FIT2-interacting proteins in early steps of LD formation. (A) Representative images of remaining LDs after delipidation and nascent LDs formed after OA treatment in control and ER tubule-forming protein–depleted COS-7 cells. Each group of cells expressing LiveDrop was delipidated for 60 h and then treated with 0.2 mM OA for 15 min to induce nascent LD formation. Quantification of LDs is shown on the right. n = 36–42 cells/group. Unpaired t test; NS, P > 0.05. Scale bar, 10 µm. (B) As in A, but with cells overexpressing Flag–Climp-63. n = 33–37 cells/group. One-way ANOVA, ***, P < 0.001. Scale bar, 10 µm. (C) As in A, but nascent LDs in septin 2 knockdown COS-7 cells were counted. The bottom images show enlargements of the boxed regions over the tubular ER network. n = 13–16 cells/group. Unpaired t test, ***, P < 0.001. Scale bar, 10 µm; 3 µm (inset). Error bars represent SEM. (D) Time-lapse stills of nascent LD formation in control or septin 2 knockdown COS-7 cells. The histogram on the right shows the quantification of the frequency of nascent LDs in the indicated time frame. n = 3 or 4 cells/group. Scale bar, 5 µm. siControl, small interfering Control; siFIT2, small interfering FIT2; siREEP5, small interfering siREEP5; siRtn4, small interfering Rtn4; siSEPT, small interfering SEPT.