Ephrin/Eph signaling controls neuronal polyploidy in the developing neocortex. (a) Paraffin sections of E13.5 embryos at the level of the neocortex were immunostained for CitK (red) and proteins phosphorylated on tyrosines (green). Nuclei were stained with DAPI (gray). Arrowheads indicate regions of colocalization between CitK and phosphotyrosines. Bar, 20 µm. (b) Protein lysates from E13.5 neocortex were used to immunoprecipitate tyrosine-phosphorylated proteins (Ip P-Tyr). Anti–human IgG was used as a control (Ip IgG). Whole lysate (Extract) and immunoprecipitated proteins were immunoblotted with a CitK antibody. Western blots are representative of two independent biological replicates. (c) High magnification of a dividing neural progenitor in the neocortex immunostained for CitK (red) and EphB^P-Y594 (green). DNA is stained with DAPI (gray). Bar, 20 µm. (d) Protein lysates extracted from the neocortex of E13.5 control (ctl; Efnb1^loxlox; Efnb2^loxlox) and double mutant (dcKO, Efnb1^loxlox; Efnb2^loxlox; Nestin-Cre) embryos were immunoblotted with the EphB^P-Y594 antibody. Tubulin was used as a loading control. Ratio of signal intensity is provided below. (e) Neuronal ploidy was quantified by flow cytometry in cortical neurons from control (ctl) and double conditional mutant (dcKO) P0 pups. (f) Neuronal cell death was quantified by flow cytometry in cortical neurons from control (ctl) and double-mutant (dcKO) P0 pups. Error bars correspond to SEM. Statistical p-value is indicated when significant. ns, nonsignificant.