Modulation of PTP1B expression alters ephrin-induced cell contraction. (A) EphA3/HEK293T cells stably transfected with PTP1B-shRNA and nontarget control shRNA EphA3/HEK293T cells transiently expressing GFP alone (control) or w/t GFP-PTP1B were stimulated with ephrinA5 (40 min) or left untreated. Cell morphology, observed by Alexa Fluor 647–phalloidin staining and GFP-(PTP1B) expression, was analyzed by confocal microscopy. Closed and open arrowheads mark cells with high and low PTP1B levels, respectively. (B) Thresholded fluorescence area of Alexa Fluor 647–phalloidin staining was determined in 10 low-magnification images per condition of the experiment shown in A and plotted as mean ± SE (error bars; ***, P < 0.001). (C) Relative ephrinA5-induced cell contraction was estimated from the ratio of the actin-associated fluorescence footprint in micrographs taken after and before ephrinA5 treatment: whole 18-mm-diameter coverslips (equivalent to 100 low-magnification images; A) of phalloidin-stained cells were imaged by “tile scanning” on a fluorescence microscope. Mean ± SE from five separate experiments are shown (error bars; ***, P < 0.001; *, P < 0.05).