title: Effects of cadmium exposure on expression and activity of P-glycoprotein in eastern oysters, Crassostrea virginica Gmelin.
abstract: Heavy metal pollution is a worldwide problem, and cadmium (Cd) is one of the most noxious pollutants in aquatic environments. We studied P-glycoprotein (P-gp) expression and function in control and Cd exposed (50microgL(-1) Cd, 30-40 days) oysters Crassostrea virginica as a possible mechanism of cell protection against Cd. Our data show that P-gp is expressed on cell membrane and in mitochondria of oyster gills and hepatopancreas. Inhibitor studies with verapamil, cyclosporine A and JS-2190 suggest that in the gills, mitochondrial P-gp pumps substrates from cytosol into the mitochondria, while cell membrane P-gp pumps substrates from cytosol out of the cell. Cd exposure resulted in a 2-2.5-fold increase in P-gp protein expression in cell membranes and a 3.5-7-fold increase in transport activity measured as the inhibitor-sensitive rhodamine B extrusion rate. In contrast, p-gp mRNA levels were similar in control and Cd-exposed oysters. No difference in P-gp protein expression was observed between mitochondria of control and Cd-exposed oysters but the apparent transport activity was higher in mitochondria from Cd-exposed oysters. Overall, a stronger increase in substrate transport activity in Cd-exposed oysters compared to a relatively weaker change in P-gp protein levels suggests that P-gp activity is post-translationally regulated. Our data show that direct determination of P-gp transport activity may be the best measure of the xenobiotic-resistant phenotype, whereas p-gp mRNA levels are not a good marker due to the likely involvement of multiple post-transcriptional regulatory steps. Cd exposure resulted in a significantly elevated rate of oxygen consumption of isolated oyster gills by 46%. Specific inhibitors of ATPase function of P-gp (cyclosporine A and JS-2190) had no significant effect on tissue oxygen consumption indicating that P-gp contribution to energy budget is negligible and supporting indirect estimates based on the ATP stoichiometry of substrate transport that also suggest low energy demand for P-gp function.
authors.affiliation: Biology Department, University of North Carolina at Charlotte, 9201 University City Blvd., Charlotte, NC 28223, USA.
authors.forename: Anna V
authors.initials: AV
authors.lastname: Ivanina
authors.forename: Inna M
authors.initials: IM
authors.lastname: Sokolova
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics* / Q000378:metabolism*; D000818:Animals; D002104:Cadmium / Q000633:toxicity*; D019069:Cell Respiration / Q000187:drug effects; D049896:Crassostrea / Q000187:drug effects* / Q000201:enzymology* / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects*; D005880:Gills / Q000187:drug effects / Q000201:enzymology; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology; D014874:Water Pollutants, Chemical / Q000633:toxicity*
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D014874:Water Pollutants, Chemical; D002104:Cadmium