Visualizing Caveolae Dynamics and Dynamin Regulation

Fig. S1 shows mean squared displacement of stable caveolae and the uptake of Tf-647 in Cav1-mCh HeLa FlpIn cells treated with ctrl or Dyn2 siRNA as well as caveolae dynamic parameters following Dyn1 depletion. Fig. S2 provides additional data on the colocalization of Dyn2-GFP and Cav1-mCh imaged by SIM. Fig. S3 shows Dyn2-GFP localization after EHD2 or Pac2 depletion and characterization of GFP-EHD2-Cav1-mCh and Pac2-GFP-Cav1-mCh HeLa FlpIn cells as well as representative FLIM-FRET images of GFP-fusion proteins and Cav1-mCh in FlpIn HeLa cells. Fig. S4 shows the protein titration of the HeLa FlpIn cell lines expressing K44A Dyn2-GFP and Cav1-mCh or I684K Dyn2-GFP and Cav1-mCh after Dox induction. Fig. S5 provides additional information on the effect that Dyngo 4a has on the Dyn2-GFP-Cav1-mCh HeLa FlpIn cells. Video 1 shows single particle tracking of Cav1-mCh in a Cav1-mCh HeLa FlpIn cell treated with ctrl siRNA imaged on TIRF. Video 2 shows single particle tracking of Cav1-mCh in a Cav1-mCh HeLa FlpIn cell treated with EHD2 siRNA imaged on TIRF. Video 3 shows single particle tracking of Cav1-mCh in a Cav1-mCh HeLa FlpIn cell treated with Dyn2 siRNA imaged on TIRF. Video 4 shows live cell imaging of a Dyn2-GFP-Cav1-mCh HeLa FlpIn cell using SIM (SIM² algorithm). Video 5 shows live-cell TIRF imaging of Dyn2-GFP-Cav1-mCh HeLa FlpIn cells treated 30 min with 30 μΜ οf Dyngo 4a.