title: The hypoxia-inducible miR-429 regulates hypoxia-inducible factor-1α expression in human endothelial cells through a negative feedback loop.
abstract: Hypoxia-inducible factors (HIFs) 1 and 2 are dimeric α/β transcription factors that regulate cellular responses to low oxygen. HIF-1 is induced first, whereas HIF-2 is associated with chronic hypoxia. To determine how HIF1A mRNA, the inducible subunit of HIF-1, is regulated during hypoxia, we followed HIF1A mRNA levels in primary HUVECs over 24 hours using quantitative PCR. HIF1A and VEGF A (VEGFA) mRNA, a transcriptional target of HIF-1, increased ∼ 2.5- and 8-fold at 2-4 hours, respectively. To determine how the mRNAs were regulated, we identified a microRNA (miRNA), miR-429, that destabilized HIF1A message and decreased VEGFA mRNA by inhibiting HIF1A. Target protector analysis, which interferes with miRNA-mRNA complex formation, confirmed that miR-429 targeted HIF1A message. Desferoxamine treatment, which inhibits the hydroxylases that promote HIF-1α protein degradation, stabilized HIF-1 activity during normoxic conditions and elevated miR-429 levels, demonstrating that HIF-1 promotes miR-429 expression. RNA-sequencing-based transcriptome analysis indicated that inhibition of miRNA-429 in HUVECs up-regulated 209 mRNAs, a number of which regulate angiogenesis. The results demonstrate that HIF-1 is in a negative regulatory loop with miR-429, that miR-429 attenuates HIF-1 activity by decreasing HIF1A message during the early stages of hypoxia before HIF-2 is activated, and this regulatory network helps explain the HIF-1 transition to HIF-2 during chronic hypoxia in endothelial cells.
authors.affiliation: Departments of *Inorganic Chemistry and Biology and Pharmaceutical Botany, Medical University of Gdansk, Gdansk, Poland; and Department of Cell, Developmental and Integrative Biology, University of Alabama, Birmingham, Birmingham, Alabama, USA.
authors.forename: Sylwia
authors.initials: S
authors.lastname: Bartoszewska
authors.affiliation: Departments of *Inorganic Chemistry and Biology and Pharmaceutical Botany, Medical University of Gdansk, Gdansk, Poland; and Department of Cell, Developmental and Integrative Biology, University of Alabama, Birmingham, Birmingham, Alabama, USA.
authors.forename: Kinga
authors.initials: K
authors.lastname: Kochan
authors.affiliation: Departments of *Inorganic Chemistry and Biology and Pharmaceutical Botany, Medical University of Gdansk, Gdansk, Poland; and Department of Cell, Developmental and Integrative Biology, University of Alabama, Birmingham, Birmingham, Alabama, USA.
authors.forename: Arkadiusz
authors.initials: A
authors.lastname: Piotrowski
authors.affiliation: Departments of *Inorganic Chemistry and Biology and Pharmaceutical Botany, Medical University of Gdansk, Gdansk, Poland; and Department of Cell, Developmental and Integrative Biology, University of Alabama, Birmingham, Birmingham, Alabama, USA.
authors.forename: Wojciech
authors.initials: W
authors.lastname: Kamysz
authors.affiliation: Departments of *Inorganic Chemistry and Biology and Pharmaceutical Botany, Medical University of Gdansk, Gdansk, Poland; and Department of Cell, Developmental and Integrative Biology, University of Alabama, Birmingham, Birmingham, Alabama, USA.
authors.forename: Renata J
authors.initials: RJ
authors.lastname: Ochocka
authors.affiliation: Departments of *Inorganic Chemistry and Biology and Pharmaceutical Botany, Medical University of Gdansk, Gdansk, Poland; and Department of Cell, Developmental and Integrative Biology, University of Alabama, Birmingham, Birmingham, Alabama, USA jcollawn@uab.edu rafalbar@gumed.edu.pl.
authors.forename: James F
authors.initials: JF
authors.lastname: Collawn
authors.affiliation: Departments of *Inorganic Chemistry and Biology and Pharmaceutical Botany, Medical University of Gdansk, Gdansk, Poland; and Department of Cell, Developmental and Integrative Biology, University of Alabama, Birmingham, Birmingham, Alabama, USA jcollawn@uab.edu rafalbar@gumed.edu.pl.
authors.forename: Rafal
authors.initials: R
authors.lastname: Bartoszewski
mesh_terms: D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D015687:Cell Hypoxia / Q000235:genetics* / Q000502:physiology*; D025461:Feedback, Physiological; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases / Q000235:genetics; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000235:genetics; D008954:Models, Biological; D018919:Neovascularization, Physiologic; D059748:Proteolysis; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D035683:MicroRNAs; D006899:Mixed Function Oxygenases; D012333:RNA, Messenger; D012097:Repressor Proteins; D042461:Vascular Endothelial Growth Factor A; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human; C546363:MIRN429 microRNA, human; C467484:VEGFA protein, human; C439790:HIF1AN protein, human; C490190:EGLN1 protein, human
keywords: HIF-2; VEGF A; angiogenesis; hypoxamiR; miRNA