Cellular responses to environmental contaminants in amoebic cells of the slime mould Dictyostelium discoideum.
Amoebic Dictyostelium discoideum cells were employed in a bioassay to evaluate stress responses after exposures to the polyaromatic hydrocarbon benzo[a]pyrene (B[a]P) and two heavy metals (copper and mercury). Furthermore, we developed a recombinant cell line expressing a labile Green Fluorescent Protein (GFP) variant expressed under the control of an actin promoter to monitor stress-related protein degradation. Finally, cell viability was monitored to discriminate lethal exposure concentrations. The results demonstrated that exposure to sub-micromolar concentrations of mercury rendered significant changes in all studied physiological parameters, whereas B[a]P became toxic at low micromolar, and copper at high micromolar concentrations. Exposure to 0.5 microM mercury significantly reduced lysosomal membrane stability (LMS), endocytosis rate, GFP expression, and further resulted in the elevation of cytosolic free Ca(2+) ([Ca(2+)](i)). LMS in mercury-treated cells that had been pre-incubated with a specific Ca(2+)-dependent phospholipase A2 blocking agent was however not affected by the exposure, indicating that the toxic action of mercury is linked to the activation of phospholipase A2 via a Ca(2+)-signaling pathway. Exposure to 20 microM B[a]P significantly reduced LMS, endocytosis rate, and GFP expression, however without affecting [Ca(2+)](i), suggesting a calcium-independent route of toxicity for this compound. None of the physiological parameters were significantly affected by copper exposure at concentrations <400 microM, demonstrating a high resistance to this metal. Our results further showed that neither cell growth nor viability was affected by concentrations altering the studied physiological parameters. LMS, endocytosis rate, and [Ca(2+)](I), therefore, appear sensitive biomarkers of pollutant-related stress in amoebic cells.
Dondero
Francesco
F
Department of Environmental and Life Sciences (DiSAV), University of Piemonte Orientale, Alessandria, Italy. fdondero@unipmn.it
Jonsson
Henrik
H
Rebelo
Mauro
M
Pesce
Gabriella
G
Berti
Elena
E
Pons
Giovanni
G
Viarengo
Aldo
A
D000818:Animals; D001564:Benzo(a)pyrene / Q000633:toxicity; D001681:Biological Assay / Q000379:methods; D002118:Calcium / Q000378:metabolism; D003300:Copper / Q000633:toxicity; D004023:Dictyostelium / Q000187:drug effects* / Q000254:growth & development / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis; D004784:Environmental Monitoring / Q000379:methods; D004785:Environmental Pollutants / Q000633:toxicity*; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007425:Intracellular Membranes / Q000187:drug effects*; D008247:Lysosomes / Q000187:drug effects*; D008628:Mercury / Q000633:toxicity; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors
D004785:Environmental Pollutants; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D001564:Benzo(a)pyrene; D003300:Copper; D008628:Mercury; D002118:Calcium