NPM/ALK expression induces changes in proteins associated with cytoskeleton, and affects cell adhesion and invasion. (A) HEK293T cells transfected with vector, wild-type (Wt), and kinase-defective mutant (K210R) NPM/ALK were evaluated by confocal immunofluorescence microscopy for the expression of the cytoskeletal proteins RhoA and paxillin. ALK (green) is expressed in Wt NPM/ALK-transfected HEK293T cells but not in vector-transfected HEK293T cells. The level of ALK expression is slightly decreased in the K210R-transfected cells. The Texas Red–phalloidin (red) staining of the vector-, Wt-, and K210R mutant–transfected cells does not show significant differences in actin expression. The expression of RhoA, paxillin, and phospho-Tyr proteins is significantly reduced in the K210R compared with Wt. (B) HEK293T cells transfected with vector, Wt, or the K210R NPM/ALK were assayed for adhesion to fibronectin. Data represent the mean of 2 experiments with error bars representing SD of triplicate mean. The unpaired Student t test was used (Wt vs K210R, P = .017). (C) The effect of NPM/ALK on invasive properties was evaluated using NIH3T3 cells transfected with vector or Wt NPM/ALK plated onto BioCoat Matrigel invasion chambers, and cells present at the opposite side of the coated membranes were stained with crystal violet. Cells per high-power field (400× magnification) were counted. The graphs represent the results of replicate independent assays.