FIGURE 7.

Autism‐associated mutant tomosyn fails to restore dendritic arborization and spine loss in tomosyn knockdown neurons. (a) An illustration of the location of L412V and Y502C mutations in tomosyn. (b) Representative images of reconstructed dendritic trees and (c) Sholl analysis, (d) branch number, and (e) total dendrite length from mouse hippocampal neurons transfected with scrambled shRNA + GFP, shTomosyn + GFP, shTomosyn + WT^r‐Tom‐GFP, shTomosyn + L412V^r‐Tom‐GFP, or shTomosyn + Y502C^r‐Tom‐GFP. Scale bar, 100 µm. Both L412V and Y502C mutant tomosyn failed to rescue total dendrite length compared to scrambled shRNA. Only Y502C failed to rescue branch number compared to scramble. ^# p < 0.05, ^## p < 0.01, ^### p < 0.001, ^#### p < 0.0001 when compared to scramble; *p < 0.05, **p < 0.01, ****p < 0.0001 when compared to shTomosyn, one‐way ANOVA with Tukey multiple comparisons test. n = 30 scramble, 31 shTomosyn, 27 WT rescue, 33 L412V rescue, and 31 Y502C rescue. (f) Representative confocal images and (g) the quantification of spine density in cultured hippocampal neurons transfected with different tomosyn constructs as described in b, c, d, and e. Scale bar, 2 µm. ^#### p < 0.0001 when compared to scramble; ****p < 0.0001 when compared to shTomosyn, one‐way ANOVA with Dunnett's multiple comparison tests. n = 31 scramble, 30 shTomosyn, 29 WT rescue, 31 L412V rescue, and 31 Y502C rescue. (h) A reduction of GluR1 surface expression in dendritic segments in neurons co‐expressing shTomosyn‐L412V^r‐Tom‐RFP, compared to shTomosyn‐WT^r‐Tom‐RFP, was observed. **p < 0.01 when compared to shTomosyn; ^## p < 0.01, ^### p < 0.001 when compared to shTomosyn‐L412V^r‐Tom‐RFP by one‐way ANOVA with Tukey's multiple comparisons test. n = 28 scramble, 21 shTomosyn, 22 WT rescue, 24 L412V rescue, 20 Y502C rescue