Figure 4.

Integrin β1 is dispensable for the establishment but necessary for the maintenance of apicobasal polarity

(A) Distribution of the polarity marker PAR3 in mESCs at 1- and 2-cell stage (24 h of culture in −2iLIF). Wild-type cells (fl/fl, top) recruit PAR3 toward the AMIS similarly to mutant cells (Δ/Δ, bottom).

(B) Quantification of the orientation of apicobasal polarity at 24 h: Fisher’s exact test: p = ns (number of mESC spheroids n = 42 [fl/fl], n = 39 [Δ/Δ]).

(C and D) Distribution of centrosomes, as shown by γ-tubulin staining, at 2-cell stage and quantification of the angles along the nuclear-centrosome axis. Centrosome-nuclear axis angle: means ± SEMs (red). Test: Mann-Whitney test: p = ns (number of mESC spheroids n = 26 [fl/fl], n = 31 [Δ/Δ]).

(E and F) Assessment of polarization in agarose at 24 h by Golgi and PAR3 localization. Fisher’s exact test: p = ns (number of spheroids n = 38 [fl/fl], 38 [Δ/Δ]). Both wild-type and mutant cells display correct apicobasal polarity at 24 h of culture, even in the absence of ECM components.

(G) Assessment of polarization at 48 h. PAR6 is recruited at the apical site where actomyosin accumulated in wild type. PAR6 is recruited basally at the site of actomyosin accumulation in mutant cells.

(H) In wild type, podocalyxin vesicles are recruited apically at PAR6 site. In mutant, PAR6 localizes basally, and podocalyxin vesicles are secreted basally at this latter site.

(I) Quantification of the orientation of apicobasal polarity at 48 h, assessed by Golgi and PAR6 localization. Fisher’s exact test: ^∗∗∗∗p < 0.0001 (n = 30 [fl/fl], n = 29 [Δ/Δ]). Scale bars: 5 μm (A, C, and E) and 10 μm (G and H).