Supplemental Material

This article contains the following supporting material:

• Video1 - Confocal time-lapse series of a B16/F1 cell expressing EGFP-Myo9b^GAP-. Note the dynamic localization of Myo9b in the extending lamellipodium and in circular membrane ruffles. Scale bar 10 μm.
• Video2 - Confocal time-lapse series of a B16/F1 cell expressing EGFP-Myo 9b-^ins-/GAP--. The video demonstrates the accumulation of EGFP-Myo 9b-^ins-/GAP-- in the tips of protruding and retracting filopodia. Scale bar, 10 μm.
• Supplemental Figure 1 - Expression of endogenous Myo9b and various exogenous Myo9b constructs in B16/F1 cells. Cells were transfected with the indicated constructs of Myo9b and cell homogenates were analysed by immunoblotting using antibodies directed against GFP (A). In B) identical samples were analysed with antibodies directed against Myo9b and GFP as indicated on the right. Constructs are indicated above each lane. The star in A) indicates a cross-reactive band of the employed GFP antibody. Myo9b was detected with the monoclonal antibody FP3/F8 and GFP either with a monoclonal antibody from mouse (A) or a polyclonal antibody from rabbit (B).
• Supplemental Figure 2 - Fluorescence recovery kinetics of EGFP-Myo9bGAP- and mRFP-actin after treatment of cells with jasplakinolide. B16/F1 cells were co-transfected with EGFP-Myo9b^GAP- and mRFP-actin and treated with 1μM jasplakinolide 3- 15 min. before the FRAP experiment. The graph shows the result of a typical FRAP experiment. Jasplakinolide inhibits F-actin turnover and accelerates the recovery kinetics of EGFP-Myo9b^GAP-.
• Supplemental Materials