Figure 1.

Dendrite arborization is disrupted by charge-reversal mutagenesis of E177 in β5-loop 8 of Khc. (A) Engineered Khc locus in which Khc is replaced by an attP site. (B) Cartoon of kinesin-1 highlighting β5-loop 8 (green), loop 11 (orange), and loop 12 (blue), which were replaced by equivalent loops from unc-104. β5-loop 8 is highly conserved between flies and humans. TERF sequence is underlined; arrow indicates Khc E177, also highlighted in red. (C–H) Representative images of control (con) and mutant neurons at 72 and 96 h AEL. Bar, 50 µm. (I) Quantification of dendrite length and branch number (mean ± SD) at 72 h AEL of 11 (Khc^E177A/−), 13 (Khc^E177K/−), and 16 (control, Khc-RNAi) neurons in at least three larvae and at 96 h AEL of 10 neurons (control, Khc-RNAi, Khc^E177K/−) in at least three larvae. ****, P < 0.0001 relative to control and evaluated by one-way ANOVA and Tukey post hoc test (top and middle). Sholl analysis (bottom) of dendrite arbors at 72 h AEL (mean ± SEM). Critical radius and maximum branches reported in Table 2. (J) Zoomed-in view of proximal axons. The axons of Khc^E177K/− and Khc-RNAi–expressing neurons frequently sprout ectopic branches (arrows). Bar, 10 µm. (K) Percentage of axons with ectopic branches in control, Khc-RNAi, and Khc^E177K/− neurons (n = 18, all genotypes).