Fig. 4. Mash1 cells primarily become dI3 and dI5 neurons. Immunofluorescence on transverse sections of Mash1^+/– (A,C,E) or Mash1^–/– (B,D,F) mouse embryos at E11.5 containing the transgene M1-GIC (see Fig. S1 in the supplementary material) and the Cre reporter allele R26R-YFP. (A-F) Anti-GFP antibodies detect both GFP from M1-GIC and YFP when R26R-YFP reports the Cre activity from M1-GIC. These green labeled cells reflect cells with Mash1 or progeny of Mash1+ cells. (A,B) Brn3a;Lhx1/5 (pink) label dI2. These cells would appear white if they co-labeled with antibodies to GFP/YFP. The absence of white cells indicate dI2 do not derive from Mash1+ precursors (A). (C,D) Brn3a;Isl1 (pink) label dI3. White cells indicate co-labeling with antibodies to GFP/YFP and demonstrate Mash1+ cells give rise to dI3 neurons (C, arrowheads) and these cells are absent in the null (D). (E,F) Lmx1b (red) labels dI5. The co-labeling with GFP/YFP (yellow) indicates dI5 are derived from Mash1+ cells (E, arrowheads) and these cells are absent in the null (F). Lhx1/5 (blue) labels dI4. Co-labeling with GFP/YFP (turquoise) indicates rare dI4 cells derived from Mash1+ cells (E, arrow), and these cells do not increase in Mash1^–/– (F). The increase in dI2 and dI4 seen at E10.5 cannot be attributed to fate switching of dI3 and dI5 precursor cells. The color of each neuronal population is shown on the left without and with (black background) co-labeling with the GFP/YFP. The total number of cells for each neuronal subtype and the number of each that co-labels with GFP/YFP are shown in the table. (G) Individual populations in each mutant were counted on at least three sections from at least three embryos each. Scale bar: 25 µm.