Fig. 7

CCP5 and CCP6 synergistically inhibit cilia formation, but not the length of cilia. A Co-depletion of CCP5 and CCP6 in cycling hTERT-RPE1 cells synergized the abnormal cilia formation. hTERT-RPE1 cells transfected with control (siNC), CCP5, or CCP6 siRNA or both in the presence of serum were immunostained with ARL13B (red) to detect cilia and nuclei were visualized with DAPI (blue). B Quantification of the ciliation in siRNA-treated cells exemplified in A (3 independent experiments; at least 65 cells analyzed per experimental condition, Additional file 2). C Depletion of CCP6 or CCP5 partially rescued the inhibited cilia formation caused by respective CCP5 and CCP6 overexpression in quiescent hTERT-RPE1 cells. Cells transfected with GFP, GFP-tagged CCP5 or CCP6 and indicated siRNAs were serum-starved for 24 h and then immunostained with GFP (green), ARL13B (red), and nuclei were visualized with DAPI (blue). D Quantification of the ciliation in GFP-positive cells exemplified in C (3 independent experiments; at least 30 cells analyzed per experimental condition, Additional file 2). E Ciliary axoneme of quiescent hTERT-RPE1 cells transfected with control (siNC), CCP5 (siCCP5), or CCP6 (siCCP6) siRNA or both siRNAs were visualized by immunoreactivity for acetylated-tubulin (Ac-Tub, green) or ARL13B (red). Depletion of either CCP5 or CCP6 increased the ciliary length compared to the control, but co-depletion of CCP5 and CCP6 did not lead to further increase in cilia length. F Quantification of the ciliated cells recognized by ARL13B immunoreactivity in conditions shown in E (3 independent experiments; at least 45 cells analyzed per experimental condition, Additional file 2). G Quantitative analysis of the cilia length detected by ARL13B immunoreactivity as exemplified in E (Additional file 2). There is no significant difference between the CCP5 and CCP6 co-depleted group with that depleted either alone. Each dot represents one cell. Scale bars: 20 µm (A); 10 µm (C); 2 µm (E); 5 µm (H). Error bars represent s.d.. ∗ , P < 0.05; ∗  ∗ , P < 0.01; ns, no significant difference, Student’s t test