Three pools of plasma membrane cholesterol and their relation to cholesterol homeostasis
- University of Texas Southwestern Medical Center, United States
Figures
Figure 1
Movement of LDL-derived cholesterol from lysosomes to cell surface and to ER in human fibroblasts.
On day 0, SV-589 cells were set up in medium A at 1 × 105 cells per 60-mm dish. On day 2, cells were switched to lipoprotein-deficient medium C. On day 3, cells were refed with medium C containing …
Figure 2
Kinetics of transport of LDL-derived cholesterol from lysosomes to PMs and ER.
On day 0, SV-589 cells were set up in medium A at 1 × 105 cells per 60-mm. On day 2, cells were switched to lipoprotein-deficient medium C. On day 3, cells were treated with fresh medium B …
Figure 3 with 1 supplement
Effect of SMase treatment of human fibroblasts on amount of cell surface binding of 125I-PFO* (A) and PM content of SM and ceramide (B–D).
On day 0, SV-589 cells were set up in medium A at 1 × 105 cells per 60-mm dish (A–D). On day 2, cells were switched to lipoprotein-deficient medium C. On day 3, cells were treated with fresh medium …
Figure 3—figure supplement 1
Movement of FCS-derived cholesterol and effect of SMase treatment in hamster cells.
(A) Movement of FCS-derived cholesterol from lysosomes to cell surface and to ER in hamster cells. On day 0, CHO-7 cells were set up in lipoprotein-deficient medium G at 3 × 105 cells per 60-mm …
Figure 4
Effect of LDL on SM-sequestered pool of PM cholesterol.
(A and B) On day 0, SV-589 cells were setup in medium A at 1 × 105 cells per 60-mm dish. On day 2, cells were switched to lipoprotein-deficient medium C. On day 3, cells were treated with fresh …
Figure 5
Effect of paraformaldehyde on 125I-PFO* binding to human fibroblasts after treatment with SMase.
On day 0, SV-589 cells were setup in medium A at 1 × 105 cells per 60-mm dish. On day 2, cells were switched to lipoprotein-deficient medium C. On day 3, cells were treated with fresh medium C …
Figure 6
Time course of 125I-PFO* binding and cholesterol esterification in human fibroblasts after treatment with SMase.
On day 0, SV-589 cells were setup in medium A at 1 × 105 cells per 60-mm dish. On day 2, cells were switched to lipoprotein-deficient medium C. On day 3, cells were treated with fresh medium C …
Figure 7
Prior incubation of human fibroblasts with SMase stimulates transport of cholesterol from PM to ER.
On day 0, SV-589 cells were set up in medium A at 1 × 105 cells per 60-mm dish. On day 2, cells were switched to lipoprotein-deficient medium C. On day 3, cells were treated with fresh medium C …
Figure 8
Reduced threshold for 125I-PFO* binding after treatment of human fibroblasts with SMase.
On day 0, SV-589 cells were set up in medium A at 1 × 105 cells per 60-mm dish. On day 3, cells were refed with medium B. On day 4, cells were treated for 1 hr at 37°C in medium B containing 50 µM …
Figure 9
Schematic diagram illustrating the three pools of cholesterol in the PM of human fibroblasts under different conditions.
https://doi.org/10.7554/eLife.02882.013Tables
Table 1
Major subspecies of sphingomyelin and ceramide in sterol-repleted (−compactin) and sterol-depleted (+compactin) PMs from SV-589 cells treated without and with SMase
| Lipid | − Compactin | + Compactin | ||
|---|---|---|---|---|
| Species | − SMase | + SMase | − SMase | + SMase |
| mole % of total PM lipids | ||||
| Sphingomyelin | ||||
| C16:0 | 5.38 ± 0.19 | 0.32 ± 0.02 | 8.04 ± 0.48 | 0.58 ± 0.12 |
| C18:0 | 0.11 ± 0.00 | 0.01 ± 0.00 | 0.16 ± 0.01 | 0.01 ± 0.00 |
| C24:0 | 0.35 ± 0.01 | 0.01 ± 0.00 | 0.55 ± 0.03 | 0.02 ± 0.00 |
| C24:1 | 2.59 ± 0.22 | 0.14 ± 0.01 | 4.41 ± 0.23 | 0.27 ± 0.05 |
| Ceramide | ||||
| C16:0 | 0.04 ± 0.00 | 2.95 ± 0.50 | 0.07 ± 0.01 | 3.93 ± 1.22 |
| C18:0 | 0.00 ± 0.00 | 0.11 ± 0.01 | 0.01 ± 0.00 | 0.15 ± 0.04 |
| C24:0 | 0.01 ± 0.00 | 0.25 ± 0.04 | 0.01 ± 0.00 | 0.36 ± 0.11 |
| C24:1 | 0.02 ± 0.00 | 1.39 ± 0.16 | 0.04 ± 0.00 | 2.14 ± 0.60 |
-
Lipids from the purified PMs isolated from the cells used in Figure 3 were extracted with 85:15 (vol/vol) ethyl acetate: isopropanol, and the contents of the four indicated acyl chain subspecies of SM and ceramide were quantified as described in ‘Materials and methods’. The data are expressed as mole % of total PM lipids and represent the mean ± SEM obtained from three independent experiments with duplicate measurements of each sample. Levels of SM and ceramide with oleoyl (18:1), arachidoyl (20:0), and behenoyl (22:0) acyl chains are not included in this table as their levels were less than 0.1% of total PM lipids.
