Fig. 5.
scat interacts genetically with Rab5, Rab7, and Rab11 to control NMJ development. (A) NMJ phenotypes caused by the motor neuron-specific knockdown of scat expression by RNAi are suppressed by overexpression of wild-type and dominant-negative transgenes for Rab5, Rab7 and Rab11 (Rab11 images are not shown). An inducible transgenic shRNA targeting luciferase (UAS-LUC.VALIUM10) or scat (UAS-TRiPHMS01910) was expressed in motor neuron using the C380-Gal4 driver in combination with an inducible YFP-tagged wild-type or dominant negative Rab5, Rab7, or Rab11 (UAS-YFP:Rab). NMJs at muscle 6/7 in body segment A3 in wandering third instar larvae were stained with antibodies targeting Dlg (green) and HRP (red). Images show maximum Z-projections. The boxed areas are blown up in C to show altered synaptic bouton or PSD morphologies. Scale bar: 20 µm. (B) As shown in Fig. 2, the total bouton number/MSA (normalized to the respective control) are significantly increased by presynaptic scat knockdown. This phenotype is suppressed by co-expression of wild-type and dominant negative Rabs. C380>scat shRNA, Rab7 (DN) double mutant NMJs are significantly smaller. N=18, 12, 16, 13, 22, 17, 21, 17, 18, 20, 20, 22, 21 and 23. (C) Boxed areas in A. Many C380>scat shRNA, Rab5 (WT) NMJs have a clustered bouton phenotype similar to many endocytic mutants. Dlg staining and bouton morphology is significantly disrupted in C380>scat shRNA, Rab7 (DN) double mutants. Data are represented as the mean±s.e.m. Unless otherwise indicated, all comparisons have been made to the control. Statistical analysis was done using a one-way ANOVA followed by a Holm-Sidak multiple comparison test. *P<0.05, **P<0.01, ***P<0.001. ****P<0.0001.