Rapid changes in heat-shock cognate 70 levels, heat-shock cognate phosphorylation state, heat-shock transcription factor, and metal transcription factor activity levels in response to heavy metal exposure during sea urchin embryonic development

Abstract

The aim of the present study was to analyze and compare the effects of several metals on the embryos of the sea urchin Paracentrotus lividus, a key species within the Mediterranean Sea ecosystem. Embryos were continuously exposed from fertilization to the following metals: 0.6 mg/l copper, 3 mg/l lead, and 6 mg/l nickel. The embryos were then monitored for metal responses at the gastrula stage, which occurred 24 h after exposure. A biochemical multi-experimental approach was taken and involved the investigation of the levels of HSC70 expression and the involvement of heat shock factor (HSF) and/or metal transcription factor (MTF) in the response. Immunoblotting assays and electrophoretic mobility shift assays (EMSA) were used to detect stress protein levels and to study the interaction between DNA and specific transcription factors, respectively. In the 1 h during exposure to heavy metals, changes in HSC70 levels and HSC70 a phosphorylation state were observed. Rapid changes in HSF and MTF DNA-binding activity also occurred during the early stages of heavy metal exposure. In contrast, few developmental abnormalities were observed at the gastrula stage but more abnormalities were observed 48 h after metal exposure. These data demonstrate that changes in HSC70 levels and phosphorylation state as well as in HSF and MTF binding activities may be used to rapidly detect responses to heavy metal exposure. Detection of biochemical and molecular changes in response to metal exposure before manifestation of morpho-pathological effects are important for the prediction of morbidity, and these markers will be useful for determining the response to exposure as part of a toxicological exposure-response experiment and for determining responses for an impact assessment.

Sea urchin immune cells as sentinels of environmental stress

“…The stress proteins belonging to the Hsp70 family are known to serve as critical indicators of changes in the steady state homeostasis of cells, tissues and organs (Lindquist and Craig, 1988). The highly conserved Hsp70 family includes Hsc70, which is constitutively synthesized and shows moderate modulation upon mild stress, and Hsp70, which is usually not present in cells, but is highly induced by heavy stress (Deane and Woo, 2006;Franzelletti and Fabbri, 2005;Pinsino et al, 2008Pinsino et al, , 2010Pinsino et al, , 2011. Both Hsc70 and Hsp70 have chaperone functions, participating in i) protein synthesis and maturation, ii) folding, assembly, and disassembly of nascent proteins, iii) refolding of mature proteins, and iv) proteolysis and intracellular trafficking (Lindquist and Craig, 1988).…”

“…The Hsc70 levels were evaluated by immunoblotting with an anti-Hsp70 antibody that recognized both Hsc70 and Hsp70 forms of the stress proteins, but also exhibits a strong reactivity with the constitutive form present in both sea urchin embryonic and immune cells (Geraci et al, 2004;Matranga et al, 2006;Pinsino et al, 2008Pinsino et al, , 2010Pinsino et al, , 2011. The increased Hsc70 levels have been explained as an activated thermo-tolerance capability of the sea urchins, in agreement with the Hsp70-mediated thermo-tolerance described in studies on P. lividus sea urchin embryos (Giudice et al, 1999;Roccheri et al, 1995) and embryos or adult organs of other echinoderm species (Dong et al, 2011;Hammond and Hofmann, 2010).…”

Potential toxicity of improperly discarded exhausted photovoltaic cells

“…In both cases the observed effects can likely be related to nickel toxicity. In daphnids it is known that nickel interferes with protein metabolism (Vandenbrouck et al, 2009) while in the sea urchin it induces changes in gene expression (Geraci et al, 2004;Pinsino et al, 2011).…”

The Future of Human Embryo Culture Media – Or Have We Reached the Ceiling?

“…Alternatively, embryos were grown in tissue culture media that were designed to support immortal cell lines in culture and were more complex containing salts supplemented with carbohydrates, amino acids, vitamins, nucleic acids and metal ions and include media such as Ham's F-10, MEM or TCM-199 (Lane 2001;Menezo et al 1984). None of these media were designed to support embryo development and they contained many components that have subsequently been shown to be detrimental to embryo development in vitro such as high levels of glucose, metal ions and hormones (Bowden et al 1993;Brown and Whittingham 1991;Pinsino et al 2010;Quinn 1995;Takahashi and First 1992;Van Winkle and Campione 1982;Vidal and Hidalgo 1993). In fact early attempts to culture the human embryo for prolonged periods in these media resulted in low blastocyst development rates and implantation rates between 5-10%.…”