Fig. 7: p.Ile678Leu hKIF21B variant impairs ipsilateral axon collaterals formation. | Nature Communications

Fig. 7: p.Ile678Leu hKIF21B variant impairs ipsilateral axon collaterals formation.

From: Mutations in the KIF21B kinesin gene cause neurodevelopmental disorders through imbalanced canonical motor activity

Fig. 7

a Representative images of ipsilateral P8 cortices after IUE, with pCAG2-Scarlet and indicated NeuroD-IRES-GFP constructs. Rescue experiments were done by co-expressing mutated p.Ile678Leu hKIF21B together with increasing amount of WT-hKIF21B (at 0.25 µg/µL (WT 0.25) or 1 µg/µL (WT 1)) or equivalent amount of hKIF21B-ΔATP or hKIF21B-T96N, that both lost their processivity. b Close-up views of red boxed area in (a) showing axon ipsilateral branching within layer V for all the indicated conditions. c–e Histograms (means ± S.E.M.) showing (c, d) the quantification of the ipsilateral branching in layer V (intensity of scarlet signal in layer V (blue box) normalized on the intensity of the scarlet signal in the corpus callosum (red box) — as shown on the schematic in the upper panel) and (e) the distribution of electroporated neurons in three different regions (Up CP (upper cortical plate), Lo CP (lower cortical plate), and WM (White matter)). Data from were analyzed by (c, d) one-way ANOVA or (e) two-ways ANOVA (Bonferroni’s multiple comparisons test). Number of pups analyzed: c, d Empty vector, n = 5; WT, n = 7; p.Gln313Lys, n = 5; p.Ile678Leu, n = 14; p.Ile678Leu + WT 0.25 µg/µL, n = 7; p.Ile678Leu + WT 1 µg/µL, n = 5; p.Ile678Leu + ΔATP, n = 8; p.Ile678Leu + T96N, n = 8; p.Ile678LeuΔATP, n = 5; mKif21bΔrCC, n = 6; e Empty vector, n = 5; p.Ile678Leu, n = 6; p.Ile678Leu + WT 1 µg/µL, n = 4; p.Ile678LeuΔATP, n = 3; mKif21bΔrCC, n = 4. f Representative DIV5 cortical neurons transfected at DIV2 with pCAG2-Scarlet together with the indicated pcDNA-HA-hKIF21B constructs. g Distribution (means ± S.E.M.) of axonal collateral branches length at DIV5. Significance was calculated by two-way ANOVA (Bonferroni’s multiple comparisons test). Number of collaterals analyzed: Empty vector, n = 265; p.Ile678Leu, n = 385; from 51 (Empty vector) and 66 (p.Ile678Leu) neurons from five independent experiments. ns non-significant, *P < 0.05; **P < 0.005; ***P < 0.001; ****P < 0.0001. Scale bars, (a, b) 250 µm, (f) 150 µm. Source data are provided in the Source Data file.

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