Loss of PI4KIIIα causes perturbations of PtdIns4P- and PtdIns(4,5)P2-dependent processes at the plasma membrane. (A–H) Control (A and B) and PI4KIIIα KO MEFs (C–H) were transfected with STIM1-mRFP. For rescue experiments, KO MEFs were transfected with GFP-M1*-PI4KIIIα (E and F) or GFP-M1-PI4KIIIα (G and H). Plasma membrane–associated STIM1-mRFP fluorescence was imaged by TIRF microscopy before (−Tg) or 8 min after (+Tg) the addition of 1 µm thapsigargin (Tg). (I) F-actin in control and PI4KIIIα KO MEFs as revealed by staining with fluorescently labeled phalloidin. (J and K) Altered assembly of the endocytic clathrin machinery in control and PI4KIIIα KO MEFs as assessed by immunofluorescence labeling of AP-2 (J) and epsin 1 (K). Insets show a representative region at higher magnification. Bars: (A–H) 10 µm; (I–K, full size images) 20 µm; (J and K, insets) 5 µm.
