Golgi complexes are in close position to the pre-DMS and contribute to DMS growth. MKs were cultured for 4 hours or for 1 or 3 days. The cells were permeabilized and immunostained for the Golgi marker giantin (red) and GPIbβ (green). (A-C) Representative confocal images are shown at different developmental stages of the DMS. (A) Giantin staining is confined to the perinuclear region and lies close to the pre-DMS (inset of boxed area). (B) Close association but no colocalization of giantin with the intermediate DMS. Note that the size of the giantin spots increases at these later stages (white arrowheads). (C) Giantin spots become randomly distributed in MKs at the latest stages of maturation (white arrowheads). Bars, 10 µm. (D-E) Confocal images of MKs cultured for 20 hours and treated as indicated. Note the absence of Golgi staining after BFA treatment. Dotted lines indicate cell perimeters. Bars, 10 µm. (F) Quantification of the number of MKs displaying typical pre-DMS areas. (G-H) Electron micrographs showing the presence of Golgi complexes (arrowheads) in proximity to the pre-DMS between (G) 2 nuclear lobes or (H) the intermediate DMS. Bars, 500 nm. (I) A typical FIB/SEM analysis illustrating the 3D visualization of all the Golgi complexes (red) detected in a large volume (∼34 µm3) of a stage II MK. (J) Higher magnifications of the FIB/SEM analysis showing close apposition of Golgi-derived vesicles at the boundary of the DMS. (K) Segmentation of the DMS (yellow) and the Golgi (red). (L) 3D reconstruction. *DMS. Bars, 1 µm. g, Golgi stacks; n, nucleus.
