Figure 6.
Silencing of Rab35 causes the loss of the transversal apical membrane bulkheads and formation of spherical cysts via a cell self-organization process. (a) In the cells treated with Rab35 siRNA, the lumina tend to grow as spheres instead of elongating as tubes. Images from the live-cell time-lapse microscopy experiment showing two neighboring differentiating hepatoblasts expressing LifeAct-EGFP under Rab35 siRNA conditions. The white star indicates the forming lumen between the two cells. Note that the typical transverse striped actin pattern observed in the tubular BC is absent. Scale bar: 10 µm. See also Video 8. (b) Multicellular cyst-like structures form by cell rearrangements. Images from the live-cell time-lapse microscopy experiment. The cells self-organize in such a way that the three separate lumina (black star) eventually fuse into one large spherical lumen, in the absence of cell division. Scale bar: 10 µm. See also Video 9. (c) EM analysis of a cyst-like lumen resulting from the Rab35 knock-down. A series of longitudinal 90-nm sections of the lumen formed between five cells. The bulkheads typical for hepatocyte BC are absent in the lumen. Arrows indicate TJs. GG, glycogen granules. Scale bar: 5 µm. (d) Longitudinal view (left) through the middle of a 3D model of the lumen based on rendering plasma membranes and TJs (red) on serial sections. The five cells forming the lumen are represented in different colors. Red arrows point to the TJs at which the cyst is cut open to reveal the sagittal view (right). The lumen has a circular profile, and TJs do not protrude into the lumen.

Silencing of Rab35 causes the loss of the transversal apical membrane bulkheads and formation of spherical cysts via a cell self-organization process. (a) In the cells treated with Rab35 siRNA, the lumina tend to grow as spheres instead of elongating as tubes. Images from the live-cell time-lapse microscopy experiment showing two neighboring differentiating hepatoblasts expressing LifeAct-EGFP under Rab35 siRNA conditions. The white star indicates the forming lumen between the two cells. Note that the typical transverse striped actin pattern observed in the tubular BC is absent. Scale bar: 10 µm. See also Video 8. (b) Multicellular cyst-like structures form by cell rearrangements. Images from the live-cell time-lapse microscopy experiment. The cells self-organize in such a way that the three separate lumina (black star) eventually fuse into one large spherical lumen, in the absence of cell division. Scale bar: 10 µm. See also Video 9. (c) EM analysis of a cyst-like lumen resulting from the Rab35 knock-down. A series of longitudinal 90-nm sections of the lumen formed between five cells. The bulkheads typical for hepatocyte BC are absent in the lumen. Arrows indicate TJs. GG, glycogen granules. Scale bar: 5 µm. (d) Longitudinal view (left) through the middle of a 3D model of the lumen based on rendering plasma membranes and TJs (red) on serial sections. The five cells forming the lumen are represented in different colors. Red arrows point to the TJs at which the cyst is cut open to reveal the sagittal view (right). The lumen has a circular profile, and TJs do not protrude into the lumen.

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