Cloning and characterization of metallothionein cDNAs in the mussel Mytilus edulis L. digestive gland

Metallothioneins are small metal-binding proteins found in all species of a nimals and are transcriptionally-induced by heavy metal ions, oxidative str esses, and inflammation. In the blue sea mussel, Mytilus edulis, several ap parent subtypes of each isoform have been purified and biochemically sequen ced. To determine whether the high number of metallothionein forms present in M. edulis were specific to the digestive gland, and to understand how th ese proteins evolved, we cloned five variants of metallothionein from M. ed ulis. MT10 and MT20 isoform fragments were amplified by PCR, and used as ra diolabelled probes to screen digestive gland cDNA libraries. The MT10 trans cripts were 321-353 nucleotides long and the MT20 transcripts, 513-555 nucl eotides. Previously identified primary structures of MT10 subtypes were con firmed and, in addition, a novel subtype was identified. Expression of MT10 and MT20 isoforms shown by clonal representation and Northern blot analysi s indicated that the MT10 message was more prevalent than the MT20 message. Only the MT20 II transcript could be identified among the MT20 clones. The high degree of untranslated region similarity between each isoform indicat es that these additional forms are recent gene duplication events in the My tilus lineage. Exposure of 0.4 mg l(-1) of cadmium to the mussels resulted in a marked increase in both mRNAs suggesting that the MT20 isoform represe nts a primarily inducible metallothionein not highly expressed under basal conditions. (C) 1999 Elsevier Science Inc. All rights reserved.