Disruption of centrobin–tubulin interaction destabilizes the centrioles. (A–I) Centrobin–tubulin interaction confers stability to the centrioles. HeLa (A, B, E, and F) and U2OS (C and D) cells pretreated with HU and transfected with either control or GFP–centrobin-TuBD were stained with anti–centrin 3 (A–D) or anti–α-tubulin and CP110 antibodies (E and F), and the percentage of cells with the indicated number of respectively stained centrioles is presented. (G and H) Increasing concentration of full-length centrobin partially rescued the inhibitory effect of centrobin-TuBD on centrobin recruitment and centriole elongation. HU-pretreated HeLa cells were transfected with centrobin-TuBD, full-length centrobin (pMini-myc-centrobin), or a combination as indicated. Cells were stained with anti-centrobin (G) and centrin 3 (H) antibodies, and the percentage of centrobin (G)- and centrin 3 (H)–positive centrioles is presented. (I) The schematic depicts the experimental method in A–H. (J) EM analysis of centrobin-TuBD cells. HeLa cells pretreated with HU and transfected with control or GFP–centrobin-TuBD were processed for EM, and 250-nm-thick sections were analyzed to detect centrosomes. (K) Overexpression of centrobin-TuBD destabilizes the preformed PLSs. U2OS-CPAP cells preinduced with doxycycline for 96 h were transfected with control or GFP–centrobin-TuBD along with dsRed–centrin 1 vector. dsRed-centrin–positive cells were evaluated for the presence of elongated or nonelongated PLSs by staining with anti-acetylated tubulin antibody. (L) The schematic illustrates the destabilization effect of centrobin-TuBD in HU-treated cells (panel a) or U2OS-CPAP–overexpressing cells (panel b). Histograms are plotted as mean ± SEM (n = 3). Asterisks denote that the difference is significant, and P < 0.001. Bars: (A and C) 1 µm; (E) 3 µm; (J) 100 nm.