Endogenous ACSL3 colocalize with HPos in pre- and emerging LDs. (A) Starved GFP-HPos–transfected cells were fixed and labeled with an antibody for ACSL3. Green arrows indicate colocalization, white arrows indicate equivalent structures to pre-LDs found in nontransfected cells, and blue arrows indicate pre-LDs in central regions of the cell. (B) Distribution of ACSL3, Sec61, and Plin2 in starved cells (top) and cells additionally treated with OA for 24 h (bottom) and fractionated in sucrose density gradients. (C) Distribution of ACSL3 in cells treated with OA for 24 h. (D) Starved GFP-HPos–transfected cells were additionally treated with OA for 7.5 or 15 min, and cells were fixed and labeled with the ACSL3 antibody. Green arrows indicate colocalization. (E) ACSL3 protein levels detected by Western blotting of homogenates of normal cells (control), starved cells (24 h stv), and cells additionally treated with OA for 24 h (24 h OA). Bars: (A and C) 20 µm; (D) 1 µm.