{"title":"乙醇诱导酵母质膜的通透性屏障","authors":"Haruhiko Mizoguchi, Shodo Hara","doi":"10.1016/S0922-338X(97)80348-0","DOIUrl":null,"url":null,"abstract":"<div><p>An increase in extracellular nucleotides appeared to conform to simple diffusion, when <em>Saccharomyces cerevisiae</em> cells were suspended in 20% ethanol at 15°C. However, in the case of cells grown in the presence of 8% ethanol, the leakage of nucleotide appeared to be repressed significantly in the initial phase. The addition of glucose led to a continuation of the repression of the leakage. Under this condition, the addition of iodoacetamide as an inhibitor in glycolytic pathway, or stilbestrol as an inhibitor of plasma membrane ATPase, resulted in a rapid increase in the leakage of nucleotides, indicating that the membrane permeability barrier is dependent on membrane ATPase. Ouabain, an inhibitor of the <span><math><mtext>Na</mtext><msup><mi></mi><mn>+</mn></msup><mtext>K</mtext><msup><mi></mi><mn>+</mn></msup><mtext>-ATPase</mtext></math></span>, had no effect. The addition of 10 mM CaCl<sub>2</sub> for inducing lateral phase separation in the inner membrane, which caused a faster release of nucleotides in general, had a little effect on nucleotide leakage, when cells grown in the presence of 8% ethanol were suspended in 20% ethanol containing glucose. Moreover, the addition of 10 mM KCl, together with CaCl<sub>2</sub>, had almost no effect on leakage. Ca<sup>2+</sup> influx was found to be smaller in cells grown in the presence of ethanol when compared to cells grown in the absence of ethanol, when fura-2 loaded cells were suspended in K-MOPS buffer containing 50 mM CaCl<sub>2</sub>. Divalent cations such as Ba<sup>2+</sup>, Mg<sup>2+</sup> and Mn<sup>2+</sup> had effects similar to Ca<sup>2+</sup> on membrane permeability. The decrease in cell viability corresponded to the amount of leakage of nucleotides over the experimental time course. These results suggest that the high activity of membrane ATPase which is induced by growth in the presence of 8% ethanol ensures homeostasis of ions in the cytoplasm at high concentrations of ethanol, and reduces the effects of membrane surface-acting substances such as divalent cations on the membrane integrity. Thus the maintenance of the cell membrane as a permeability barrier appears to lead to a high ethanol-endurability.</p></div>","PeriodicalId":15696,"journal":{"name":"Journal of Fermentation and Bioengineering","volume":"85 1","pages":"Pages 25-29"},"PeriodicalIF":0.0000,"publicationDate":"1998-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S0922-338X(97)80348-0","citationCount":"24","resultStr":"{\"title\":\"Permeability barrier of the yeast plasma membrane induced by ethanol\",\"authors\":\"Haruhiko Mizoguchi, Shodo Hara\",\"doi\":\"10.1016/S0922-338X(97)80348-0\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>An increase in extracellular nucleotides appeared to conform to simple diffusion, when <em>Saccharomyces cerevisiae</em> cells were suspended in 20% ethanol at 15°C. However, in the case of cells grown in the presence of 8% ethanol, the leakage of nucleotide appeared to be repressed significantly in the initial phase. The addition of glucose led to a continuation of the repression of the leakage. Under this condition, the addition of iodoacetamide as an inhibitor in glycolytic pathway, or stilbestrol as an inhibitor of plasma membrane ATPase, resulted in a rapid increase in the leakage of nucleotides, indicating that the membrane permeability barrier is dependent on membrane ATPase. Ouabain, an inhibitor of the <span><math><mtext>Na</mtext><msup><mi></mi><mn>+</mn></msup><mtext>K</mtext><msup><mi></mi><mn>+</mn></msup><mtext>-ATPase</mtext></math></span>, had no effect. The addition of 10 mM CaCl<sub>2</sub> for inducing lateral phase separation in the inner membrane, which caused a faster release of nucleotides in general, had a little effect on nucleotide leakage, when cells grown in the presence of 8% ethanol were suspended in 20% ethanol containing glucose. Moreover, the addition of 10 mM KCl, together with CaCl<sub>2</sub>, had almost no effect on leakage. Ca<sup>2+</sup> influx