Single-particle motility of a SHE complex consisting of ASH1-3′+ GFP-She2p (ΔhE) + She3p + Myo4p. In this montage, SHE complexes containing the She3p-binding mutant GFP-She2p (ΔhE) (right) are compared with complexes containing wild-type GFP-She2p and no RNA (left). To ensure comparability between both experiments, image sequences start at the exact same time when each complex has been applied to the flow cell (i.e., 3 min). GFP is shown in green, and TMR-labeled actin filaments are in red. TIRFM images were taken with a TIRF microscope (IX50; Olympus) at room temperature at 4 frames/s and are displayed at 16 frames/s. As a result of size limitation, only every third frame is displayed. Bar, 5 µm. The time stamp indicates the total elapsed time since the beginning of the acquisition. The video was collected during the first 8 min of data acquisition. A full-size version of the video can be accessed via the JCB DataViewer. Video is related to Fig. 4 F. Time stamp indicates minutes and seconds.
