Figure 4. Rab3GAP is required for ER... | Rockefeller University Press

$Figure 4. Rab3GAP is required for ER localization of Rab18. (a) HeLa cells expressing GFP-Rab18 were depleted of Rab3GAP subunits (siGAP1 and siGAP2) for 72 h and then stained with a Rab3GAP1 antibody. The enlarged region from the yellow boxed area shows details of Rab18 localization. (b) The presence of GFP-Rab18 on ER tubules was scored for all conditions and plotted in the bar graph. Error bars indicate the standard deviation of the mean (n = 3 independent experiments). (c) HeLa cells were depleted of Rab3GAP subunits (GAP1 and GAP2) for 72 h. Western blots show the distribution of endogenous Rab18 to the membrane pellet (P) and soluble (S) cytosol fractions marked by the Golgi membrane protein GM130 and tubulin, respectively. Rab3GAP1 depletion was confirmed by Western blotting. (d) COS7 cells expressing a GFP-tagged ER marker and mCherry-tagged Rab18 were depleted of Rab3GAP subunits alone or in combination or Rab18 for 72 h then imaged at 1.5-s intervals using a spinning-disk confocal microscope. The entire cell is shown for t = 0, and a time series from the yellow boxed area is depicted in the enlarged regions. Bars are marked in the figure.$

Figure 4.

Rab3GAP is required for ER localization of Rab18. (a) HeLa cells expressing GFP-Rab18 were depleted of Rab3GAP subunits (siGAP1 and siGAP2) for 72 h and then stained with a Rab3GAP1 antibody. The enlarged region from the yellow boxed area shows details of Rab18 localization. (b) The presence of GFP-Rab18 on ER tubules was scored for all conditions and plotted in the bar graph. Error bars indicate the standard deviation of the mean (n = 3 independent experiments). (c) HeLa cells were depleted of Rab3GAP subunits (GAP1 and GAP2) for 72 h. Western blots show the distribution of endogenous Rab18 to the membrane pellet (P) and soluble (S) cytosol fractions marked by the Golgi membrane protein GM130 and tubulin, respectively. Rab3GAP1 depletion was confirmed by Western blotting. (d) COS7 cells expressing a GFP-tagged ER marker and mCherry-tagged Rab18 were depleted of Rab3GAP subunits alone or in combination or Rab18 for 72 h then imaged at 1.5-s intervals using a spinning-disk confocal microscope. The entire cell is shown for t = 0, and a time series from the yellow boxed area is depicted in the enlarged regions. Bars are marked in the figure.

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