Supplemental Material

This article contains the following supporting material:

• Movie 1 - Time-lapse series of images showing cellular distribution of a (left) GFP-myo1c-tail^IQ1-3, (center) GFP-K892A, and (right) GFP-903A during a TIRF/FRAP experiment in live NRK cells, as shown in Figure 3. The 1 s bleach pulse occurred at 31 s. The arrow identifies the location of the bleach pulse and the time stamp is min:sec.

• Movie 2 - Time-lapse series of images showing cellular distribution of GFP-myo1c-tail^IQ1-3 during a TIRF/FRAP experiment in live NRK cells. The size of the bleach spot was larger, and the acquisition time for each frame was longer than images acquired for analysis in Figure 3 and Movie 1. The acquisition conditions for this experiment allowed for better resolution of the membrane-attached myosin, but resulted in poor kinetic resolution.

• Movie 3 - Time lapse series of epifluorescence images showing the cellular distribution of full-length GFP-myo1c in live NRK cells, as shown in Figure 4. The red box is the region expanded on the right side of the sequence. Note the transient localization of GFP-myo1c to membrane ruffles and macropinocytic structures at the cell margins. The scale bar is 15 μm, and the time stamp is min:sec.

• Movie 4 - Time lapse series of epifluorescence images showing the cellular distribution of GFP-myo1c-K892A in live NRK cells, as shown in Figure 4. The red box is the region expanded on the right side of the sequence. Note the cytoplasmic localization of GFP-myo1c-K892A and the absence of localization to membrane ruffles. The scale bar is 15 μm, and the time stamp is min:sec.

• Movie 5 - Time lapse series of epifluorescence images showing the cellular distribution of GFP-myo1c-R903A in live NRK cells, as shown in Figure 4. The red box is the region expanded on the right side of the sequence. Note the cytoplasmic localization of GFP-myo1c-R903A and the absence of localization to membrane ruffles. The scale bar is 15 μm, and the time stamp is min:sec.

• Movie 6 - Rotation of the calculated structure of the myo1c-tail region (orange) compared with the Pdk1 structure (green).