Figure 5

Dynamic behaviour of LD is both microtubule and microfilament dependent. (A) LD size distribution in embryos after culture for 22 hours in cytochalasin D and nocodazol. Control embryos were cultured in DMSO (carrier). There is no difference in the distribution of LD sizes (n = 4 embryo for each condition). (B, C, D) Extended focus projections of THG signal from representative control, Cytochalasin D and Nocodazole treated embryos, showing no obvious difference in LD. (E, E') Extended focus projections of confocal scans of fluorescence from DAPI stained nuclei (magenta) in control (E) and nocodazole (E') treated embryos. As expected, nuclei in the later are arrested at the metaphase stage, as seen by the condensed chromosomes. (F, F') Optical confocal section of fluorescence from F-actin stained with Phalloidin (grey) and nuclei stained with DAPI (magenta), in control and cytochalasin D treated embryos. As expected, F-actin localisation is disrupted in the latter. (G, H) Change in average LD size and number over time in embryos cultured in the presence of cytochalasin D and nocodazole (n = 12 sections from 4 embryos for each condition). Both inhibitors cause a clear delay in the formation of larger LD. The inset in (H) shows the estimated average velocity of LD. Treatment with the two inhibitors causes a significant reduction to LD velocity as compared to control LD (p < 0.0001, Students T-test). Scale bar in (B) = 20 μm and applies to (B, C and D). Scale bar in (E) = 20 μm and applies to (E, E', F and F')