Fig. 7

Plin4 AH expression in Drosophila S2 cells rescues the defect in LD size due to PC depletion. a Plin4-12mer-GFP localizes to LDs. Expression was induced with copper for 48 h. For the last 24 h, cells were either supplemented with oleic acid to induce LDs (+OA) or were kept in normal growth medium (−OA). LDs were stained with Autodot dye. Images were obtained with a spinning disc microscope and are presented as projected z-stacks. The merged images show a ×3 zoom of the area marked in the Plin4 images with LDs in magenta and Plin4 in green. Scale bar: 5 µm. b Western blot analysis of Plin4-12mer-GFP expression in non-induced cells (no copper addition, lane 1), or in copper-induced cells (lanes 2–5) that were subjected to the indicated treatments. Plin4-12mer levels were not affected by oleic acid treatment or by RNAi against CCT1 (c). Tubulin was used as loading control (uncropped membranes in Supplementary Fig. 7). White arrowheads indicate the migration of 70 kDa and 55 kDa molecular weight standards, respectively. c, d Cells were treated with RNAi against CCT1 to deplete PC or with control RNAi, followed by Plin4-12mer-GFP induction and oleic acid treatment; c images show a comparison of LD size between Plin4-transfected and non-transfected cells in the same field, with enlarged images of the merged channels; d graphs show quantification of LD size in individual cells in a representative of three experiments (shown in Supplementary Fig. 6b, c), with bars depicting median values