Upstream regulatory elements in chick heme oxygenase-1 promoter: a study in primary cultures of chick embryo liver cells

Mol Cell Biochem. 2000 Jun;209(1-2):17-27. doi: 10.1023/a:1007025505842.


Previously, chick heme oxygenase-1 (cHO-1) gene was cloned by us and two regions important for induction by sodium arsenite were identified. These two regions were found to contain consensus sequences of an AP-1 (-1580 to -1573) and a MRE/cMyc complex (-52 to -41). In the current study, the roles of these two elements in mediating the sodium arsenite or cobalt chloride dependent induction of cHO-1 were investigated further. DNA binding studies and site-directed mutagenesis studies indicated that both the AP-1 and MRE/cMyc elements are important for the sodium arsenite induction, while cobalt chloride induction involves only the AP-1 element. Electrophoretic mobility shift assays showed that nuclear protein binding to the AP-1 element was increased by both sodium arsenite or cobalt chloride treatment, whereas the binding of proteins to the MRE/cMyc element showed a high basal expression in untreated cells and the binding activity was only slightly increased by sodium arsenite treatment. Site-directed mutagenesis studies showed that, to completely abolish sodium arsenite induction, both the AP-1 and MRE/cMyc elements must be mutated; mutation of either element alone resulted in only a partial effect. In contrast, a single mutation at AP-1 element was sufficient to reduce the cobalt chloride induction almost completely. The MRE/cMyc complex plays a major role in the basal level expression, and shares some similarities to the upstream stimulatory factor element (USF) identified in the promoter regions of mammalian HO-1 genes and other stress regulated genes. Because sodium arsenite is known to cause oxidative stress and because activation of AP-1 proteins has been shown to be a key step in the oxidative stress response pathway, we also explored the possibility that the induction of the cHO-1 gene by sodium arsenite is mediated through oxidative stress pathway(s) by activation of AP-1 proteins. We found that pretreatment with antioxidants (N-acetyl cysteine or quercetin) reduced the induction of the endogenous cHO-1 message or cHO-1 reporter construct activities induced by sodium arsenite or cobalt chloride. These antioxidants also reduced the protein binding activities to the AP-1 element in the electrophoretic mobility shift assays. In summary, induction of the cHO-1 gene by sodium arsenite or cobalt chloride is mediated by activation of the AP-1 element located at -1,573 to -1,580 of the 5'UTR.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Arsenites / pharmacology
  • Base Sequence
  • Binding Sites
  • Cells, Cultured
  • Chick Embryo
  • Chickens
  • Chloramphenicol O-Acetyltransferase / genetics
  • Cobalt / pharmacology
  • Consensus Sequence
  • Enzyme Induction / drug effects
  • Gene Expression Regulation, Enzymologic / drug effects
  • Heme Oxygenase (Decyclizing) / biosynthesis
  • Heme Oxygenase (Decyclizing) / genetics*
  • Heme Oxygenase-1
  • Liver / cytology
  • Liver / enzymology*
  • Mammals
  • Mutagenesis, Site-Directed
  • Promoter Regions, Genetic*
  • Recombinant Fusion Proteins / biosynthesis
  • Regulatory Sequences, Nucleic Acid*
  • Sodium Compounds / pharmacology
  • Transfection
  • beta-Galactosidase / genetics


  • Arsenites
  • Recombinant Fusion Proteins
  • Sodium Compounds
  • Cobalt
  • sodium arsenite
  • Heme Oxygenase (Decyclizing)
  • Heme Oxygenase-1
  • Chloramphenicol O-Acetyltransferase
  • beta-Galactosidase
  • cobaltous chloride