Fig. 4

SiRNA-downregulation of menin in BON1 cells enhances Rapamycin-induced Akt phosphorylation and migration. Control or menin-siRNA treated BON1 cells were pretreated with or without rapamycin (100 nM) for indicated time periods upon starvation (a), PDGF-BB (b), or serum condition (c), as specified. Total cell lysate were analyzed by immunoblotting with antibodies specific for menin, Akt phosphorylated on S473, as well as S6 phosphorylation, and the expression of total protein Akt and S6. The relative protein phosphorylations were quantified for a representative experiment. The migration of BON1 cells was carried out as explained in Methods. The 12 well plate were field with medium containing 10% serum as attractant, 200 × 103 pretreated BON1 cells with either control or menin-siRNA, as indicated, were placed on the insert filters to migrate overnight at 37 °C. The amount of the migrated cells is given as percentage relative of corresponding control (d). Only in control siRNA treated BON1 cells rapamycin treatment significantly reduced cell migration (P < 0.01). Data are presented as mean values (± SEM) in three independent experiments, each performed in triplicate