Supplemental Figure 1CD
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Fig. S1. (A,C) Subcellular localisation of MARK1 and MARK4 after PKCζ co-expression. Wild-type (wt) or indicated mutant forms of GST-MARK isoforms were expressed in HEK 293 cells, in the absence or presence of kinase inactive (D394/A, ki) or wild-type (wt) Flag-PKCζ, which were fractionated into a cytosolic and a membrane fraction. Subcellular fractions, including solubulised portions of the homogenates were analysed by western blot with regards to MARK isoforms (αGST), PKCζ (αFlag), the cytosolic marker GAPDH (αGAPDH) and the plasma membrane marker Na-K-ATPase (αNa-K-ATPase). (B,D). As above, except that GFP fusions of MARK isoforms were employed and cells were fixed in 3% paraformaldehyde 24 hours post transfection. The cells were subsequently permeabilized and stained with anti-Flag and a 594 nm (red)-labelled anti-mouse antibody. The fluorescence was analysed by confocal fluorescence microscopy. The cells shown are representative of images obtained in two separate experiments. Bars; 10 μm.
