Supplemental Materials

This article contains the following supporting material:

• Movie 01 - Spreading Phenotypes by Phase-Contrast Microscopy.
  (1) Smooth Edged, (2) Filopodial, and (3) Ruffling spreading phenotypes are shown by live-cell phase-contrast microscopy. All movies were taken over a time period of 10-20 minutes during the spreading phase.
• Movie 02
• Movie 03
• Supplemental Figure 1 - Localization of VASP mutants at Filopodial Tips.
  Localization of EGFP-tagged mutant VASP proteins in filopodia in MV^D7 (A-F). For each mutant, a series of the actin distribution (right), EGFP- mutant VASP protein (middle) and merged (right) images presented. (A) MV^D7-VASP, (B) MV^D7-Mena. (C) MV^D7-EVH2(VASP), (D) MV^D7-GAB mutant (VASP), (E) MV^D7- ΔFAB(VASP), (F) MV^D7-ΔCOCO. Scale bar 5 μm.
• Supplemental Figure 2 - Quantification of FRAP in MV^D7 derivative cell lines.
  (A-C) Examples of plots of FRAP of MV^D7 derivative cell lines. (A) MV^D7-VASP, (B) MV^D7-GAB mutant (VASP) and (C) MV^D7-ΔFAB(VASP). Whereas 100% of VASP cells (n=14 out of 14) and 90% of VASP-ΔFAB cells (n=18 out of 20) showed no recovery, 56% of MV^D7-GAB mutant (VASP) cells (n=14 out of 26) showed partial recovery. The data which showed significant recovery were curve-fitted to a single exponential function as described in Materials and Methods. An example of the curve-fitting was presented in (B). (D-E) Analysis of fluorescence-recovered samples of MV^D7-GAB mutant and MV^D7-ΔFAB(VASP) (14/26 and 2/20, respectively). Bar graphs of the half-times of recovery (D) and the final recovery (E) calculated from curve-fitting to a single exponential function. Mean ± standard deviation.