Hook1 localizes to distal axon, while other dynein effectors are enriched in other compartments. (A) Representative images of neurons expressing indicated dynein effectors. White arrows point to effector puncta. Dashed line represents the line used for line scan plots , but line is shifted below the axon so as to not obscure primary data. Line scan graphs of mid- and distal axons are shown to the right of the image. Black arrows point to corresponding puncta peaks. (B) Quantification of percentage of neurons with puncta in soma. Bar graph shows mean ± SEM; one-way ANOVA (***, P ≤ 0.0001; ns, P = 0.3964). n = 3–4 individual experimental averages, 23–26 cells. (C) Graphs of dynein effector enrichment in mid versus distal axons. Red lines indicate enrichment in the distal region of axon. Black lines indicate enrichment in the mid-axon. BICD2: n = 21 neurons; RILP: n = 27 neurons; Hook1: n = 27 neurons. (D) Graph of dynein effector puncta per growth cone. Scatter plot shows mean ± SEM; one-way ANOVA (***, P < 0.0001; ns, P = 0.8901). n = 16–23 growth cones. (E) Representative images of proximity ligation assay of Hook1-Halo–expressing distal axons with and without anti-DIC. Hook1–DIC complexes are presented in green and GFP in magenta. Arrow indicates Hook1–DIC complexes. (A and E) Bars, 10 µm. (F) Quantification of distal axons with Hook1–DIC PLA puncta present. Bar graph shows mean ± SEM; unpaired t test (*, P = 0.0119). n = 2 individual experimental averages, 19–38 neurons.