Correlation between the level of the potential biomarker, heat-shock protein, and the occurrence of DNA damage in the dab, Limanda limanda: a field study in the North Sea and the English Channel.

In the present study, heat-shock protein of M(r) 70 kDa (HSP70), a marker of cellular stress response, was validated as a potential biomarker under field conditions. The dab, Limanda limanda (female, size > or = 25 cm, spawning maturity stage 2) was used as the indicator organism. The data on HSP level were correlated with the occurrence of DNA damage, measured in the same specimens of L. limanda, to prove the usefulness of the method. The area under investigation was the North Sea. Four locations were selected: station N01, close to Heligoland, in the North Sea; station N04 at the Dogger Bank; station N06 at the Firth of Forth; and station G08 in the English Channel. Ten animals from each location were selected and their livers used for the experiments. The results show that the highest levels of HSP70 (consisting of two forms of M(r) 75 and 73 kDa) were in fish from station N04, while low values were measured in livers from L. limanda collected at station N01. Intermediate levels were seen in the animals from the two other locations. By application of a novel technique, it was found that the extent of DNA damage (single-strand breaks and alkaline labile sites) in fish liver parallels the levels of both HSP70 forms. Our results suggest that L. limanda may be a useful bioindicator and heat-shock proteins, a useful biomarker for monitoring of environmental pollution.

Induction of DNA strand breaks and expression of HSP70 and GRP78 homolog by cadmium in the marine sponge Suberites domuncula.

The marine sponge Suberites domuncula was used as a bioindicator to study the effects of cadmium on the occurrence of DNA strand breakage and on the induction of the expression of the stress biomarkers, heat shock protein 70 (HSP70) and glucose-regulated protein 78 (GRP78) homolog. The cDNA encoding GRP78 homolog from S. domuncula was isolated and characterized. The GRP78 cDNA has a length of 2.1 kb and displays characteristic features of the HSP70 family; it encodes an aa sequence of Mr 72,000. Exposure of S. domuncula to 1 mg/L of cadmium chloride for 24 h caused a strong (16. 6-fold) increase in cadmium content to 7.7 microg/g wet weight of sponge tissue; after an incubation period of 6 days, the accumulation was 20.4-fold. The increase in cadmium content was paralleled by a transient decrease in zinc content at days 1 and 3. Exposure of S. domuncula to cadmium chloride also resulted in a marked increase in the number of DNA single strand breaks, as assessed by a recently developed fast and sensitive microplate assay. The maximum increase in DNA damage was observed after an incubation of 12 h in the presence of 1 mg/L of cadmium chloride; after longer incubation, the number of damaged sites decreased, most likely due to DNA repair. Quantitative analysis of the expression of HSP70 (Mr 73 kDa) revealed that onset of maximal levels of HSP70 depends on the concentration of cadmium chloride in the ambient seawater. Maximal induction (8.9-fold increase compared to control) of HSP70 following exposure to 1 mg/L of cadmium chloride was found after 12 h, while longer incubation periods (3-6 days) were needed to reach maximum levels of HSP70 in the presence of lower concentrations of cadmium chloride (0.1 mg/L and 0.01 mg/L). Northern blot analysis revealed that the level of the 2.0 kb sponge GRP78 homolog mRNA transiently increased under cadmium stress; the maximum increase in the presence of 0.1 mg/L of cadmium chloride was observed at day 3. Our results suggest that sponges are useful indicator organisms to assess the genotoxic risks of cadmium pollution in marine environments.