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Fig 5. Altered F-actin rearrangement in F cells. (A) F cells expressing the mutant L1106A exhibit altered F-actin rearrangement and focal adhesion formation. F cells, expressing WT ßPDGFR or the L1106A mutant in serum-free medium, were plated on FN-coated coverslips for 90 minutes and fixed. Cells were then stained with anti-paxillin (secondary antibody: Alexa Fluor 488-conjugated goat anti-mouse IgG) to examine focal adhesion formation and with Alexa Fluor 594-conjugated phalloidin to examine the actin cytoskeleton. F cells expressing the mutant L1106A exhibit a nonpolar spreading-morphology, a dense ring of cortical F-actin and an increased number and density of focal adhesions. Scale bars, 10 µm. (B) F cells expressing WT ßPDGFR and the C-terminus of NHERF display delayed spreading on FN. Serum-deprived F cells expressing WT ßPDGFR together with a HA-tagged C-terminal fragment of NHERF (aa 270-358), which encompasses the MERM binding domain and EGFP, or F cells expressing WT ßPDGFR and EGFP alone (vector), were plated on FN-coated coverslips. After 90 minutes, the cells were fixed and stained for F-actin with Alexa Fluor 594-conjugated phalloidin. GFP expression identifies transfected cells. Scale bars, 20 µm.

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