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As it is well-established that protein extraction constitutes a crucial step for two-dimensional electrophoresis (2DE), this work was done as a prerequisite to further the study of alterations in the proteome in gills of the shore crab Carcinus maenas under contrasted environmental conditions. Because of the presence of a chitin layer, shore crab gills have an unusual structure. Consequently, they are considered as a hard tissue and represent a challenge for optimal protein extraction. In this study, we compared three published extraction procedures for subsequent applications to 2DE: the first one uses homogenization process, the second one included an additional TCA-acetone precipitation step, and finally, the third one associated grinding in liquid nitrogen (N 2 ) and TCA-acetone precipitation. Extracted proteins were then resolved using 1DE and 2DE. Although interesting patterns were obtained using 1DE with the three methods, only the one involving grinding in liquid N 2 and TCA-acetone precipitation led to proper resolution after 2DE, showing a good level of reproducibility at technical (85%) and biological (84%) levels. This last method is therefore proposed for analysis of gill proteomes in the shore crab.

Estrogenic potential of environmental samples is frequently assessed using receptor-based functional assays. Using the yeast estrogen screen (YES) developed by Routledge and Sumpter, we assessed the ability of cadmium to activate the estrogen receptor-mediated response. No induced transcriptional activity was observed with a range of CdCl2 concentrations (1 nm-1mM). But, when combining cadmium with the model compound 17beta-estradiol, cadmium was able to significantly potentiate the induced estrogenic response for concentrations ranging from 15 nM to 1 microM. A maximal effect was observed at 0.5 microM with a ten fold reduction of the 17beta-estradiol EC50.