Fig. 9. The role of calcineurin in nuclear translocation of PYK2. (A) PC12 cells were transfected with a tagged control protein (Flag-protein, FP), a phosphatase-dead form of calcineurin A (Flag-PD-CnA), or a constitutively active calcineurin A (Flag-CA-CnA) and treated for 3 minutes with a control or depolarizing (High K⁺, 40 mM) solution. Immunofluorescence was analyzed for Flag (left), PYK2 (middle) and merged with DAPI (right). Bars, 10 µm. (B) Control of expression levels of Flag-PD-CnA or Flag-CA-CnA by immunoblotting with an anti-Flag antibody. (C) Quantification of results in A. Values are means ± s.e.m. (four to six per group). Two-way ANOVA: interaction between depolarization and transfection F_(2,26)=14.6, P<0.0001, depolarization effect F_(1,26)=44.5, P<0.0001, transfection effect F_(1,26)=26.3, P<0.0001. Newman-Keuls test: FP control vs FP K⁺, ^***P<0.001, PD-CnA K⁺ vs FP K⁺, P<0.001, CA-CnA control vs FP control, °°P<0.01, CA-CnA K⁺ vs FP K⁺, P<0.01.