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. 2017 Apr 15;28(8):1111–1122. doi: 10.1091/mbc.E16-07-0499

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© 2017 Iaea et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 6: — Expression of wild-type, cytosolic-localized STARD4 restores sterol transport between ERC and plasma membrane. U2OS-SRA cells were transfected with mCherry-FKBP-STARD4 with or without MitoTrap 48 h before the experiment. STARD4 CRISPR constructs were transfected for 18 h before the experiment. To redistribute STARD4 from the cytosol to the mitochondria, cells expressing MitoTrap were treated with 1 μM rapamycin for 10 min before imaging. After rapamycin treatment, cells were washed, and fresh medium 2 was added. (A) FRAP measurements for single cells expressing mCherry-FKBP-STARD4 and MitoTrap in cells expressing STARD4 CRISPR constructs. Control, t_1/2 = 14.5 ± 1.5 min; SD4 CRISPR, t_1/2 = 11.0 ± 1.6 min; SD4 CRISPR + SD4 + Mito, t_1/2 = 19.7 ± 1.5 min. (B) DHE efflux curves for cells expressing mCherry-FKBP-STARD4 and MitoTrap in cells expressing STARD4 CRISPR constructs Control, t_1/2 = 14.9 ± 0.9 min; SD4 CRISPR, t_1/2 = 12.5 ± 0.4 min; SD4 CRISPR + SD4 + Mito, t_1/2 = 19.1 ± 0.3 min. Each data point is derived from an average of at least 10 experiments (±SE). Data are fitted to single-exponential curves.