Fig. 6. dWnt4, wg double mutants display early loss of PCP orientation.

All panels show 16h APF pupal wings (anterior is up and distal to the right), with the respective polarity distributions in rosette diagrams. (a-a””) wild-type, and (b–c””) wnt4^−/−, wg^CX3/^ts double mutant pupal wings shifted at 0–1h APF to 29°C and incubated for 15 hours (about 16h APF at 25°C). (a',b') display high magnification view of boxed areas in (a) and (b). (c) Same pupal wing area of an independent wnt4^−/−, wg^CX3/^ts double mutant. Fmi staining (monochrome) was used as PCP orientation marker and polarity and strength/nematic order were plotted as yellow lines (a”, b” and c'). Cells close to wing margin are less (or not) polarized as previously reported³⁷ (the 3 margin proximal rows highlighted by pink lines were excluded from polarity analysis in rosette diagrams in a”' and b”', for comparison; all other rosette diagrams include all cells. (d) shows the sum of three independent wnt4⁻, wg^CX3/ts mutant wings. wnt4⁻, wg^CX3/ts double mutant wings showed much wider polarity distribution than wild-type (compare a”' with b”' [p=0.002 with two-sample Kolmogorov-Smirnov test, n=number of cells]; or a”” with b”” and d; p<10⁻⁶; 3/3 wnt4^−/−, wg^CX3/ts wing discs quantified displayed very similar phenotypes; 2 examples shown in b and c panels, d shows sum of all three). Overall polarization/nematic order level was markedly reduced from wild-type to the wnt4⁻, wg^CX3/ts double mutants (wt: 59.13+/−36.65 vs mutant: 35.68+/− 20.00 p=10⁻²⁶ with t-test; average nematic order +/− s.d.). All double mutant wings analyzed show complete lack of orientation and reduced overall polarization (3 out of 3 quantified wnt4^−/−, wg^CX3/ts and all others not quantified in detail displayed visually very similar phenotypes).