Heme oxygenase-1 gene enhancer manifests silencing activity in a chromatin environment prior to oxidative stress.
The expression of heme oxygenase-1 (HO-1) is regulated by E1 and E2 enhancers, both of which contain multiple Maf recognition elements (MAREs). In living cells, MAREs are bound by Bach1/MafK heterodimers, hence maintaining a quiescent state of the HO-1 gene (hmox-1). However, in transient transfection assays, they act as transcriptional enhancers. Therefore MAREs may manifest their function only in a chromatin environment. By using NIH3T3 cell pools stably transfected with EGFP reporter genes driven by the wild-type or mutated E2 enhancer, we demonstrate that the E2 MAREs function as transcriptional silencers depending on the binding of Bach1/MafK heterodimer in vivo only in a chromatin environment. After cadmium treatment, they switched into transcriptional enhancers. Surprisingly, single MARE site did not exhibit such function. Furthermore, by using DNase I hypersensitivity assay, we demonstrate that simple chromatin condensations were not involved in the Bach1-mediated repression. We conclude that, in a chromatin environment, the E2 MAREs function as transcriptional silencers depending on binding of Bach1/MafK heterodimer.
Department of Biomedical Chemistry, Hiroshima University Graduate School of Biomedical Science, Hiroshima, Japan.
Yoshihiro
Y
Dohi
Jawed
J
Alam
Masao
M
Yoshizumi
Jiying
J
Sun
Kazuhiko
K
Igarashi
D016475:3T3 Cells; D000818:Animals; D002843:Chromatin / Q000235:genetics*; D017931:DNA Primers; D004742:Enhancer Elements, Genetic; D005434:Flow Cytometry; D020868:Gene Silencing; D051547:Heme Oxygenase-1 / Q000235:genetics*; D051379:Mice; D018384:Oxidative Stress / Q000502:physiology*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D014158:Transcription, Genetic; D014162:Transfection
D002843:Chromatin; D017931:DNA Primers; D051547:Heme Oxygenase-1