Figure 9.

KIF3C regulates microtubule dynamics and organization in the growth cone. A, Immunofluorescence images of control or KIF3C knockdown growth cones immunostained for TUBB3. High magnification of the boxed regions shows the arrangement of microtubules within the periphery of the growth cones. Scale bar, 10 μm. B, Quantification of the percentage of control and KIF3C knockdown growth cones containing looped microtubules (n = 102 and n = 121, respectively, compiled from three independent experiments). Error bars indicate SD. ***p < 0.001. C, Spinning-disk confocal live-cell imaging of adult DRG growth cones cotransfected with control, Kif3c shRNA, or mRFP-Kif3c and MACF43-GFP constructs. Images were collected for 100 s with a 0.2 s interval between frames. Left to right, Single frame of the MACF43-GFP channel; maximum intensity projection of the MACF43-GFP channel >500 frames (100 s). D, Representative kymographs illustrating the growth of MACF43-GFP-decorated microtubule tips in control, KIF3C-depleted, and KIF3C-overexpressing growth cones. Sudden shrinkage events are seen in the control and mRFP-KIF3C recordings, whereas the KIF3C knockdown shows steady elongation with no catastrophes. E–G, Quantification of the duration of growth, frequency of catastrophes, and the instantaneous growth velocity (growth rate) in control, KIF3C-depleted, and KIF3C-overexpressing growth cones. Fifty to 150 growth episodes in 10 growth cones were analyzed in each condition. Data for mRFP control and control shRNA showed no significant differences and were pooled. Error bars indicate SEM. **p < 0.01. ***p < 0.001.