Hernández AgustÍn, Cooke David T, Lewis Mervyn, Clarkson David T
Department of Agricultural Sciences, University of Bristol, Institute of Arable Crops Research, Long Ashton Research Station, Bristol BS18 9AF, UK.
Microbiology (Reading). 1997 Oct;143(10):3165-3174. doi: 10.1099/00221287-143-10-3165.
Plasma membrane vesicles from and sterol-deficient mutants and from wild-type sporidia treated with and without inhibitors of sterol 14α-demethylase or sterol ∆-∆ isomerase (triadimenol and fenpropimorph fungicides, respectively) were purified by aqueous two-phase partitioning. Changes in plasma membrane lipid composition were mostly restricted to sterols and complex lipid-bound fatty acids (CLB fatty acids). There was a greater accumulation of abnormal sterols (14α-methyl-or ∆ unsaturated sterols) in plasma membranes from sterol-deficient mutants than from those treated with their fungicide counterparts. However, greater growth inhibition was observed on fungicide-treated wild-type than on mutants. Changes in CLB fatty acids were restricted to alterations in the relative proportion of linoleic acid (18:2) with respect to oleic acid (18:1). The 18:2 to 18:1 ratio found in CLB fatty acids in plasma membranes could be correlated to rates of sporidial growth but not to accumulation of a particular abnormal sterol or to the extent of sterol replacement. Plasma membrane permeability to protons was increased moderately in the mutants only. No changes were observed in plasma membrane fluidity. Plasma membrane H-ATPase activity was increased up to twofold in those cases with lower growth rate. It was concluded that fungicide-induced growth inhibition in was not due to accumulation of abnormal sterols in plasma membranes but probably due to intracellular ATP depletion by the H-ATPase and that changes in 18:2 to 18:1 ratio in CLB fatty acids were not directly dependent on the plasma membrane physical state or lipid composition but were possibly part of a stress adaptation mechanism.
通过水相双相分配法纯化了来自甾醇缺陷型突变体以及用甾醇14α-脱甲基酶或甾醇Δ-Δ异构酶抑制剂（分别为三唑醇和粉唑醇杀菌剂）处理和未处理的野生型担孢子的质膜囊泡。质膜脂质组成的变化主要局限于甾醇和复合脂质结合脂肪酸（CLB脂肪酸）。与用相应杀菌剂处理的相比，甾醇缺陷型突变体质膜中异常甾醇（14α-甲基或Δ不饱和甾醇）的积累更多。然而，观察到杀菌剂处理的野生型比突变体有更大的生长抑制。CLB脂肪酸的变化仅限于亚油酸（18:2）相对于油酸（18:1）相对比例的改变。质膜中CLB脂肪酸的18:2与18:1比例与担孢子生长速率相关，但与特定异常甾醇的积累或甾醇替代程度无关。仅在突变体中质膜对质子的通透性适度增加。质膜流动性未观察到变化。在生长速率较低的情况下，质膜H-ATPase活性增加至两倍。得出的结论是，杀菌剂诱导的生长抑制不是由于质膜中异常甾醇的积累，而是可能由于H-ATPase导致细胞内ATP消耗，并且CLB脂肪酸中18:2与18:1比例的变化不是直接依赖于质膜的物理状态或脂质组成，而是可能是应激适应机制的一部分。
