Epistasis analysis suggests competition between EB1 and EB2 in regulation of neurite outgrowth.

<p>A: P19 cells co-transfected with neurogenic transcription factor NeuroD2 and 2pmol siRNA oligonucleotides for each target gene were stained with antibodies against neuronal β-III-tubulin (Tubb3). The total amount of siRNA oligonucleotides per well in a 384-well plate was kept constant at 4pmol (non-targeting siRNA was added, if necessary). B: Quantification of neurite outgrowth modulation analogous to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0079796#pone-0079796-g002" target="_blank">Figure 2C</a>. (*: p<0.05; ***: p<0.001; one-way ANOVA with Dunnett’s post-test using non-targeting siRNA as the control group, n = 6). C: Plot of the reduction of EB2 protein levels measured via western blot vs the increase of neurite outgrowth for different siRNAs targeting EB2. Error bars represent the standard error of the mean (n = 3). Additional western blot analyses are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0079796#pone.0079796.s003" target="_blank">Figure S3</a>. D: Confocal z-projections of partially extracted, methanol fixed neuronal differentiated P19 cells stained with antibodies against neuronal β-III-tubulin (Tubb3) and EB1. E: Quantification of average EB1 signals within neurites of P19 cells. Confocal z-projections of EB1 signals were masked based on the neuronal β-III-tubulin signal and average intensities were calculated within those masked regions (**: p<0.01; Student’s t-test, data obtained from 4 independent knock-down experiments).