Keratinocyte differentiation induces loss of centrosomal γ-tubulin and Nedd1. (A–D) Cryosections of neonatal mouse backskin were stained for centrosomal proteins, as indicated. Dashed lines represent the basement membrane separating underlying dermis from the epidermis. Insets of boxed regions show centrosomes at higher magnification. Bars: (main) 10 µm; (insets) 1 µm. (E) Quantification of fluorescence intensity of centrosomal proteins during epidermal differentiation. Proliferative indicates basal cells, the second time point is the first suprabasal cell layer, the third time point is upper spinous cells, and the fourth is granular cells. (F) Cultured mouse keratinocytes were grown under proliferative conditions or were induced to differentiate and then were stained for centrosomal proteins, as indicated. Arrows indicate low levels of residual centrosomal γ-tubulin and Nedd1 in differentiated keratinocytes. Bar, 5 µm. (G) Quantification of fluorescence intensity of centrosomal proteins in proliferative and differentiated cultured keratinocytes (n ≥ 280 centrosomes from three independent experiments). (H) Western blots of whole-cell lysates prepared from proliferative keratinocytes and keratinocytes at indicated time points after differentiation induction, blotted with antibodies as indicated. n.s., not significant; *, P < 0.05; ***, P < 0.001. Data are presented as mean ± SEM.