Movie 2

EFA6A depletion leads to Arl13B positive Distal Appendage Vesicles. EFA6A siRNA treated RPE-1 cells were serum-starved for 24h before processed for STED 3D imaging using anti gamma tubulin (magenta) and anti Arl13B (red) or anti CEP164 (yellow) antibodies. All the samples were mounted in Abberrior Mount Solid Antifade (Abberior GmbH, Göttingen). STED images were acquired using a Leica SP8 STED 3X (Leica Microsystems, Nanterre), at 700Hz through a 93x/1.3 NA Glyc objective using the LAS X software (Leica Microsystems,Nanterre). 3D STED image were obtained with a 561 and 633nm excitations and the depletion at 775nm for both fluorophores (20-30% of its power. STED images had a 30-nm pixel size and all the images were deconvolved using Huygens Professional (version 18.10, Scientific Volume Imaging, The Netherlands, http://svi.nl), using the CMLE algorithm, with respectively SNR:100 with 5 iterations.