Fig 1: The minimal sequences of the EGF3 domain activates downstream pathways of Netrin-1. (A) Illustrations of the peptides. (B) Cortical neurons (DIV3) were stimulated with gradient concentrations of peptide E1. Cell lysates were incubated with anti-p-FAK-861, anti-p-SFK-418 and anti-p-ERK-202/204. (C) Cultured cortical neurons were treated with gradient concentrations of peptide E2. Lysates were collected and incubated with the indicated antibodies to measure the phosphorylation of FAK, SFK and ERK. (D–F) Quantification of the extent of FAK, SFK and ERK phosphorylation induced by peptide E1 in neurons is shown in (B). n = 3. (G–I) Quantification of the extent of FAK, SFK and ERK phosphorylation induced by peptide E2 is shown in (C). n = 3. (J) Cortical neurons derived from DCC wild-type and homozygote mutant embryos were stimulated with Netrin-1 or Netrin-1-derived peptides and were lysed. The resulting lysates were subjected to immunoblotting. (K) Quantification of FAK PY861 and DCC levels in DCC mutant neurons is shown in (J). n = 3. The data are presented as the means ± SEMs. One-way ANOVA was used for all statistical analyses shown in this figure (* p < 0.05; ns, not significant). Details of data analysis are seen in Supplementary Table S1.