ERK is activated in response of H₂O₂ and contributes to H₂O₂-induced protrusion dynamics.

<p>(A) Cell lysates from starved PtK1 cells treated with 500 µM H₂O₂ alone or in combination with ROS scavenger (5 mM) for 0-15-30-45-60 min were immunoblotted with antibodies against pERK and ERK. In (B), the graph represents the averaged normalized pERK values. Data are from four and three independent experiments for H₂O₂ and H₂O₂+ROS scavenger, respectively. Error bars represent s.e.m. *, p<0.05 compared to 0 min H₂O₂ and 15 min H₂O₂+scavenger. (C) Immunolocalization of phosphorylated ERK (pERK, green) and F-actin phalloidin staining (red) in starved PtK1 cells treated with H₂O₂ 500 µM for the indicated times. The scale bar is 10 µm. (D and E) Fluorescence intensity of pERK (D) and F-actin (E) in cells treated with H₂O₂ 500 µM, measured from the cell edge (0 µm) into the cell center (10 µm). (F) pERK/F-actin fluorescence intensity ratio in cells treated with H₂O₂ 500 µM, measured from the cell edge (0 µm) into the cell center (10 µm). In (D)–(F), the data shown represent one experiment and are averaged from at least 13 cells for each condition. The experiment was repeated three times with similar results (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0041342#pone.0041342.s002" target="_blank">Figure S2D</a>). Protrusion width (G), persistence of protrusion (H) and protrusion/retraction velocities (I) in starved Ptk1 cells incubated for 45 min with control media (+DMSO) or media containing 500 µM H₂O₂ alone or in combination with UO126, a MEK inhibitor. In (G)–(I), the data shown result from the analysis of at least 23 cells and 115 kymographs per condition. Error bars represent s.e.m. ***, p<0.001 compared to control and H₂O₂+UO126.