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. 2016 Mar 30;15(5):678–688. doi: 10.1080/15384101.2016.1147632

Figure 3.

Figure 3.

PKN2 is a CDK10/CycM substrate and a ciliogenesis regulator (A) Quantification of 33P protein labeling in in vitro kinase assays conducted with CDK10/CycM alone, PKN2 alone or CDK10/CycM plus PKN2. CPM: counts per minute. (B) Western blot analysis of PKN2, CDK10 and CycM in anti-PKN2 or control rabbit IgG immunoprecipitates from serum-starved hTERT-RPE1 cells. Input is 1/25 of the total extract used for the immunoprecipitation. (C-F) hTERT-RPE1 cells were transfected with control (siCTL) or PKN2 (siPKN2) siRNAs, subjected to serum starvation, and analyzed 72 hour post transfection. (C) Western blot analysis of PKN2 protein levels. (D) Immunofluorescent visualization and XZ optical projections of primary cilia (acetylated-tubulin staining, shown in green) and F-actin (Rhodamine-Phalloidin staining, shown in red). Scale bar, 25 μm. (E, F) Percent ciliated cells and measurement of cilia length (quantified as described in Fig. 1 legend). (G,H) hTERT-RPE1 cells were serum starved for 24 hours, transfected with EGFP, EGFP-PKN2WT or EGFP-PKN2TATA and analyzed 16 hours post transfection. (G) Western blot analysis of PKN2 protein levels. (H) Percent ciliated cells (quantified as described in Fig. 1B) expression.