Modulation of endoplasmic reticulum-bound cholesterol regulatory enzymes by iron/ascorbate-mediated lipid peroxidation

Mammalian sterol regulatory enzymes are integral membrane proteins of the e ndoplasmic reticulum. They play a critical role in Liver cholesterol homeos tasis and the maintenance of overall cholesterol balance in different speci es. Because lipid peroxidation has been implicated in hepatic dysfunction a nd atherosclerosis, we hypothesized that its occurrence could alter the com position and properties of the bilayer lipid environment, and thereby affec t the functions of these membrane proteins. Preincubation of rat liver micr osomes with iron (Fe)/ascorbate (50 mu M/200 mu M), known to induce peroxid ation, resulted in a significant inhibition of (i) the rate-limiting enzyme in cholesterol biosynthesis, HMG-CoA reductase (46%, p < .01), (ii) the cr ucial enzyme controlling the conversion of cholesterol in bile acids, chole sterol 7 alpha-hydroxylase (48%, p < .001), and (iii) the central enzyme fo r cholesterol esterification: Acyl-CoA:cholesterol acyltransferase (ACAT, 8 0%, p < .0001). The disturbances of these key enzymes took place concomitan tly with the high production of malondialdehyde (350%, p < .007) and the lo ss of polyunsaturated fatty acids (36.19 +/- 1.06% vs. 44.24 +/- 0.41 in co ntrols, p < .0008). While alpha-tocopherol simultaneously neutralized lipid peroxidation, preserved microsomal fatty acid status, and restored ACAT ac tivity, ii was not effective in preventing Fe/ascorbate-induced inactivatio n of both HMG-CoA reductase (44%, p < .01) and cholesterol 7 alpha-hydroxyl ase (71%, p < .0001). These results indicate that Fe/ascorbate alters the a ctivity of the rate-determining steps in liver cholesterol metabolism, eith er directly or via lipid peroxidation, capable of modifying their membrane environment. The present data also suggest that the three regulatory enzyme s respond differently when exposed to Fe/ascorbate or antioxidants, which m ay be due to dissimilar mechanisms. (C) 2000 Elsevier Science Inc.