Presence of Ecotoxicologically Relevant Pgp and MRP Transcripts and Proteins in Cyprinid Fish

Roberta Klobučar,Damjan Franjević,Tvrtko Smital,Roko Žaja,Anamaria Brozović

doi:10.2478/10004-1254-61-2010-2008

Roberta Klobučar, Damjan Franjević + Show 3 more

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https://doi.org/10.2478/10004-1254-61-2010-2008

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Abstract

One of the most intriguing defence strategies which aquatic organisms developed through evolution is multixenobiotic resistance (MXR). The key mediators of MXR activity are ATP-binding cassette (ABC) transport proteins. They provide resistance of aquatic organisms by binding xenobiotics and extruding them from cells in an energy-dependent manner. Since Cyprinid fish species are common target in freshwater biomonitoring programs, we have studied the presence of two main MDR/MXR efflux transporters P-glycoprotein (Pgp, Abcb1) and MRP-like protein(s) (Abcc) in the liver of five Cyprinid species: common carp, European chub, sneep, barbel, and silver prussian carp. Their presence was evaluated on the mRNA and protein level. Various pairs of primers were designed to clone homologous fragments of MXR-related genes. At the protein level, we used Western blotting with specific monoclonal antibodies against human Pgp (Abcb1, Ab C219), MRP1 (Abcc1; Ab MRPm6) or MRP2 (Abcc2; Ab M2I-4). Transcripts of both key types of MXR transporters were identified in all species examined and here we provide the phylogenetic analysis of new partial sequences. Immunochemical determinations with mammalian antibodies failed to identify the presence of MRP(s), but Pgp expression was found in all five Cyprinid species. These results support that MXR is a defence system mediated by both Pgp and MRP types of ABC transport proteins.

Highlights

One of the most intriguing defence strategies which aquatic organisms developed through evolution is multixenobiotic resistance (MXR)
multidrug resistance-associated protein (MRP) sequence corresponded to Mrp1 (Abcc1) – barbel, Mrp2 (Abcc2) – chub and Prussian carp, and Mrp3 (Abcc3) – common carp
Multixenobiotic resistance phenotype expressed in aquatic organisms serves as a defence mechanism that protects these organisms by pumping structurally diverse xenobiotics out of the cells

Summary

Introduction

One of the most intriguing defence strategies which aquatic organisms developed through evolution is multixenobiotic resistance (MXR). Since Cyprinid fish species are common target in freshwater biomonitoring programs, we have studied the presence of two main MDR/MXR efflux transporters Pglycoprotein (Pgp, Abcb1) and MRP-like protein(s) (Abcc) in the liver of five Cyprinid species: common carp, European chub, sneep, barbel, and silver prussian carp. The primary goal of this study was to determine Pgp (Abcb1) and Mrp (Abcc) related mRNA transcripts in a group of freshwater cyprinidae, as follows: common carp (Cyprinus carpio), European chub (Squalius cephalus), sneep (Chondrostoma nasus), barbel (Barbus barbus), and silver Prussian carp (Carassius auratus gibelio) These species are typical representatives of the fish communities of continental European waters and are frequently used in biomonitoring programmes/studies [21, 22]. We tried to detect Pgp and MRPs on the protein level using commercially available antibodies directed against human Pgp and MRP(s)

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