Figure 6

Suppression of PP2A provokes shuttling and phosphorylation of GSK3β. F9 cells expressed Rfz1 were treated without (-) or with (+) OA for 1 hr and then stimulated without (time + 0) or with Wnt3a for the indicated time periods. At indicated time points, cell cultures were harvested, disrupted, and subjected to subcellular fractionation to plasma membrane (PM), cytoplasm (CY) and nuclei (NU) fractions, as described in the Methods. Samples of each fraction were subjected to protein determination, SDS-PAGE, and the resolved proteins blotted and stained with antibodies specific for GSK3β and with antibodies specific for Ser9 phospho-GSK3β. Left-handed panel shows the quantitative analysis of the blots for Wnt3a alone (blue line) or Wnt3a in the presence of OA (pink line). The data are displayed as fold of control (time zero set to 1). Right-handed panel displays immunoblots stained with GSK3β and phospho-GSK3β (Ser 9) antibodies as well as antibodies specific for subcellular fractions, (i.e., marker proteins). The results shown are derived from a single experiment, representative of two additional experiments.