Alfred Wegener Institute for Polar and Marine Research, Am Handelshafen 12, 27570 Bremerhaven, Germany.
Toxicon. 2011 Sep 1;58(3):247-58. doi: 10.1016/j.toxicon.2011.06.004. Epub 2011 Jun 30.
Certain allelochemicals of the marine dinoflagellate Alexandrium tamarense cause lysis of a broad spectrum of target protist cells but the lytic mechanism is poorly defined. We first hypothesized that membrane sterols serve as molecular targets of these lytic compounds, and that differences in sterol composition among donor and target cells may cause insensitivity of Alexandrium and sensitivity of targets to lytic compounds. We investigated Ca(2+) influx after application of lytic fractions to a model cell line PC12 derived from a pheochromocytoma of the rat adrenal medulla to establish how the lytic compounds affect ion flux associated with lysis of target membranes. The lytic compounds increased permeability of the cell membrane for Ca(2+) ions even during blockade of Ca(2+) channels with cadmium. Results of a liposome assay suggested that the lytic compounds did not lyse such target membranes non-specifically by means of detergent-like activity. Analysis of sterol composition of isolates of A. tamarense and of five target protistan species showed that both lytic and non-lytic A. tamarense strains contain cholesterol and dinosterol as major sterols, whereas none of the other tested species contain dinosterol. Adding sterols and phosphatidylcholine to a lysis bioassay with the cryptophyte Rhodomonas salina for evaluation of competitive binding indicated that the lytic compounds possessed apparent high affinity for free sterols and phosphatidylcholine. Lysis of protistan target cells was dose-dependently reduced by adding various sterols or phosphatidylcholine. For three tested sterols, the lytic compounds showed highest affinity towards cholesterol followed by ergosterol and brassicasterol. Cholesterol comprised a higher percentage of total sterols in plasma membrane fractions of A. tamarense than in corresponding whole cell fractions. We conclude therefore that although the molecular targets of the lytic compounds are likely to involve sterol components of membranes, A. tamarense must have a complex self-protective mechanism that still needs to be addressed.
海洋甲藻亚历山大藻的某些化感物质会导致广谱靶原生质细胞裂解，但裂解机制尚未完全阐明。我们首先假设细胞膜甾醇是这些裂解化合物的分子靶标，并且供体和靶细胞之间甾醇组成的差异可能导致亚历山大藻不敏感而靶细胞对裂解化合物敏感。我们研究了裂解物施加于大鼠肾上腺髓质嗜铬细胞瘤衍生的模型细胞系 PC12 后 Ca(2+)内流，以确定裂解化合物如何影响与靶细胞膜裂解相关的离子流。裂解化合物即使在镉阻断 Ca(2+)通道的情况下，也会增加细胞膜对 Ca(2+)离子的通透性。脂质体测定结果表明，裂解化合物并非通过类似去污剂的活性非特异性地裂解靶细胞膜。对亚历山大藻分离株和五种靶原生动物物种的甾醇组成的分析表明，裂解和非裂解亚历山大藻菌株均以胆固醇和二甾醇为主要甾醇，而其他测试物种均不含二甾醇。将甾醇和磷脂酰胆碱添加到隐藻 Rhodomonas salina 的裂解生物测定中，用于评估竞争结合，表明裂解化合物对游离甾醇和磷脂酰胆碱具有明显的高亲和力。向隐藻 Rhodomonas salina 的裂解生物测定中添加各种甾醇或磷脂酰胆碱可降低靶原生动物细胞的裂解程度。对于三种测试甾醇，裂解化合物对胆固醇的亲和力最高，其次是麦角甾醇和 brassicasterol。胆固醇在亚历山大藻质膜部分的总甾醇中所占比例高于相应的全细胞部分。因此，我们得出结论，尽管裂解化合物的分子靶标可能涉及膜的甾醇成分，但亚历山大藻必须具有复杂的自我保护机制，这仍有待解决。
