(a–c) Correlation of Wnt4-GFP expression to stages of early nephron formation defined by Yang et al (Yang et al., 2013). Staining with NCAM (white, cell membrane), Wnt4-GFP (green), and aPKC (red) shows no polarization in Wnt4-expressing PTA structures in a, one aPKC-enriched foci in the primitive RV shown in b, and a polarised luminal epithelium within a mature RV in c. (d) Schematic of the experimental approach. In Wnt4GCE;Rosa26R-LSL-tdTomato mice, committing cells within the PTA near the tip-stalk junction express GFP and become permanently labelled by tdTomato expression in the presence of tamoxifen. PTA structures mature to primitive RV and RV structures as nephrogenesis proceeds. (e) Cells were labelled with a single dose of tamoxifen at E12.5, then fixed and stained at E13.5, E14.5 or E15.5. Embryonic kidneys were stained for tdTomato protein (red), GFP (green) and nephron progenitor marker SIX2 (white) and whole-mount imaged. (f) Maximum intensity projection of a typical E15.5 dataset is shown, with a single niche marked by a dotted region. This image has been displayed on a black background for presentation purposes. (g–k) At 24 hr after tamoxifen (E13.5) tdTomato-labelled cells are restricted to the PTA (magenta arrowhead), marked by GFP expression and are absent from the cap mesenchyme marked by high SIX2 expression (dashed outline). (l–p) By 48 hr after tamoxifen (E14.5) rare tdTomato-labelled Wnt4 lineage cells (arrows) are found in the SIX2+ cap population (dashed outline). (q-u) At 72 hr after tamoxifen (E15.5) clusters of SIX2+ tdTomato-labelled cells (arrows) are found in some caps (dashed outline). Maximum intensity projections and 3D projections of confocal volumes show a cluster of cells present in a cap at E14.5 and E15.5 (arrows). Scale for all panels as in g, l, q, except for 3D projection where scale is indicated in bounding box.