Movie 5

The actin and microtubule cytoskeletons are coordinately regulated in a Rap-dependent manner during B cell spreading. Vector control and RapGAPII-expressing A20 cells that had been transfected with F-tractin-tdTomato (red) and GFP-α-tubulin (green) were allowed to attach to anti-Ig-coated coverslips. Time-lapse spinning disk confocal microscopy was used to acquire images every 40 s between 5 min and 15 min. 3D reconstruction of z-stacks (0.5 μm step size) with side and top views of the cells are played back at 5 frames per second (120X real time).