Salinity-dependent copper accumulation in the guppy Poecilia vivipara is associated with CTR1 and ATP7B transcriptional regulation

Abstract

Copper (Cu) accumulation and regulation of key-genes involved in Cu homeostasis were evaluated infreshwater- and saltwater-acclimated guppies Poecilia vivipara. Fish were exposed (96 h) to environ-mentally relevant concentrations of dissolved Cu (0, 5.0, 9.0 and 20.0 g/L). In freshwater guppies, gilland liver Cu accumulation was dependent on Cu concentration in the exposure medium. In saltwaterguppies, this dependence was observed only in the gut. These findings indicate that Cu accumulation wassalinity- and tissue-dependent. Key genes involved in Cu metabolism were sequenced for the first timein P. vivipara. Transcripts coding for the high-affinity copper transporter (CTR1) and copper-transportingATPase (ATP7B) were identified using polymerase chain reaction (PCR) and gene sequencing. The full-length CTR1 open reading frame (1560 bp) and a partial ATP7B (690 bp) were discovered. Predicted aminoacid sequences shared high identities with the CTR1 of Fundulus heteroclitus (81%) and the ATP7B of Sparusaurata (87%). Basal transcriptional levels addressed by RT-qPCR in control fish indicate that CTR1 andATP7B was highly transcribed in liver of freshwater guppies while CTR1 was highly transcribed in gut ofsaltwater guppies. This could explain the higher Cu accumulation observed in liver of freshwater guppiesand in gut of saltwater guppies, because CTR1 is involved in Cu uptake. Reduced gill mRNA expression ofCTR1 was observed in freshwater guppies exposed to 20.0 g/L Cu and in saltwater guppies exposed to5.0 g/L Cu. In turn, reduced mRNA expression of gut ATP7B was observed in freshwater and salt waterguppies exposed to 9.0 and 20.0 g/L Cu. Liver CTR1 and ATP7B transcription were not affected by Cuexposure. These findings suggest that gill CTR1 and gut ATP7B are down-regulated to limit Cu absorptionafter exposure to dissolved Cu, while liver CTR1 and ATP7B levels are maintained to allow Cu storageand detoxification. In conclusion, findings reported here indicate that Cu accumulation in the euryhalineguppy P. vivipara is tissue specific and dependent on water salinity. They also suggest that Cu homeostasisinvolves a differential transcriptional regulation of the newly identified Cu transporters, CTR1 and ATP7B.