Extended Data Fig. 10: PGP depletion-induced Golgi fragmentation can be restored by glycerol in mammalian cells.
From: Glycerol mediates crosstalk between metabolism and trafficking through the golgin Imh1
(a) Human PGP was knocked down by two individual siRNAs (siPGP#6 and siPGP#7) in HeLa cells to observe the subcellular localization of p230 (red) and B4GALT1 (green). Glycerol (0.1 M) was applied to the indicated groups for an additional 1 h. DAPI is the nuclear stain. Scale bar, 20 mm. PGP-knockdown efficiency was detected by western blotting. Golgi fragmentation (p230 and B4GALT1) was quantified (N = 3, n = 15). Data are reported as the mean ± SD of three independent experiments. (One-way ANOVA with Dunnett’s post-hoc multiple comparison test) (b) HeLa cells were treated with control siRNA (siCtrl) and supplied with 0.1 M glycerol for 1 h. Subcellular localization of p230 (red), B4GALT1 (green), and DAPI (blue) were stained. Scale bar, 20 mm. Golgi fragmentation (p230 and B4GALT1) was quantified (N = 3, n = 20). Data are reported as the mean ± SD of three independent experiments. (N.S. not significant; two-sided unpaired t test). (a-b) Golgi fragmentation was defined by calculating the number of disconnected Golgi marker signals (shown as fragments here) detected by Imaris software. Each statistical spot represents the average vesicles/cell in an image captured at 400X magnification that containing ~35 cells. Detailed settings are described in the Methods. Data collected from the same biological replicates were labeled in the same color.
