Figure 3.
PCIF1 suppresses ciliogenesis through BICD2. (A and B) RPE-1 cells treated with the indicated siRNAs for 48 h were subjected to the tandem mass tag quantitative proteomics analysis. Venn diagram shows the overlapping upregulated proteins in RPE-1 cells treated with PCIF1 siRNAs (A). The overlapping upregulated proteins ranked by fold change are listed (B). (C) The heatmap of the normalized ratio of ciliated cells transfected with the indicated siRNAs targeting 11 overlapping upregulated proteins from TMT proteomics analysis. The normalized ratio of ciliated cells represents the quotient of the ciliated rate induced by siRNA relative to the ciliated rate induced by siNC. (D–F) RPE-1 cells transfected with control or BICD2 siRNAs for 48 h were treated with serum starvation for another 24 h, and then subjected to western analysis or immunofluorescence. (G–I) RPE-1 cells were treated with control or BICD2 plasmid for 48 h under normal culture conditions, and then applied for Western blotting or immunofluorescence analysis. (J–L) RPE-1 cells were treated with the indicated siRNAs for 48 h under normal culture conditions, and then applied for Western blotting or immunofluorescence analysis. Western blotting analysis of BICD2 and PCIF1 protein (D, G, and J). Actin is the internal control. Immunofluorescence images of RPE-1 cells with anti-ARL13B (green) and γ-tubulin (red) antibodies (E, H, and K). DNA was stained with DAPI (blue). Cilia were indicated by white arrows. Scale bar, 10 μm. The quantification analysis of ciliation percentages is presented in SuperPlots (F, I, and L). Each color in SuperPlots signifies an independent biological replicate. Smaller symbols indicate the ciliation percentage under each field of view, while larger symbols represent the mean for each replicate. All error bars represent means ± SD. Student’s t test, *P < 0.05, **P < 0.01, ***P < 0.001. Source data are available for this figure: SourceData F3.

PCIF1 suppresses ciliogenesis through BICD2. (A and B) RPE-1 cells treated with the indicated siRNAs for 48 h were subjected to the tandem mass tag quantitative proteomics analysis. Venn diagram shows the overlapping upregulated proteins in RPE-1 cells treated with PCIF1 siRNAs (A). The overlapping upregulated proteins ranked by fold change are listed (B). (C) The heatmap of the normalized ratio of ciliated cells transfected with the indicated siRNAs targeting 11 overlapping upregulated proteins from TMT proteomics analysis. The normalized ratio of ciliated cells represents the quotient of the ciliated rate induced by siRNA relative to the ciliated rate induced by siNC. (D–F) RPE-1 cells transfected with control or BICD2 siRNAs for 48 h were treated with serum starvation for another 24 h, and then subjected to western analysis or immunofluorescence. (G–I) RPE-1 cells were treated with control or BICD2 plasmid for 48 h under normal culture conditions, and then applied for Western blotting or immunofluorescence analysis. (J–L) RPE-1 cells were treated with the indicated siRNAs for 48 h under normal culture conditions, and then applied for Western blotting or immunofluorescence analysis. Western blotting analysis of BICD2 and PCIF1 protein (D, G, and J). Actin is the internal control. Immunofluorescence images of RPE-1 cells with anti-ARL13B (green) and γ-tubulin (red) antibodies (E, H, and K). DNA was stained with DAPI (blue). Cilia were indicated by white arrows. Scale bar, 10 μm. The quantification analysis of ciliation percentages is presented in SuperPlots (F, I, and L). Each color in SuperPlots signifies an independent biological replicate. Smaller symbols indicate the ciliation percentage under each field of view, while larger symbols represent the mean for each replicate. All error bars represent means ± SD. Student’s t test, *P < 0.05, **P < 0.01, ***P < 0.001. Source data are available for this figure: SourceData F3.

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