PTEN and Crm1 form an S6K mediated complex upon S1P stimulation.

<p>A) PPC1 cells were transfected with WT-PTEN and FLAG-Crm1. Cells were collected after 2 hours stimulation with 500nM S1P or PBS and submitted to immunoprecipitation of the FLAG-Crm1 protein. The negative control (Neg) indicates lysate from cells not transfected with FLAG-Crm1. B) PPC1 cells were transfected with FLAG-PTEN and collected after 2 hour stimulation with 500nM S1P or PBS and submitted to immunoprecipitation of the FLAG-PTEN protein. The negative control (Neg) indicates lysate from cells not transfected with FLAG-PTEN. C) PPC1 cells were infected with Ad-AC or Ad-GFP and analyzed for S6K phosphorylation. D) PPC1 cells were treated with 500nM S1P for 2 hours and analyzed for S6K phosphorylation. E) PPC1 cells transfected with WT-PTEN were treated with water (NT) or 2.5 µM S6K1 inhibitor DG2 (S6Ki) for 24 hours then stimulated with 500 nM S1P for 2 hours prior to immunostaining for PTEN (red) and nuclei (blue). F) The percentage of cells from (E) which had nuclear PTEN in each treatment. G) PPC1 cells were transfected with WT-PTEN and FLAG-Crm1 and treated for 24 hours with 2.5 µM S6Ki. Cells were collected after 2 hour stimulation with 500nM S1P or PBS and submitted to immunoprecipitation of the FLAG-Crm1 protein. One way ANOVA with Bonferroni correction, *p<.05**p<.01.