Common carp metallothionein-1 gene: cDNA cloning, gene structure and expression studies

Biochim Biophys Acta. 2004 Jan 20;1676(2):162-71. doi: 10.1016/j.bbaexp.2003.11.009.

Abstract

Metallothionein-1 (MT-1) cDNA clones were isolated from a common carp (Cyprinus carpio) uninduced hepatopancreas cDNA library. Northern blot assay using the common carp (cc) MT-1 cDNA as a probe showed high fold induction of ccMT mRNA levels in the intestine and kidney following exposure to Cd2+ and Zn2+. Using polymerase chain reaction (PCR), primers designed from the cDNA sequences allowed the isolation of ccMT-1 gene fragments including the 5'-flanking region. The 600 bp 5'-flanking region of ccMT-1 gene carries four putative metal regulatory regions, one AP1, two SP1, one c-Jun site, and a TATA box. The 5'-flanking region of the ccMT-1 gene obtained was a functional promoter responding to the administration of various metal ions as well as hydrogen peroxide (H2O2) and lipopolysaccharide (LPS). When tested in primary cultures of cc hepatocytes, Zn2+ had the highest fold (20 times) induction of the 600 bp cloned ccMT-1 gene promoter, followed by Cu2+, Hg2+, Ni2+ and Pb2+ (4-5-fold inductions); H2O2 and LPS had a 6-7-fold induction. In conclusion, the ccMT-1 is a constitutively expressed MT and its gene promoter is inducible by various metal ions and chemical agents.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Blotting, Northern
  • Blotting, Southern
  • Carps / genetics*
  • Cloning, Molecular
  • DNA Primers
  • DNA, Complementary / biosynthesis
  • DNA, Complementary / genetics
  • DNA, Complementary / isolation & purification*
  • Hepatocytes / enzymology
  • Hepatopancreas / enzymology*
  • Metallothionein / biosynthesis
  • Metallothionein / genetics*
  • Metallothionein / isolation & purification
  • Molecular Sequence Data
  • Polymerase Chain Reaction
  • Promoter Regions, Genetic
  • RNA, Messenger / analysis
  • RNA, Messenger / biosynthesis

Substances

  • DNA Primers
  • DNA, Complementary
  • RNA, Messenger
  • Metallothionein

Associated data

  • GENBANK/AF001983
  • GENBANK/AF002161
  • GENBANK/AF002162