Figure 2.

Rab7 is required for serum-induced disassembly of primary cilia. (A) Schematic diagram summarizing the strategy used for the cilia disassembly experiments. (B) Quiescent RPE-1 cells transfected with control or Rab7 siRNA were subjected to serum restimulation for the indicated times. Primary cilia were scored at 0, 2, and 24 h after serum readdition. Cilia were stained with anti-Arl13b (green), and nuclei were stained with Hoechst (blue). Scale bar, 10 µm. (C) RPE-1 cells transfected with the indicated siRNAs were subjected to cilia disassembly assay. The percentage of cells with primary cilia was quantified. Data are means ± SD of three independent experiments. Student’s t test was performed. **P < 0.01, ***P < 0.001. From left to right, cell n = 426, 402, 395; 489, 450, 389; 407, 372, 390; and 407, 382, 324. (D) Immunoblots of RPE-1 cell lysates in B with the indicated antibodies. GAPDH was used as a loading control. (E) Western analysis of Rab7 in RPE-1 cells after serum starvation and serum restimulation for the indicated times. AurA, HDAC6, and GAPDH were used as positive, negative, and loading controls, respectively. Quantification of relative amounts of Rab7, AurA, and HDAC6, normalized to respective GAPDH levels, is shown at the bottom of each immunoblot.