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Fig. 5. Cytochalasin D treatment promotes processes outgrowth in Drosophila S2 cells. Microtubule staining (A) and phalloidin (B) staining of S2 cells on concanavalin (ConA)-coated coverslips in the present of cytochalasin D. These processes contained long and unfragmented microtubules, as in A. Furthermore, the presence of actin patches confirmed that cytochalasin D destroys the network of actin filament, as in B. (Scale bar, 10 m m.)

Movie 1. dFMR-containing RNP granules move bidirectionally in thin processes formed by S2 cells in the presence of cytochalasin D. The time-lapse movies were taken at 2-sec intervals. Anterograde and retrograde movement and bidirectional oscillations can be observed in this sequence.

Movie 2. The movements of dFMR-containing RNP granules were dramatically reduced when KHC was knocked down by RNAi.

Fig. 6. Microtubule staining on S2 cells in various RNAi treatment. The staining shows that RNAi has no effect on microtubule distribution: Mock (A), KHC-RNAi (B), and DHC-RNAi (C). (Scale bar, 10 m m.)

Fig. 7. Movements of GFP-peroxisome in various RNAi treatments. (a) Immunoblotting confirmed the specificity and efficiency of RNAi treatment. Control extracts were diluted to one-half, one-fourth, one-eighth, 1/16th, and 1/32nd to estimate the knockdown efficiency. The antibodies used were indicated at the left. The target proteins were consistently knockdown to less than 10%. Histograms of anterograde velocity (b), and retrograde velocity (c) for GFP-peroxisome in various RNAi treatments. Two controls, Lac-I and Klp61F-RNAi, were used and showed comparable results, confirming the specificity of RNAi effect. Either KHC or KLC knockdown is sufficient to impede the movement bidirectionally, indicating peroxisome translocation requires both KHC and KLC. Note that peroxisome movement is slightly biased toward anterograde movement.