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Transcriptional Regulation of the Menkes Copper ATPase ( Atp7a ) Gene by Hypoxia Inducible Factor (HIF) During Iron Deficiency
Author(s) -
Xie Liwei,
Collins James F.
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.229.5
Subject(s) - atp7a , dmt1 , ferroportin , hepcidin , microbiology and biotechnology , chromatin immunoprecipitation , luciferase , promoter , chemistry , biology , gene expression , gene , biochemistry , transporter , transfection , immunology , inflammation
Several intestinal genes are induced by HIF‐2α during iron deficiency, including divalent metal transporter 1 ( Dmt1 ) and duodenal cytochrome B ( DcytB ). We previously observed that Atp7a was strongly upregulated in the duodenum of iron deficient rats. We thus sought to test the hypothesis that Atp7a is regulated by one of the HIFs during iron deprivation. To mimic hypoxia, rat intestinal epithelial (IEC‐6) cells were treated with CoCl 2 or an iron chelator plus copper; both treatments led to induction of Atp7a mRNA expression. Cells were further exposed to low oxygen conditions, causing a ~4‐fold increase in Atp7a expression. To decipher the molecular mechanism of this induction, several Atp7a promoter fragments (ranging in size from 3000 to 50 bp) were PCR amplified and cloned into a firefly luciferase reporter vector. A 224 bp basal promoter fragment with full activity was identified, which contained evolutionarily conserved HIF binding sites. HIF‐1α and HIF‐2α over‐expression led to a significant increase in Atp7a promoter activity (2‐ and 4‐fold, respectively). Overall, this study demonstrates that Atp7a is regulated by HIF during iron deprivation, in parallel to other iron and copper related genes. Future experiments using chromatin immunoprecipitation (ChIP) and gel mobility shift assay are underway to determine precisely which HIF isoform (HIF‐1α or ‐2α) regulates Atp7a expression. Grant Funding Source: 1R01 DK074867