Fig. 2. Overexpression of c-Gcm1 leads to depletion of proliferative neural progenitors. (A-F,J-P) Transverse sections after electroporation with control or c-Gcm1 expression vectors at the level of the truncal neural tube of E1.5 chicken embryos. For all images, the electroporated side is on the left; electroporated cells were detected by GFP immunolabeling or by GFP fluorescence (green in both cases). (A-B') GFP immunolabeling (A,B) and in situ hybridization using Sox2 (A') and NeuroM (B') probes. Note the decrease in Sox2 (A') and the increase in NeuroM (B') expression 6 hours after c-Gcm1 overexpression. (C-D') Ngn2 expression domain is not modified upon c-Gcm1 overexpression (C,C'), whereas Ngn2 overexpression induces c-Gcm1 expression (D,D') 24 hours after electroporation. Note that signal development was stopped before detection of endogenous c-Gcm1 mRNA in D'. (E,F) BrdU (red) and GFP colabeling in embryos subjected to a 1 hour BrdU pulse performed 30 hours after electroporation of control (E) or c-Gcm1 (F) expression vectors. Many of the cells electroporated with the control expression vector are in a proliferative state (arrows in E), whereas overexpression of c-Gcm1 markedly reduces the number of transfected BrdU-positive cells (F). Pax6 (J-M) and Pax7 (N-P) immunolabeling performed 24 hours after electroporation of control (J,J',L,O) or c-Gcm1 (K,K',M,N,N',P) expression vectors. c-Gcm1 overexpression strongly reduces the number of Pax6 (K,K',M) and Pax7 (N,N',P) expressing progenitors in the intermediate and dorsal neural tube, respectively. High magnifications of neural tube sections show GFP (L,M,O,P) and Pax6 (red in L,M) or Pax7 (red in O,P) colabeling. Neural progenitors electroporated with control vector maintain the expression of Pax6 and Pax7 (arrows in L,O), whereas neural progenitors electroporated with c-Gcm1 do not (arrows in M,P). (G-I) Percentage of transfected cells expressing Sox3 (G), incorporating BrdU (H) or expressing Pax6 (I), after electroporation of control or c-Gcm1 expression vectors. Asterisks indicate significant differences (P<0.05 in G, P<0.0001 in H; P<0.01 in I). Scale bars: 40 µm in A-F,J-K',N-O; 10 µm in L,M,O,P.