Fig 1: Role of Rbfox2 and Rbfox3 in neuronal migration during mouse brain development.(a) Characterization of pSuper-mRbfox2 and –mRbfox3 vectors. pCS-MT-mRbfox2 or –mRbfox3 was cotransfected into COS7 cells with pSuper-mRbfox2 or –mRbfox3 in various combinations. After 48 h, cells were harvested and subjected to western blotting with anti-Myc. Anti-Sept11 was used for a loading control. Relative band intensity was also shown. (b) Knockdown of Rbfox2 or Rbfox3 in migrating neurons. pCAG-EGFP was coelectroporated with pSuper-H1.shLuc (Control), pSuper-mRbfox2 or -mRbfox3 into cerebral cortices at E14.5 and fixed at P3. Coronal sections were immunostained with anti-GFP (white) and DAPI (blue). Scale bar, 100 µm. (c) Quantification of the distribution of Rbfox2- or Rbfox3-deficient neurons in distinct parts of the cerebral cortex (bin 1–5, and IZ) for each condition shown in (b). Error bars indicate SD (n = 3). (d) Effects of Rbfox1-iso1-knockdown on the expression of Rbfox2. A coronal section prepared at P7 as in Fig. 2g was stained for GFP (green) with Rbfox2 (magenta). Double-positive cells for GFP and Rbfox2 were indicated by arrowheads. Quantification of Rbfox2 expression was performed as in Fig. 2e. Note that Rbfox2 expression was comparable to surrounding internal control neurons. Bars, 10 µm. (e) Characterization of anti-Rbfox2 antibody. Lysates (20 µg of protein per lane) from COS7 cells transiently expressing Myc-Rbfox1-iso1, -Rbfox2 or –Rbfox3 were subjected to western blotting with anti-Rbfox2.