FIG 7.

Cellular phosphatases are required for MCPyV ST-mediated microtubule destabilization and cell motility. (A) MCC13 cells were transfected with either EGFP, EGFP-ST, EGFP-R7A, or EGFP-Δ95-111 expression vector. After 24 h, cells were fixed and permeabilized, and GFP fluorescence was analyzed by direct visualization, whereas endogenous β-tubulin was identified by indirect immunofluorescence using a β-tubulin-specific antibody. (B) MCC13 cell lysates transfected with either EGFP, EGFP-ST, EGFP-R7A, or EGFP-Δ95-111 expression vector were analyzed by immunoblotting using stathmin-, phosphorylated stathmin-, acetylated tubulin-, and EGFP-specific antibodies. GAPDH was used as a measure of equal loading. (C) (i) MCC13 cells were transfected with either EGFP, EGFP-ST, EGFP-R7A, or EGFP-Δ95-111 expression vector. After 24 h, a scratch was created by scraping the monolayer, migration of cells toward the scratch was observed over a 24-h period, and images were taken at 0 and 24 h under a Zeiss light microscope at ×4 magnification. (ii) The size of the wound was measured at 0 and 24 h. Scratch assays were performed in triplicate.