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FASEB J. 2015 Apr;29(4):1467-79. doi: 10.1096/fj.14-267054. Epub 2014 Dec 30.

The hypoxia-inducible miR-429 regulates hypoxia-inducible factor-1α expression in human endothelial cells through a negative feedback loop.

Author information

1
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.
2
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.

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.

KEYWORDS:

HIF-2; VEGF A; angiogenesis; hypoxamiR; miRNA

PMID:
25550463
PMCID:
PMC4396612
DOI:
10.1096/fj.14-267054
[Indexed for MEDLINE]
Free PMC Article

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