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. 2016 Nov 22;35(24):2699–2716. doi: 10.15252/embj.201695170

Figure EV2. Supporting information for Figure 2 .

Figure EV2

  1. A431 WT cells with seipin tagged at its chromosomal locus with sfGFP were treated with OA for 1 h and imaged live in the presence of LipidTox Deep Red. Airyscan video snapshots are shown. Orange arrowheads indicate seipin puncta stably associated with LDs.
  2. In relation to Fig 2C–E. WT and SKO cells were treated with OA for 45–75 min, LDs were stained with LD540, and cells were imaged live with widefield microscopy. Shown are two subsequent frames (400 ms apart) and their overlap.
  3. Maximum velocities (average between three subsequent frames) of the tracked LDs in Fig 2E.
  4. WT and SKO cells were grown in standard growth medium, LDs were stained with LD540, and cells were imaged live with widefield microscopy. Shown are two subsequent frames (400 ms apart) and their overlap.
  5. Analysis of (D), bars: 1 – [Pearson's colocalization coefficient between subsequent frames/cell], ± SEM, n = 114–127 cells, two experiments, **P < 0.005 (Mann–Whitney test).
  6. Exemplary LD trajectories of cells treated and imaged as in (D); total duration of tracks is 40 s.
  7. Maximum velocities of LDs (averaged between three subsequent frames) of LDs tracked in Fig 2H, mean ± SEM, n = 29–42 LDs/group (70–91 LDs/genotype), two experiments, *P < 0.05, **P < 0.005 (Kruskal–Wallis test, followed by Dunn test).
  8. SKO cells stably expressing WT‐ or A212P‐seipin‐GFP were transfected with ER marker plasmid (sfGFP‐ER‐3), delipidated overnight, incubated with OA and LipidTox Deep Red for 15–75 min and imaged live with Airyscan microscopy. Phenotyping of LD‐ER‐associated movement as in Fig 2F, bars: % of LDs/ROI displaying indicated mode of movement ± SEM, n = 12 ROIs/genotype (75–65 LDs), one experiment, *P < 0.05 (unpaired t‐test).