Fig. 4

Kif20b mutant axons have reduced enrichment of Shootin1 in the growth cone. A Immunostaining for endogenous Shootin1 with anti-Shootin1 antibody [15] shows Shootin1 enriched in the axonal growth cone of a control (+/+) Stage 3 neuron. A’ Linescan of Shootin1 staining intensity from image in A, starting from the tip of the axon and extending 20 μm shows a peak at the growth cone that flattens in the axon shaft. B Shootin1 immunostaining reveals Shootin1 in the soma of a polarized Kif20b mutant (-/-) neuron, but little enrichment in the distal axon. B’ Linescan of Shootin1 staining intensity from image in b shows a severely reduced peak of Shootin1 at the tip of the mutant axon. C Averaged line scans of anti-Shootin1 signal intensity of 78 +/+, 101 +/-, and 100 -/- axons from 3 independent experiments show significantly bigger peaks in control axons than mutants. (+/+, black circles; +/-, gray triangles; -/-, light gray squares). *, p <0.05; **, p <0.01; *** p <0.001, t-test. Solid brackets compare wild-type (+/+) with mutant (-/-) for all points under each bracket. Dashed bracket compares heterozygous controls (+/-) with mutant (-/-) for all points under the bracket. Line compares +/+ with +/- for all points under the line. D, E, and F Tau immunostaining and axonal linescans reveal no significant difference in distribution or intensity between control and mutant neurons. D’, E’, and G. DCX (doublecortin) immunostaining and axonal linescans show similar high distal distributions and intensities in control and mutant neurons. n = 40 +/- and 40 -/- cells for both Tau and DCX linescans from two independent culture experiments (2 animals each, 2 coverslips from each animal). Scale bar = 10 μm for A and B. Scale bar =20 μm for D and E n.s., not significant, t-test