Fig. 3: FOXJ3 is essential for proper cell fate determination and cortical lamination in deeper-layer and Layer 4 neurons.

A Electroporation of shCtrl or shFoxj3, together with the GFP (green) construct, was performed at E12.5; brains were harvested at P7. Immunostaining of P7 cortices for cortical neuron markers TBR1 (magenta), CTIP2 (red), and SATB2 (red). Enlarged views show colocalization of GFP+ cells with these markers. Brain slices were counterstained with DAPI (blue) to indicate nuclei. Scale bars: 200 μm (top panels), 50 μm (lower panels). B Bar graphs show the percentage of GFP+ cells colocalized with TBR1, CTIP2, or SATB2. Error bars represent S.E.M. **: p < 0.01, ***: p < 0.001 (n = 3 independent experiments; two-tailed unpaired t-test; p = 0.0012, 0.0008, 0.0043). C Axon projection patterns of GFP+ cells (green) electroporated with shCtrl or shFoxj3. DAPI (blue) was used to counterstain cell nuclei. Scale bars: 400 μm (top panels), 100 μm (lower panels). D Electroporation of shCtrl or shFoxj3 with GFP at E13.75 with fixation at P7. Immunostaining for cortical layer markers RORβ (red) and BRN2 (red) in P7 cortices. Enlarged panels show colocalization of GFP+ cells (green) with layer markers. Brain slices were counterstained with DAPI (blue) for nuclei. Scale bars: 200 μm (top panels), 50 μm (lower panels). E Quantification of GFP+ cells colocalized with RORβ, and BRN2. Error bars represent S.E.M. ***: p < 0.001, ****: p < 0.0001 (n = 3 independent experiments; two-tailed unpaired t-test; p < 0.0001, p = 0.0006) F Callosal projection patterns of GFP+ cells (green) after electroporation. Scale bars: 750 μm (top panels), 100 μm (lower panels). G Quantified intensity of callosal projections of GFP+ cells. Error bars represent S.E.M. *: p < 0.05 (n = 3 independent experiments; two-tailed unpaired t-test; p = 0.0184).