Sterol transport is independent of vesicle trafficking. (A) Treatment of cells with 300 µM BFA leads to collapse of Anp1-GFP–marked Golgi to the ER, but has no effect on mCherry-D4H distribution at the PM. (B) Cells pretreated with BFA for 30 min to induce Golgi collapse and then incubated in the presence of BFA and CK-666 to induce sterol internalization display three principal phenotypes: (i) mCherry-D4H at the PM with occasional internal dot-like structure that coincided with Syb1 (arrowheads), (ii) mCherry-D4H at the PM and internal haze, with dot like-structures that coincided with Syb1, (iii) weak mCherry-D4H at the PM with internal haze and vacuolar staining. The right panel shows control treatments. (C) Cells were treated for 1 h with CK-666, and then 1 h more with CK-666, DMSO, or BFA. (D) mCherry-D4H localization during steady-state growth (DMSO), upon 1 h CK-666 treatment, and after 1 h recovery in exomer-deficient mutants (bch1Δ cfr1Δ), mutants lacking the AP-1 adaptor complex subunit (apm1Δ), and triple mutant (bch1Δ cfr1Δ apm1Δ). (E) mCherry-D4H localization during steady-state growth (DMSO), upon 1 h CK-666 treatment, and after 1 h recovery in cells in which sec8 expression (under the control of the p^nmt81 promoter) has been repressed for 20 h. Scale bars, 2 µm.