FIGURE 3.

Cer gel solubilization by Chol is reversible. A, Cer domain formation upon Chol oxidation by Chol oxidase. DPH and t-PnA steady-state fluorescence anisotropy at 24 °C in POPC/Chol (60:40) mixtures containing 12 mol % PCer, before (green) and after (orange) Chol oxidase treatment. The samples were treated with 1.5 units/ml at 37 °C. In control samples, enzyme was replaced by buffer (blue). The data correspond to the average of at least three independent experiments. Comparison of means was made by Student's t test. Statistical significance was considered for p < 0.05 (indicated by * and by ** when p < 0.001). No significant differences were found between control and untreated samples. B–F, temperature effects on PCer-rich gel domain formation and solubilization are shown. Fluorescence lifetime components of t-PnA, POPC/Chol (3:2) (B); POPC/Chol (3:2) with 12 mol % PCer (C); PSM/POPC/Chol (1:1:1) (D), and PSM/POPC/Chol (1:1:1) with 4 mol % PCer (E) at initial temperature (24 °C, green), after cooling (15 °C, orange), and after reheating (24 °C*, blue). When Cer gel is formed, an extra long component (τ4) is necessary to properly describe the decay. F, Cer domain solubilization and re-formation by heating-cooling. Fluorescence lifetime components of t-PnA are shown in PSM/POPC/Chol (1:1:1) with 8 mol % PCer at initial temperature (24 °C, green), after heating (45 °C, orange), and after recooling (24 °C*, blue).