SCAP knockdown does not disrupt cholesterol metabolism under cholesterol-fed conditions. (A) siRNA-mediated knockdown of SCAP in HeLa cells. The graph shows determination of the expression levels of SCAP at 72 h after siRNA transfection. Data are means ± SEM (n = 3 independent experiments; ***, P < 0.001; unpaired two-tailed Student’s t test). (B) Determination of mRNA levels of the indicated genes in control (Cont) and SCAP knockdown cells by quantitative real-time PCR. Data are means ± SEM (n = 4 independent experiments; **, P < 0.01; ***, P < 0.005; unpaired two-tailed Student’s t test). (C) Total cholesterol (Chol) levels in control and SCAP knockdown cells. Data are means ± SEM (n = 6 independent experiments; unpaired two-tailed Student’s t test). (D and E) shRNA-mediated knockdown of SCAP in HeLa cells. (E) Expression levels of SCAP, HMGR, and LDLR in parental HeLa (control) and shSCAP HeLa cells. Data are means ± SEM (n = 4 independent experiments; *, P < 0.05; ****, P < 0.001; unpaired two-tailed Student’s t test). (F) Total cholesterol levels in parental HeLa and shSCAP HeLa cells. Data are means ± SEM (n = 3 independent experiments; unpaired two-tailed Student’s t test). Asterisks on the Western blots denote nonspecific bands. IB, immunoblotted.