Two metallothioneins in the fresh-water fish, crucian carp (Carassius cuvieri): cDNA cloning and assignment of their expression isoforms

Abstract

Two metallothionein cDNAs (MT-A and MT-B) in the fresh-water fish crucian carp (Carassius cuvieri TEMMINCK et SCHLEGEL) were cloned. Sequence analysis of both cDNAs gave the structure of a coding region corresponding to 60 amino acid residues. The homology of their deduced amino acid sequences was completely conserved at the positions of the cysteine residue, but a significant difference existed in the size of their 3'-untranslated regions (130 base pairs for MT-A and 280 base pairs for MT-B). Direct amino acid sequencing of the MT-II isoform purified by HPLC was accomplished for up to 30 residues and its sequence was identical to that deduced from MT-B cDNA. This is the first case in vertebrates that N-terminal methionine in crucian carp MT-II was not blocked. By northern blot analysis, basal and cadmium chloride- or dexamethasone-induced MT-B (MT-II) mRNAs were detected time dependently after treatment. On the other hand, the expression of MT-A mRNA was extremely low. These results indicate that the MT isoform II in crucian carp is coded by the MT-B gene, and that the MT-B-dominant expression of mRNA in crucian carp may be due to the difference in the 3'-untranslated regions of MT mRNAs.