Figure 2
Overexpression of PELO affects actin stress fibers, cell growth and spreading in stable PELO-overexpressing cell lines. A, Hep2G cells were transfected with Myc-PELO fusion constructs or empty vector. Protein extracts from stable control (LC) and Myc-PELO expressing cell lines (L1, L3, L6 and L7) were analyzed by immunoblotting with monoclonal anti-Myc tag antibody. The membrane was subsequently reprobed with α-tubulin antibody. B-C, Cells of Myc-PELO-overexpressing L3 (B) and L6 (C) cell lines were stained with FITC-phalloidin. Cell showed diffusely cytoplasmic actin are labelled with arrows and that containing stress actin with arrowheads. D, In vitro growth properties of control and PELO-overexpressing cells. Growth curves of mock-transfected Hep2G (LC), and PELO-transfected clone L3 and L6. E, Growth of the same cell lines in soft agar. 0.2 × 104 cells were plated per dish. 100% (LC) = 4.4 × 102 colonies. The data points represent the means ± SD of three independent experiments. F, Control cells (LC) as well as L3 and L6 that expressed PELO at low and high levels, respectively, were seeded onto fibronectin-coated slides for 30 and 120 min. Cells were then counted and scored for spreading. White bars represent L3, black bars L6 and grey bars LC cells. Results are presented as percentage of cells that adapted spread morphology at 30 and 120 min after seeding. 100 cells per well were counted in three separate experiments. Data represent means ± SD.