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. Author manuscript; available in PMC: 2014 Apr 15.
Published in final edited form as: Dev Cell. 2008 Aug;15(2):187–197. doi: 10.1016/j.devcel.2008.07.004

Figure 2.

Figure 2

RNAi-mediated suppression of CEP290 inhibits primary cilia formation. (A) Western blotting of CP110 and CEP290 in RPE-1 cells treated with control, CP110, CEP290, or both siRNAs. α-tubulin was used as a loading control. (B) U2OS cells were arrested at S phase with hydroxyurea for 48 hr after siRNA transfection. The percentages of cells with more than two γ-tubulin dots, indicative of centrosome over-duplication, were determined. (C) The percentages of U2OS cells with well-separated γ-tubulin dots, indicative of premature centrosome separation, were determined. (D) The percentages of binucleate WI38 cells, indicative of cytokinesis failure, were determined. (E) The percentages of growing RPE-1 cells with primary cilia were determined using acetylated tubulin as a marker. (F) RPE-1 cells transiently transfected with control or CEP290 siRNAs, induced to quiescence, stained with antibodies to glutamylated tubulin (GT335, green), CEP290 (red), and with DAPI (blue). Bar: 10 μM; insets: 2 μM. (G) The percentages of quiescent RPE-1 cells with primary cilia were determined using either acetylated tubulin (Ac. tub.) or gltuamylated tubulin (GT335) as a marker. In (B), (C), (D), (E) or (G), average data obtained from three independent experiments were shown. Error bars represent +/- S.D. About 200 cells for each siRNA condition were scored each time.