Institute of Aquaculture, University of Stirling, Stirling FK94LA, Scotland, UK.
Aquat Toxicol. 2010 Apr 1;97(1):23-33. doi: 10.1016/j.aquatox.2009.11.017. Epub 2009 Dec 16.
Copper (Cu) is an essential metal, although in excess is highly toxic due to its redox properties and, therefore intracellular Cu homeostasis is a highly regulated process. Cu-ATPases are pivotal regulatory, proteins of intracellular and bodily Cu homeostasis. Two Cu-ATPases, ATP7A and ATP7B with distinct, functions are found in mammals and herein we report the structure and expression under Cu stress of, homologues of ATP7A and ATP7B in gilthead sea bream (Sparus aurata), the first such report for any, fish. The deduced protein sequences of S. aurata ATP7A (saATP7A) and ATP7B (saATP7B), displayed 63% and 75% identity respectively to their human homologues. All characteristic structural, features of Cu-ATPases were conserved between fish and mammals, although the number of Cu-binding, domains was less in fish ATP7B than in mammalian ATP7B. The tissue expression of sea bream, Cu-ATPases was similar to that observed in mammals, saATP7A being ubiquitously expressed, although low in liver, whilst saATP7B was mainly expressed in the intestine and liver. By analysis of the sequenced genomes of other species we have confirmed the presence of ATP7A and ATP7B genes in fish and propose that the presence of two Cu-ATPase genes in vertebrates represents a retention and neo-functionalization of a duplicated ancestral gene coincident with the development of a closed circulatory system and discrete hepato-biliary system. Expression of Cu-ATPase mRNA was changed after exposure to excess Cu in a manner dependent on exposure route and tissue type. Excess dietary Cu (130mgkg(-1) Cu dry diet) reduced saATP7A mRNA levels in intestine, gill, kidney and liver, and increased hepatic saATP7B mRNA consistent with increased biliary excretion. Whilst after waterborne Cu exposure (0.3mgL(-1) Cu), expression of ATP7A mRNA was increased in intestine and liver and toxic responses were observed in gill and liver. Our results indicate that Cu-ATPases in both fish and mammals have similar functions in maintenance of Cu homeostasis and are consistent with previous physiological evidence from various fish species for the involvement of multiple Cu-ATPases in Cu transport. Furthermore, our evidence suggests that fish can detoxify excess dietary Cu relatively efficiently but are unable to cope with excess dissolved Cu in the water, demonstrating that the exposure route is critical to toxicity.
铜（Cu）是一种必需的金属，但由于其氧化还原性质，过量的铜是高度有毒的，因此细胞内的铜稳态是一个高度调节的过程。Cu-ATPases 是细胞内和体内铜稳态的重要调节蛋白。哺乳动物中存在两种具有不同功能的 Cu-ATPases，即 ATP7A 和 ATP7B，而我们在此报告了金头鲷（Sparus aurata）中 ATP7A 和 ATP7B 的同源物在 Cu 胁迫下的结构和表达，这是鱼类中首次进行此类报告。金头鲷的 ATP7A（saATP7A）和 ATP7B（saATP7B）的推导蛋白序列分别与人同源物具有 63%和 75%的同一性。鱼类和哺乳动物之间 Cu-ATPases 的所有特征结构特征均得以保守，尽管鱼类的 Cu-ATPase 中的 Cu 结合结构域数量少于哺乳动物的 ATP7B。金头鲷的组织表达与哺乳动物观察到的相似，saATP7A 广泛表达，尽管在肝脏中含量较低，而 saATP7B 主要在肠和肝脏中表达。通过对其他物种的基因组序列分析，我们已经证实鱼类中存在 ATP7A 和 ATP7B 基因，并提出脊椎动物中存在两种 Cu-ATPase 基因是与封闭循环系统和离散的肝胆系统的发展相吻合的对祖先基因的保留和新功能化。暴露于过量铜后，Cu-ATPase mRNA 的表达发生变化，这取决于暴露途径和组织类型。过量的饮食铜（130mgkg(-1) Cu 干饮食）降低了肠、鳃、肾和肝中的 saATP7A mRNA 水平，并增加了肝中的 saATP7B mRNA，这与胆汁排泄增加一致。而在经水暴露（0.3mgL(-1) Cu）后，肠和肝中的 ATP7A mRNA 表达增加，并且在鳃和肝中观察到毒性反应。我们的结果表明，鱼类和哺乳动物中的 Cu-ATPase 在维持铜稳态方面具有相似的功能，并且与来自各种鱼类的先前生理证据一致，即多种 Cu-ATPase 参与 Cu 转运。此外，我们的证据表明，鱼类可以相对有效地解毒过量的饮食铜，但无法应对水中过量的溶解铜，这表明暴露途径对毒性至关重要。
