Figure 4. Loss of basal cell polarity in K14-Cre;Nf2^lox/lox embryonic skin.

(A, B) In contrast to the apical concentration of aPKC (green) at the basal:suprabasal interface in the wild-type E15 epidermis, aPKC is diffusely localized throughout basal cells of the K14-Cre;Nf2^lox/lox epidermis. DAPI (blue), and anti-β4-integrin (red) were used to label nuclei and basal lamina. Bars, 20 μm.

(C) Despite similar total levels of aPKC (right), very little aPKC immunoprecipitated with Par3 in the K14-Cre;Nf2^lox/lox neonatal epidermis relative to wild-type. Quantitation of three independent experiments using ImageJ software is shown on the bottom left. Values = mean +/− SD.

(D, E) Immunostaining with an anti-β-tubulin antibody (green) shows representative spindle orientations in wild-type and K14-Cre;Nf2^lox/lox basal cells. Co-staining with DAPI (blue) and anti-β4-integrin antibodies (red) labels nuclei and basal lamina, respectively. Bars, 10 μm.

(F) Calculation of the percentage of basal cells undergoing asymmetric division (mitotic spindles at a 90° angle relative to the basement membrane), compared to those undergoing non-asymmetric divisions (less than a 90° angle relative to the basement membrane). Values = mean +/− SD; n = >300 mitoses/genotype.

(G) Plotting of individual mitotic spindle orientations revealed that they were either parallel (symmetric) or perpendicular (asymmetric) in wild-type basal cells, as expected. In contrast, mitotic spindles were randomly oriented in K14-Cre;Nf2^lox/lox embryonic epidermal basal cells.