Fig. 4. TGFß signaling through Smad3 promotes neurogenesis in the spinal cord. HH stage 14-16 embryos electroporated with pCIG-based vectors driving the expression of nuclear GFP as reported, were harvested 48 hours later. (A-D) Control embryos electroporated with the empty pCIG vector. (E-H) Electroporation of Smad3 causes mediolateral displacement of GFP-expressing cells by >55% (E), decreases BrdU incorporation by >80% (F), cell autonomously increases p27^kip1 expression by >30% (G) and cell-autonomously induces Tuj1 by >65% (H) compared with the empty pCIG vector transfection. (I-L) Electroporation of TßR-I causes similar phenotype changes to Smad3 electroporation. (M-P) Electroporation of Smad3-3S/A reverts all aspects of the Smad3 phenotype; lateral GFP+ cells (M) are reduced by 40%, transfected cells incorporating BrdU (N) are increased four- to five-fold and transfected cells co-expressing p27^kip1 (O) or Tuj1 (P) are reduced by more than 27% and 53% respectively. (Q-T) Percentages of transfected cells at lateral positions (Q), double labeled GFP/BrdU (R), co-expressing GFP/p27^kip1 (S) and co-expressing GFP/Tuj1 (T). The medial (m) or lateral (l) position of transfected cells was defined by cell location in relation to p27^kip1+ cells. Histograms show data points as mean values ± s.d. (n=6 embryos, >6 sections were assessed in each group of experiments) ^*P<0.05; ^**P<0.01; ^***P<0.001.