Video 2. Super-resolution 3D-SIM FRAP experiments reveal the differences between how DSpd-2 and Cnn are incorporated into the PCM.
(Related to Figure 4). All Videos shown here are maximum intensity projections of image stacks. Live cell SD-SIM illustrating the dynamic behaviour of DSpd-2-GFP (A) and GFP-Cnn (B) at centrosomes in Drosophila embryos. The employed OMX Blaze 3D-SIM system enables sub-diffraction live cell imaging at high frame rates with ∼two-fold better xy- and z-resolution compared to conventional microscopy. Time before and after photobleaching (t = 0 s) is shown at the top right of each panel. Note how, prior to photobleaching, GFP-Cnn has a broader distribution that DSpd-2-GFP within the PCM. Immediately after photobleaching, DSpd-2-GFP fluorescence recovers in the shape of a toroid around the centriole, supporting our conclusion that Asl, which is distributed as a toroid around the centriole, is the major recruiter of DSpd-2 to centrosomes. In contrast, GFP-Cnn fluorescence recovers in a broader region around the centrioles, supporting our conclusion that DSpd-2 is the major recruiter of Cnn to centrosomes.