was found to be smaller in cells grown in the presence of ethanol when compared to cells grown in the absence of ethanol, when fura-2 loaded cells were suspended in K-MOPS buffer containing 50 mM CaCl<sub>2</sub>. Divalent cations such as Ba<sup>2+</sup>, Mg<sup>2+</sup> and Mn<sup>2+</sup> had effects similar to Ca<sup>2+</sup> on membrane permeability. The decrease in cell viability corresponded to the amount of leakage of nucleotides over the experimental time course. These results suggest that the high activity of membrane ATPase which is induced by growth in the presence of 8% ethanol ensures homeostasis of ions in the cytoplasm at high concentrations of ethanol, and reduces the effects of membrane surface-acting substances such as divalent cations on the membrane integrity. Thus the maintenance of the cell membrane as a permeability barrier appears to lead to a high ethanol-endurability.</p></div>\",\"PeriodicalId\":15696,\"journal\":{\"name\":\"Journal of Fermentation and Bioengineering\",\"volume\":\"85 1\",\"pages\":\"Pages 25-29\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"1998-01-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1016/S0922-338X(97)80348-0\",\"citationCount\":\"24\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of Fermentation and Bioengineering\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0922338X97803480\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Fermentation and Bioengineering","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0922338X97803480","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 24
摘要
当酿酒酵母细胞悬浮在15°C的20%乙醇中时,细胞外核苷酸的增加似乎符合简单扩散。然而,在8%乙醇存在下生长的细胞中,核苷酸的泄漏似乎在初始阶段被显著抑制。葡萄糖的加入导致了对渗漏的持续抑制。在此条件下,加入碘乙酰胺作为糖酵解途径的抑制剂,或烯雌酚作为质膜atp酶的抑制剂,导致核苷酸泄漏量迅速增加,表明膜通透性屏障依赖于膜atp酶。而Na+K+- atp酶抑制剂瓦巴因则没有作用。当在8%乙醇中生长的细胞悬浮在含葡萄糖的20%乙醇中时,加入10 mM CaCl2诱导内膜侧相分离,通常会导致核苷酸释放更快,但对核苷酸泄漏的影响很小。此外,添加10 mM KCl和CaCl2对泄漏几乎没有影响。当载fura-2的细胞悬浮在含有50 mM CaCl2的K-MOPS缓冲液中时,发现在有乙醇存在的细胞中生长的Ca2+内流比在没有乙醇的情况下生长的细胞要小。Ba2+、Mg2+、Mn2+等二价阳离子对膜通透性的影响与Ca2+相似。细胞活力的下降与实验时间过程中核苷酸的泄漏量相对应。这些结果表明,在8%乙醇的存在下,细胞膜atp酶的高活性确保了高浓度乙醇下细胞质中离子的稳态,并减少了膜表面作用物质(如二价阳离子)对膜完整性的影响。因此,细胞膜作为渗透性屏障的维持似乎导致了高乙醇耐受性。
Permeability barrier of the yeast plasma membrane induced by ethanol
An increase in extracellular nucleotides appeared to conform to simple diffusion, when Saccharomyces cerevisiae cells were suspended in 20% ethanol at 15°C. However, in the case of cells grown in the presence of 8% ethanol, the leakage of nucleotide appeared to be repressed significantly in the initial phase. The addition of glucose led to a continuation of the repression of the leakage. Under this condition, the addition of iodoacetamide as an inhibitor in glycolytic pathway, or stilbestrol as an inhibitor of plasma membrane ATPase, resulted in a rapid increase in the leakage of nucleotides, indicating that the membrane permeability barrier is dependent on membrane ATPase. Ouabain, an inhibitor of the , had no effect. The addition of 10 mM CaCl2 for inducing lateral phase separation in the inner membrane, which caused a faster release of nucleotides in general, had a little effect on nucleotide leakage, when cells grown in the presence of 8% ethanol were suspended in 20% ethanol containing glucose. Moreover, the addition of 10 mM KCl, together with CaCl2, had almost no effect on leakage. Ca2+ influx was found to be smaller in cells grown in the presence of ethanol when compared to cells grown in the absence of ethanol, when fura-2 loaded cells were suspended in K-MOPS buffer containing 50 mM CaCl2. Divalent cations such as Ba2+, Mg2+ and Mn2+ had effects similar to Ca2+ on membrane permeability. The decrease in cell viability corresponded to the amount of leakage of nucleotides over the experimental time course. These results suggest that the high activity of membrane ATPase which is induced by growth in the presence of 8% ethanol ensures homeostasis of ions in the cytoplasm at high concentrations of ethanol, and reduces the effects of membrane surface-acting substances such as divalent cations on the membrane integrity. Thus the maintenance of the cell membrane as a permeability barrier appears to lead to a high ethanol-endurability.