mean ± SEM). Statistical analysis based on one-way ANOVA with post hoc Tukey’s test for multiple comparisons. Hsp, heat shock protein; NMJ, neuromuscular junction; ns, not significant; RNAi, RNA interference; sHSP, small HSP.