Figure 5.

PPARA and NR1H4 reciprocally control ciliogenesis. (A and B) RPE1-SMO-GFP or HK2 cells were treated with 1, 3, or 6 μΜ GW 4064 for 24 h, as indicated, and immunostained with anti-ARL13B Alexa Fluor 488-conjugated antibody. The cells were observed under fluorescence microscope. Data shown represent mean ± s. d. percentage of cells with primary cilia for 200 cells per well in triplicate samples. (C) HK2 cells were transfected with scrambled control or NR1H4 siRNAs. After 48 h, cells were treated with 0.5 μΜ GW 7647 as indicated, and immunostained with anti-ARL13B Alexa Fluor 568-conjugated antibody. Data shown represent mean ± s. d. percentage of cells with primary cilia for 200 cells per well in triplicate samples. (D and E) HK2 cells were transfected with control or NR1H4 siRNAs for 48 h followed by treatment with 0.5 μΜ GW 7647 for 12 or 24 h. Total RNA or protein was extracted and analyzed by quantitative real-time polymerase chain reaction (qPCR) or immunoblotted, respectively. (F) HK2 cells were pretreated with 0.5 μΜ GW 7647 for 30 min, treated with different doses of GW 4064, and ciliated cells were visualized by immunofluorescence using anti-ARL13B Alexa Fluor 568-conjugated antibody. Data shown represent mean ± s. d. percentage of cells with primary cilia for 200 cells per well in triplicate samples. All the experiments were performed 3 times. *P < 0.05, **P < 0.01 and ***P < 0.001, one-way ANOVA, compared to that of cells in normal, serum-free media or GW 7647 only. CTRL; scrambled control.