Figure 5.

Recruitment of FilGAP to bead complexes. (A) HEK cells were grown on coverslips, transfected with FilGAP, and subjected to mechanical force as described in text. FilGAP was visualized by staining with anti-HA antibody. Accumulation of FilGAP in the bead complexes is indicated by arrows. (B) HEK cells, grown on 10-cm cell culture dishes, were transfected with FilGAP or FilGAP^ΔCC and either subjected to mechanical force as described in text or left untreated. Proteins associated with beads were obtained by bead isolation technique as described in Materials and Methods and analyzed by immunoblotting. (C) Quantification of bead-associated FilGAP and FLNa. Quantities of bead-associated FilGAP are represented as ratios relative to the levels of FilGAP in crude cell lysates. Bead-associated FLNa is presented by peak intensity. (D) FLNa-KD HEK cells were transfected with HA-tagged FilGAP and either full-length FLNa or truncated FLNa^Δ23 construct. Cells were subjected to force and bead-associated proteins were obtained as described previously. (E) Quantification of bead-associated FilGAP and FLNa was done as described in C. Intensities of all bands were measured using ImageJ densitometry software. Data are expressed as the means and SDs of three independent experiments. *p < 0.03 versus respective untreated control.