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Fig. 5. aPKC regulates the distribution of the acetylated microtubule cytoskeleton and Vangl2-VAMP1 islands. (A) Histological section of a follicle-free (collagenase-treated) stage 6 Xenopus oocyte stained for aPKC protein. The nuclear staining is non-specific, as it is also observed in aPKC-depleted oocytes (not shown). (B) High magnification images of a histological section of the animal cortex of a control stage 6 oocyte co-immunostained for VAMP1, Vangl2 (VG) and aPKC. Chevron indicates a follicle cell. (C) Western blot showing aPKC protein levels in control and aPKC antisense oligonucleotide-injected (aPKC–) oocytes. (D) Western blot for aPKC using lysates from membrane and cytosolic fractions. Total α-tubulin is used as a marker for the cytosolic fraction and C-cadherin (C-cad) is used as a membrane fraction marker. (E,E′) Sections of control (upper panels) and aPKC-depleted (lower panels) stage 6 oocytes immunostained for acetylated tubulin (E) and Vangl2 (green) and VAMP1 (red) (E′). (F) Colocalization of Vangl2 (green) and VAMP1(red) proteins in aPKC-depleted oocyte cytoplasm. Right-hand panel shows incubation with secondary antibody only. (G) Co-immunoprecipitation analyses of lysates of control stage 6 (St. VI) oocytes showing that endogenous Vangl2 protein interacts with aPKC and that this interaction is not reduced by nocodazole (Noco) treatment (5 μg/ml) but is reduced by taxol treatment (2 μM/ml). |
