A Polybasic Motif Allows N-WASP to Act as a Sensor of PIP2 Density
Figure S1. PIP2 binding assays
Supplemental Figure S2. Actin polymerization assays
Supplemental Figure S3. PIP2 activation curve for deltaEVH1-N-WASP
Supplemental Figure S4. Activation of N-WASP by PIP3
Supplemental Figure S5. Co-activation by Cdc42
Movies were made from timelapse images taken every 5 s over a time period of 5–10 min. Live cells were visualized by fluorescence microscopy under 600× magnification 24 hr after transfection with the appropriate constructs (N-WASP variant and actin-GFP). Supplemental Movie 1 depicts a cell transfected with 14K N-WASP and actin-GFP. Actin comet tails can be seen rocketing around the cytoplasm. For comparison, Supplemental Movie 2 depicts an example of a cell transfected with 9K N-WASP and actin-GFP. No actin rocketing is observed. Only slight fluctuations of vacuoles are observed. Played with Windows Media Player, these movies run 15 s (14 K) and 8 s (9 K) each. The movies have been accelerated by a factor of approximately 35-fold above real time. See Experimental Procedures for further details.