Pantoja O, Smith J A C
Department of Plant Sciences, University of Oxford, South Parks Road Oxford, OX1 3RB, United Kingdom.
J Membr Biol. 2002 Mar 1;186(1):31-42. doi: 10.1007/s00232-001-0132-z.
The organic anion malate is accumulated in the central vacuole of most plant cells. Malate has several important roles in plant vacuoles, such as the maintenance of charge balance and pH regulation, as an osmolyte involved in the generation of cell turgor, and as a storage form of CO2. Transport of malate across the vacuolar membrane is important for the regulation of cytoplasmic pH and the control of cellular metabolism, particularly in plants showing crassulacean acid metabolism (CAM), in which large fluxes of malate occur during the day/night cycle. By applying the patch-clamp technique, in the whole-vacuole configuration, to isolated vacuoles from leaf mesophyll cells of the CAM plant Kalanchoë daigremontiana, we studied the regulation of the vacuolar malate channel by pH and Ca2+, as well as its sensitivity to anion-channel blockers. Malate currents were found to be insensitive to Ca2+ on the cytoplasmic side of the membrane over a range from approximately 10(-8) M to 10(-4) M. In contrast, decreasing cytoplasmic pH below 7.5 had a significant modulatory effect on channel activity, reducing malate currents by 40%, whereas increasing cytoplasmic pH above 7.5 resulted in no change in current. Several known Cl?-channel blockers inhibited the vacuolar malate currents: niflumic acid and indanoyloxyacetic acid (IAA-94) proved to be the most effective inhibitors, exerting half-maximal effects at concentrations of approximately 20 mM, suggesting that the plant vacuolar malate channel may share certain similarities with other classes of known anion channels.
有机阴离子苹果酸在大多数植物细胞的中央液泡中积累。苹果酸在植物液泡中具有几个重要作用，比如维持电荷平衡和pH调节，作为参与细胞膨压产生的渗透剂，以及作为CO2的储存形式。苹果酸跨液泡膜的转运对于细胞质pH的调节和细胞代谢的控制很重要，特别是在具有景天酸代谢（CAM）的植物中，在昼夜循环中会出现大量苹果酸通量。通过在全液泡配置下将膜片钳技术应用于CAM植物落地生根叶片叶肉细胞分离的液泡，我们研究了pH和Ca2+对液泡苹果酸通道的调节，以及其对阴离子通道阻滞剂的敏感性。发现在膜细胞质侧，苹果酸电流在大约10^(-8) M至10^(-4) M的范围内对Ca2+不敏感。相反，将细胞质pH降低至7.5以下对通道活性有显著调节作用，使苹果酸电流降低40%，而将细胞质pH升高至7.5以上则电流无变化。几种已知的Cl-通道阻滞剂抑制液泡苹果酸电流：氟尼辛和茚满氧基乙酸（IAA-94）被证明是最有效的抑制剂，在浓度约为20 mM时发挥半数最大效应，这表明植物液泡苹果酸通道可能与其他已知阴离子通道类别有某些相似之处。
