Lavoie Michel, Le Faucheur Séverine, Fortin Claude, Campbell Peter G C
Institut National de la Recherche Scientifique-Eau, Terre et Environnement (INRS-ETE), Québec, Canada.
Aquat Toxicol. 2009 Apr 9;92(2):65-75. doi: 10.1016/j.aquatox.2008.12.007. Epub 2008 Dec 27.
The aim of this study was to evaluate whether intracellular detoxification mechanisms could explain, at least partially, the different sensitivity to Cd of two freshwater green algae, Chlamydomonas reinhardtii and Pseudokirchneriella subcapitata. Subcellular Cd distribution and the synthesis of metal-binding thiolated peptides were thus examined in both algae exposed to a range of free [Cd(2+)] from 0.7 to 253 nM. Cadmium partitioning among five subcellular fractions (cellular debris, granules, organelles, heat-denaturable proteins - HDP, and heat-stable proteins - HSP) was determined after differential centrifugation of algal homogenates. Thiolated-peptides, phytochelatins (PC(n)) and precursors, were analyzed by HPLC with pre-column monobromobimane derivatization. Cadmium accumulation per cell was 2-4 times greater for C. reinhardtii than for P. subcapitata, yet C. reinhardtii was more resistant to Cd with an EC(50) of 273 nM Cd(2+) [244-333 nM Cd(2+) CI(95%)]) compared to 127 nM Cd(2+) [111-143 nM Cd(2+) CI(95%)] for P. subcapitata. Although [Cd] generally increased in the organelle fractions when free [Cd(2+)] increased in the experimental media, their relative contributions to the total Cd cellular content decreased, suggesting that partial protection of some metal sensitive sites was achieved by the initiation of cellular detoxification mechanisms. An increase in the proportion of Cd in the granules fraction was observed for C. reinhardtii between 6 and 15 nM Cd(2+) (i.e., at [Cd(2+)]<the threshold for growth inhibition) suggesting the involvement of granules in protecting against the occurrence of toxic effects in C. reinhardtii. Both species also produced also high levels of PC(n), but with longer oligomers for C. reinhardtii. Unknown thiolated compounds (X(n)), which were not canonical or hydroxymethyl PC(n), were also found in both algae but at much higher concentrations for C. reinhardtii than for P. subcapitata. This difference in thiol synthesis could also be involved in the higher Cd resistance of C. reinhardtii with respect to P. subcapitata. This study demonstrates the importance of metal detoxification strategies in explaining the Cd sensitivity of different algal species.
本研究的目的是评估细胞内解毒机制是否至少能部分解释两种淡水绿藻莱茵衣藻（Chlamydomonas reinhardtii）和小形假微型海链藻（Pseudokirchneriella subcapitata）对镉的不同敏感性。因此，在暴露于0.7至253 nM一系列游离[Cd(2+)]的两种藻类中，检测了亚细胞镉分布和金属结合硫醇化肽的合成。在对藻类匀浆进行差速离心后，确定了镉在五个亚细胞组分（细胞碎片、颗粒、细胞器、热变性蛋白 - HDP和热稳定蛋白 - HSP）之间的分配。硫醇化肽、植物螯合肽（PC(n)）及其前体通过柱前单溴代联苯胺衍生化的高效液相色谱法进行分析。莱茵衣藻的每细胞镉积累量比小形假微型海链藻高2 - 4倍，然而莱茵衣藻对镉的抗性更强，其半数有效浓度（EC(50)）为273 nM Cd(2+) [244 - 333 nM Cd(2+) 95%置信区间（CI）]，相比之下，小形假微型海链藻的EC(50)为127 nM Cd(2+) [111 - 143 nM Cd(2+) 95% CI]。尽管当实验培养基中游离[Cd(2+)]增加时，细胞器组分中的[Cd]通常会增加，但其对细胞总镉含量的相对贡献却降低了，这表明通过启动细胞解毒机制实现了对一些金属敏感位点的部分保护。在6至15 nM Cd(2+)之间（即[Cd(2+)] <生长抑制阈值时），观察到莱茵衣藻颗粒组分中镉的比例增加，这表明颗粒参与了保护莱茵衣藻免受毒性作用的发生。两种藻类都产生了高水平的PC(n)，但莱茵衣藻的寡聚体更长。在两种藻类中还发现了未知的硫醇化化合物（X(n)），它们不是典型的或羟甲基PC(n)，但莱茵衣藻中的浓度比小形假微型海链藻高得多。硫醇合成的这种差异也可能与莱茵衣藻相对于小形假微型海链藻对镉的更高抗性有关。这项研究证明了金属解毒策略在解释不同藻类物种对镉敏感性方面的重要性。
