Fig. 4. Ectopic localization of Bmp4⁺ cells and signaling in the kidney of the Foxd1 null. At E14.5, Bmp4^lacZ expression is normally limited to the ureter and structures in the interior of the kidney (A). A Foxd1-null mutant (B) displays weak expression in the interior of the kidney, but high expression in the capsule (ca). At E12.5 (C,D), Bmp4^lacZ expression is present in the midline of both Foxd1-null and heterozygous tissue but there is a wider distribution of expression around the kidneys in the mutant. We confirmed these results at E12.5 by immunofluorescence with an anti-ß-gal antibody (red cells in E,F) and GFP expression from the Foxd1^GFP allele (green cells in E,F). Expression of GFP and lacZ did not overlap. At E18.5, the heterozygous and mutant kidney both displayed similar expression of Bmp4^lacZ (G,H). At E12.5, phospho-Smad1 expression (brown) is in the pre-tubular aggregates (pa) (I,I'), while weaker expression is displayed in the condensed mesenchyme on both sides of the UB. There is no expression in the nascent capsule layer (nca). In the Foxd1-null (J,J'), phospho-Smad1 expression is strongly expressed in the pre-tubular aggregates, but there is also expression in the nascent capsule layer. At E14.5 in wild type (K,K'), phospho-Smad1 expression is in the pre-tubular aggregates, the renal vesicles (rv) and the UB stalk, but not in the condensed mesenchyme above the UB tips, nor the part of the UB tips that face the capsule (ca). In the Foxd1 null (L,L') phospho-Smad1 expression is also in pre-tubular aggregates and renal vesicles, but these structures are ectopically positioned between the UB and the capsule. Higher magnification of the bracketed areas in I-L are displayed (I',J',K',L'). Scale bar: 200 µm for A,B,G,H,K,L; 100 µm for C-F,I,J; 75 µm for K',L'; 50 µm for I',J'.