FIGURE 5.

Knockdown of VIMP causes ER spreading. HeLa cells (A) or those stably expressing mRFP-Sec61β (B) were transfected without (Mock) or with one of the siRNAs targeting VIMP (siRNA(210) or siRNA(247)). A, after 72 h, the HeLa cells were fixed and stained with an antibody against Sec61β (left) or CLIMP-63 (right). Sec61β or CLIMP-63 fluorescence along white lines from the nucleus to the cell periphery was analyzed using ImageJ software. The average values of fluorescence brightness in five cells are shown. Scale bar = 5 μm. B, alternatively, HeLa cells stably expressing mRFP-Sec61β were incubated with 5 μg/ml nocodazole, and live cell imaging was performed. Representative images at the indicated times are shown. Arrows indicate peripheral regions, where ER structures were rarely seen before nocodazole treatment. Scale bar = 5 μm.