frizzled9b limits basal cell proliferation and directs basal protrusions to generate orthogonal tubule interconnections. (A,B) Basal new nephron invading cells are non-proliferating in wild type (A, asterisk) but show ectopic proliferation in fzd9b mutants (B, asterisk). (C,D) Basal protrusions (arrowheads) are directed orthogonally at the target distal tubule (DT) in wild type (C, arrowhead) but show a lateral displacement in fzd9b mutants (D, arrowhead). (E) Quantification of the number of EdU+ basal cells (in the bottom 6 µm) per nephron reveals a significant increase in fzd9b mutants that can be blocked by IWR1 treatment. Data are mean±s.d. (wt: 0.55±0.93, n=11; fzd9b−/−: 5.75±3.2, n=16; fzd9b−/− + IWR1: 0.38±0.52, n=8). (F) Quantification of lateral displacement of basal protrusions in fzd9b mutants. Data are mean±s.d. (wt: 0.44±0.25, n=11; fzd9b−/−: 1.06±0.67, n=11). Blue fluorescence indicates Hoechst-stained nuclei. Arrows indicate direction of fusion with the distal tubule (DT). Scale bars: 5 µm. E: ****P<0.0001; F: **P=0.0095 (ordinary one-way ANOVA).
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