Metallothionein induction in cultured fibroblasts and liver of a marine flatfish,the turbot,Scophthalmus maximus

Intraperitoneal injection of turbot with Cd induced the synthesis of a low molecular weight hepatic Cd-binding protein and a 500bp mRNA, which hybridised to a plaice metallothionein (MT) cRNA probe. The Cd-binding protein displayed cross-reactivity in a competitive ELISA with antiserum raised against rainbow trout MT and had the characteristic amino acid composition, metal stoichiometry and spectral characteristics of a Class I MT. Only one isoform was apparent on ion exchange chromatography. Southern blot analysis of DNA cleaved with four restriction enzymes suggested that only a single MT gene is present in turbot.In an established turbot fibroblast cell line, Cd induced MT mRNA and MT levels in a dose and time-dependent manner. MT was also induced by Cu, Hg and Zn but not Pb exposure. Physiological concentrations of glucocorticoids and sex hormones did not induce MT synthesis, although at high concentrations a positive response to corticosterone, dexamethasone, hydrocortisone or progesterone was observedin vitro indicating the possible presence of a functional steroid regulatory element in the fish MT gene.Abbreviations used AMP adenosine monophosphate - CHSE chinook salmon embryo - DMSO dimethyl sulphoxide - HPLC high performance liquid chromatography - KB kenacid blue - MT metallothionein - Mr molecular weight (kDaltons) - NR neutral red - PEG polyethylene glycol - RTH rainbow trout hepatoma - SDS sodium dodecyl sulphate - SSC 0.15M NaCl, 0.015M sodium citrate - TF turbot fibroblast